Glyphosate is currently the most widely used broad-spectrum herbicide.Cultivating glyphosate tolerant crops will help improve the effectiveness of chemical control on weed in farmlands,reduce the use of pesticide,and simplify preventive and control measures.To fully detect the glyphosate tolerance(GT)loci in wheat,484 germplasm resources from the Huang-huai wheat region were used to identify glyphosate toxicity.Based on the wheat 15K SNP array data,genome-wide association analysis(GWAS)was used to explore QTL related to glyphosate tolerance in wheat.The main results were as follows:the trend of changes in glyphosate tolerance of wheat varieties cultivated in different eras was slow,and the glyphosate tolerance had not significantly improved;three glyphosate tolerant wheat germplasms(including Henong 130,Jimai 782 and Taishan 23)were selected based on the phenotypic identification results of pesticide damage;seven QTL associated with the level of wheat pesticide damage were detected by GWAS,including 19 significant SNPs,distributed on wheat chromosomes 1A(0.00—30.48 Mb),1B(6.57—30.57 Mb),1D(0.00—22.98 Mb),4A(656.09—680.09 Mb),5A(508.19—532.19 Mb),6A(54.56—85.09 Mb),and 6D(12.02—36.02 Mb);the two QTL qGlyT-1A and qGlyT-6A located on wheat chromosomes 1A and 6A were the main effector sites for glyphosate tolerance in wheat,containing a total of 16 genes that may be related to glyphosate tolerance in wheat.

Plant growth and production are faced with various biological and abiotic stresses,among which salt stress seriously affects the normal growth and development,quality and yield formation of plants.Plants have evolved morphological structure,physiological and biochemical reactions and genetic basis to adapt to salt stress during the long process of evolution.In terms of morphological structure,the leaves of salt-tolerant plants have waxy layer and lower stomatal density than those of salt-sensitive plants,and salt glands,microhairs,salt vesicles,and casparian strip have salt secretion or blocking functions.In terms of physiological activity regulation,on the one hand,salt-tolerant plants have high enzymatic and non-enzymatic antioxidant substances,such as SOD,CAT,phenolic substances,on the other hand,salt-tolerant plants have a high content of osmoregulatory substances,or can synthesize osmoregulatory substances under salt stress,including soluble proteins and sugars of organic substances and inorganic ions.In terms of molecular mechanism,SOS pathway is the most clearly studied ion regulation pathway,which maintains intracellular Na⁺/K⁺ balance through the synergistic action of SOS1,SOS2 and SOS3.In addition,plant hormones and carbon metabolism pathways also play an important role in the process of plant salt tolerance.This paper summarizes the research progress of salt-tolerant plants,and discusses the potential research focus and direction of salt-tolerant plants in terms of morphological structure,physiological basis,genetic molecular basis and transgenic methods in response to salt stress,which will help researchers quickly find the breakthrough point,gradually improve the mechanism system of salt-tolerant plants,and accelerate the efficient utilization of salt-tolerant plants.

The transcription factor BnHY5-2 is associated with plant stress resistance.In order to reveal the response of Brassica napus L.transcription factor BnHY5-2 to salt alkali stress in B.napus,the response of BnHY5-2 to light and salt and alkali was analyzed by transient overexpression,qRT-PCR analysis and subcellular localization.The results revealed that under light conditions,the expression level of the BnHY5-2 gene in B.napus leaves and stems was 29.22 and 3.15 fold higher,respectively,compared to dark conditions.The higher sensitivity to light in leaves suggested that they were the primary site for light signal response.Under light conditions,the expression of BnHY5-2 in leaves and stems was significantly downregulated by 53.1% and 31.0%,respectively,when B.napus was planted in Dalian coastal saline-alkali soil;after applying saline-alkali treatment under dark conditions,the expression of BnHY5-2 was downregulated by 48.2% in the stem,while the difference in expression in the leaves was not significant,indicating organ differences,indicating that the leaves had stricter requirements for light conditions.In B.napus leaves with transient overexpression of BnHY5-2,two out of six genes related to saline-alkali stress(BnNAC32 and BnGS)showed upregulation by 1.25,3.28 fold,respectively,while the other four genes(Bnamy,BnAsp,BnNHX7,BnTPS)were downregulated by 24.8%,25.4%,71.0%,and 82.0%,respectively.Meanwhile,the content of the resistance substance betaine in B.napus increased from 0.256 to 0.573 mg/g,indicating an enhancement by 1.24 fold,suggesting that the overexpression of BnHY5-2 gene could improve the saline-alkali tolerance of B.napus.Subcellular localization results showed that the transcription factor BnHY5-2 was localized in the nucleus and regulates the expression of functional genes.Therefore,BnHY5-2 is not only related to light signaling but also participates in the saline-alkali resistance of Brassica napus L.

To investigate the role of calcium-dependent protein kinase (CDPK) in wheat growth and stress response,the TaCDPK17 gene was cloned from common wheat and its sequence structure,expression pattern,and stress resistance function were preliminarily analyzed.The results showed that the length of the TaCDPK17 gene coding region was 1 701 bp, encoding 566 amino acids and possessing typical structural features of the CDPK family, including one conserved serine/threonine kinase domain and four EF hand shaped domains. Evolutionary tree analysis of TaCDPK17 and CDPK17 from 12 other plants showed that TaCDPK17 had high homology with the CDPK17 sequence of gramineous crops,especially Aegilops tauschii and barley.The promoter region of TaCDPK17 gene contained multiple cis regulatory elements related to hormone signaling pathways,light response.Among them, there are more abscisic acid (ABA) responsive elements (ABRE) and methyl jasmonate responsive elements (CGTCA). The expression analysis based on Real-time Fluorescence Quantitative PCR showed that the expression level of TaCDPK17 increased to varying degrees after induced by 100 μmol/L ABA, 100 μmol/L methyl jasmonate, 20% PEG6000, and 250 mmol/L NaCl. Under stress conditions of 2 μmol/L ABA and 100 mmol/L NaCl, the germination rate of Arabidopsis seeds overexpressing TaCDPK17 was significantly higher than that of the wild type. Meanwhile, overexpression of TaCDPK17 alleviated the inhibitory effects of ABA or osmotic stress treatments on seedling root growth. During stomatal closure, transgenic plants overexpressing TaCDPK17 are more sensitive to ABA and exhibit a stronger stomatal closure trend compared to wild-type plants. These results indicated that TaCDPK17 plays an important role in stress response and hormone signaling in wheat.

Drought stress has a serious effect on the growth and development of maize,which leads to a decrease of maize yield.Purple acid phosphatase is a phospholipase protein involved in many physiological and biochemical functions of plants.In order to further study the role of purple acid phosphatase family genes in the process of stress resistance of maize,this paper explored the response mode of ZmPAP26b gene under drought stress,and Real-time fluorescence Quantitavive analysis was used to analyze the relative gene expression in different maize inbreeding lines under simulated drought conditions;ZmPAP26b(GenBank:NC_050104.1)was cloned from maize,and PAP genes in Zea mays,Arabidopsis thaliana,Oryza sativa L.,Triticum aestivum L.,Sorghum bicolor and Brachypodium distachyon were identified and bioinformatic analysis was performed.Meanwhile,prokaryotic overexpression strains were constructed for functional verification.The results showed that the expression of this gene decreased in drought tolerant materials and increased in drought sensitive materials under drought stress.The CDS length of this gene was 1 431 bp,encoding 476 amino acids.A total of 228 PAP genes were found in six species,divided into 4 subfamilies by phylogenetic analysis.The 19 PAP genes in maize were distributed on 9 chromosomes and had similar conserved domains.Analysis of promoter cis-acting elements showed that they contained elements responding to drought and hormones.Prokaryotic expression experiments showed that the growth of strains containing the recombinant plasmid pET28a-ZmPAP26b was inhibited compared with non-loaded strains under 10% PEG-6000 and 15%PEG-6000 simulated drought stress.In summary,it is speculated that ZmPAP26b is negatively regulated under drought stress.

Serine hydroxymethyltransferase(SHMT),as an important enzyme involved in basic metabolism,plays an important role in plant cell metabolism,photorespiration and defense activities.To understand the bioinformatics function of the SHMT gene family in watermelon,explore its gene expression characteristics under abiotic stress,and provide a basis for the functional development of watermelon SHMT and the breeding of watermelon stress-resistance genes.Bioinformatics methods were used to identify SHMT family,and RT-qPCR was used to analyze the expression patterns of ClSHMTs in different tissues and abiotic stresses.The results showed that 8 ClSHMTs gene family members were identified in the whole genome of watermelon,which were unevenly distributed on 6 chromosomes and named ClSHMT1—ClSHMT8 in turn.There were some differences in the physical and chemical properties of each gene family member,such as the number of amino acids,molecular weight,isoelectric point.The protein contained 471—585 amino acids,with molecular weight of 51.87—65.00 ku and isoelectric point of 6.57—8.52,all of which were hydrophilic proteins.The subcellular localization prediction was mainly distributed on mitochondria.Gene structure and protein conserved motifs analysis showed that the ClSHMTs structure consisted of 4—15 exons and 3—14 introns,and all ClSHMTs contained conserved SHMT domains.Furtherly,phylogenetic analysis with 6 species such as cucumber and wheat showed that 50 SHMTs were divided into 3 sub-families,Group Ⅰ—Ⅲ.Promoter of ClSHMTs contained cis-acting elements related to light response,plant hormone response and stress response.The expression pattern analysis showed that 6 ClSHMTs were expressed in different tissues of watermelon,and the expression levels of ClSHMT1,ClSHMT4,ClSHMT5,ClSHMT8 in leaves were significantly higher than those in other tissues.Under low temperature,drought and salt stress,the expression abundance of ClSHMTs varies,but the expression was mainly up-regulated.In conclusion,this study systematically analyzed the SHMT gene family in watermelon,and will provide a reference for the further study of the biological functions of ClSHMTs.

Temperature stress is one of the main nonbiological stresses that affect the quality and yield of spinach.Investigating the molecular response mechanism of spinach to temperature stress is crucial for spinach stress tolerance breeding.To provide a theoretical basis for the research of the mechanism of spinach resistant to cold stress and heat stress,this study used the cold-tolerant inbred line D3 and the heat-tolerant inbred line M10 of spinach as experimental materials and analyzed their transcriptomes and metabolomes under cold and heat stress to explore the transcriptional and metabolic mechanisms underlying spinach tolerance.Transcriptomic analysis showed that the pathways in which the DEGs were the most enriched in D3 and M10 were essentially the same under cold stress and heat stress.Metabolomics analysis showed that under cold stress,they were coenriched in the pyrimidine metabolism and lysine degradation in KEGG pathways.Under heat stress,these were mainly enriched in the tryptophan metabolism,toluene degradation,biosynthesis of various other secondary metabolites,and glycine,serine and threonine metabolism in KEGG pathways.The joint transcriptomic and metabolomic analysis indicated that through data analysis and gene annotation.SpADH(sov2g036390),SpSHMT(sov1g001130)and SpALDH-1(sov4g007150)were identified as the candidate genes for cold stress tolerance in spinach.SpALDH-1(sov4g007150),SpALDH-2(sov1g043320)and SpNPC(sov1g040610)were identified as candidate genes for heat stress in spinach.Among them,SpALDH-1(sov4g007150),which may be a regulatory gene for spinach stress tolerance,was significantly expressed under both cold and heat stress.

The adult plant resistance gene Lr12 exhibits excellent resistance in production systems.To fine map and develop reliable molecular markers for Lr12,a cross was made between the susceptible variety Thatcher and the resistant near-isogenic line RL6011 containing the Lr12 gene.The F₁ generation resulting from this cross was self-pollinated to generate F₂ individual plants and F_2∶3 families.Field evaluations were conducted using a mixture of five highly virulent leaf rust pathotypes (PHTT, THKS, THTT, PHTS, and PHKS) to inoculate F₂ individual plants and F_2∶3 families for adult plant resistance assessment and genetic analysis of resistance.Subsequently,genotyping was performed using a 16K liquid chip on 10 resistant and 10 susceptible individuals from the F₂ generation to identify SNP markers closely linked to Lr12.This enabled the determination of the chromosomal physical interval containing the resistance gene,the development of SSR molecular markers,and the construction of a genetic linkage map.The results indicate that the segregation ratio of resistance to leaf rust in 3 494 F₂ individuals derived from the RL6011(Lr12)/Thatcher cross was consistent with a 3∶1 ratio ( {\mathrm{\chi }}_{3:1}^2=0.14;P=0.71). In the assessment of 685 F_2∶3 families, the segregation ratio among resistant individuals, resistant heterozygous individuals, and susceptible individuals conformed to a 1∶2∶1 ratio ( {\mathrm{\chi }}_{1:2:1}^2=2.01;P=0.37), suggesting that Lr12 is a dominant gene and the population segregation follows Mendelian single-gene inheritance patterns. Genetic linkage map analysis localized the adult plant leaf rust resistance gene Lr12 between SSR molecular markers YK12817 and YK12928,within a genetic interval of 0.38 cM.This corresponds to a physical interval of 2.09 Mb within the physical range of 579.44 Mb to 581.53 Mb on chromosome 4BL of the Chinese Spring reference genome(IWGSC.Ref.V1.0).These findings provide a solid basis for predicting candidate genes.

The NAC(NAM,ATAF1/2,CUC2)gene is an important regulatory factor in the stress signal transduction network.Cloning the NAC gene in castor,studying its molecular characteristics and expression characteristics,aiming to provide data support for the potential function of the castor NAC gene.The RcNAC100-like gene of Tongbi No.5 was cloned by RT-PCR technology,and its molecular characteristics were analyzed,including bioinformatics,subcellular localization,expression patterns,and transcription activation domain analysis.The results showed that the full length of RcNAC100-like gene cDNA was 1 244 bp,including a 1 086 bp coding sequence(CDS),encoding 361 amino acids.The protein had more irregular coil and α-helix structures,and was a hydrophilic,non-secretory protein.Phylogenetic analysis showed that the RcNAC100-like protein was most closely related to NAC proteins in Manihot esculenta and Hevea brasiliensis,with highly similar motif composition and positioning.The subcellular localization of RcNAC100-like protein was consistent with the predicted results,located in the nucleus.The predicted cis-acting elements in the RcNAC100-like promoter region indicate the presence of multiple environmental response and growth-related elements.Expression pattern analysis showed that the RcNAC100-like gene had tissue-specific expression,with the highest relative expression level in the roots.Additionally,the gene could respond rapidly to adverse environments(drought,salt,cold,and ABA stress)and actively express,indicating that the RcNAC100-like gene might be a key gene in the castor's response to stress.Transcription activation assay results showed that the RcNAC100-like transcription factor has transcriptional activation activity in yeast.In summary,the RcNAC100-like gene may play an important role in the castor's resistance to adversity.

In order to explore the role of RPM1 in sorghum disease resistance,a sorghum SbRPM1 gene was obtained from sorghum smut resistant variety SX44B by homologous cloning method.The bioinformatics analysis results showed that the total length of the cDNA of SbRPM1 gene was 2 802 bp,encoding 933 amino acids,and its protein had a theoretical molecular weight of 106.1 ku and an isoelectric point of 7.11,which was a hydrophilic protein.The SbRPM1 protein had no transmembrane structure,and its subcellular localization was in the cytoplasm.Conservative domain analysis showed that SbRPM1 protein contained RX-CC-like,NB-ARC and LRR domains,and belonged to CNL proteins in the NLRs family.Phylogenetic analysis showed that SbRPM1 protein was most closely related to the RPM1 protein of Miscanthus lutarioriparius.The expression pattern of SbRPM1 gene was detected by Real-time quantitative PCR,and the results showed that the expression of SbRPM1 gene was higher in leaves and inflorescence,followed by roots,and the lowest in stem.The expression of SbRPM1 gene was significantly up-regulated at 24—72 h in disease-resistant varieties after inoculation with Sporisorium reilianum pathogen,suggesting that this gene could be induced by S.reilianum and played an important role in sorghum disease resistance.In this study, the CDS sequence of the SbRPM1 gene was cloned for the first time in sorghum, and the structure, nature and expression of the gene were characterized.

To study the resistance mechanism of TaHis gene to Fusarium head blight(FHB)in wheat,the full-length coding sequence of TaHis was cloned,and the bait vector pGBKT7-TaHis was constructed,which was then used as bait for screening a yeast two-hybrid library of wheat ear induced by FHB.After obtaining the interacting proteins,yeast two-hybridization and bimolecular fluorescence complementation were further used to verify the interaction between these proteins,and RT-qPCR was used to analyze the expression pattern of TaHis interacting protein induced by FHB in resistant and susceptible cultivars respectively.The results showed that the bait vector pGBKT7-TaHis was successfully constructed,and 18 yeast monoclones were obtained on the four deficient selection medium(SD/-Leu/-Trp/-His/-Ade)after yeast two-hybrid library screening.Blast analysis showed that a total of 5 proteins were obtained,and the coding sequence of serine/arginine-rich mRNA splicing factor SR45a-like(TaSR)was identified in 6 colonies.We cloned the full-length coding region of TaSR gene from Bainong 4299 and constructed pGADT7-TaSR vector.The experiment of yeast two-hybrid showed that the yeast cells co-transformed with pGADT7-TaSR and pGBKT7-TaHis grew well and appeared blue on SD/-Leu/-Trp/-His/-Ade/ X-α-Gal/AbA,indicating that TaSR and TaHis directly interacted in yeast cells.The vectors of YC-TaHis and YN-TaSR were constructed,and the bimolecular fluorescence complementary experiments were performed.The results showed that strong fluorescence signals were generated in tobacco cells co-transferred with YC-TaHis and YN-TaSR,which further verified the interaction between TaSR and TaHis.RT-qPCR analysis of TaSR gene expression showed that TaSR expression was up-regulated in resistant cultivar Bainong 4299,while down-regulated in susceptible cultivar Bainong 607 upon FHB infection,indicating a positive correlation between TaSR expression level and FHB resistance in wheat.To sum up,the interaction between wheat TaSR and TaHis was proved,and TaSR expression level was positively correlated with FHB resistance in wheat.

Abstract: Tetraploid wheat is the ancestor specie of common wheat and an important food crop.Aiming to provide new resistance sources for wheat variety breeding,the resistance tetraploid wheat germplasm was explored and their resistance genes were identified.TDI-1 is a cultivated emmer wheat that has been immune to powdery mildew in the field for many years.To determine the resistance genes carried by TDI-1,and provide a theoretical basis for genetic improvement of wheat resistance,a durum wheat TDU-1 that was susceptible to powdery mildew was used to hybridize with TDI-1,and their F₁ plants,F₂ population,and F_2:3 lines were obtained.Genetic analysis of resistance was conducted on parents TDI-1,TDU-1,and their hybrid offspring that were inoculated with powdery mildew isolate E09.Then,bulked segregant analysis method combined with molecular markers was used to map the resistance gene.The results showed that TDI-1 was susceptible to E09 during the seedling stage but immune during the adult stage.F₁ plants derived from the cross of TDI-1 and TDU-1 were immune to E09 during the adult stage.The resistance of adult F₂ individuals was separated,and the ratio of resistant and susceptible plants was 3:1($χ_{3:1}^{2}$=0.11,P=0.74);the ratio of the number of homozygous resistant,separated resistant,and homozygous susceptible F_2:3 lines was 1:2:1($χ_{1:2:1}^{2}$=0.47,P=0.79),indicating that the resistance to powdery mildew in the adult stage of TDI-1 was controlled by one dominant gene,temporarily named PmTDI-1.Subsequently,a set of molecular markers was used to amplify the parents and their F₂ population,and then four markers on chromosome 2A,including Xwmc407,NRM-2AS29,NRM-2AS45 and NRM-2AS84, confirmed to be linked to PmTDI-1. PmTDI-1 was between the flanking markers NRM-2AS45 and NRM-2AS84,with genetics distances of 1.8 cM and 4.6 cM,respectively.Therefore,the adult stage powdery mildew resistance gene PmTDI-1 was preliminarily localized on chromosome 2A.This study identified a novel dominant adult-plant-resistance powdery mildew gene PmTDI-1 from tetraploid wheat TDI-1.

Mitogen-activated protein kinase kinase (MAPKK or MKK) plays an important role in plant growth,development and stress responses.In order to identify MAPKK genes related to rust resistance in foxtail millet and provide candidate genes for the study of rust resistance mechanism and disease-resistant molecular breeding of foxtail millet,the members of MAPKK gene family (SiMKKs) in foxtail millet were identified and analyzed at the whole genome level by bioinformatics methods.Real-time PCR was used to detect the expression level of SiMKKs gene in different tissues,under the stress of rust fungus and exogenous hormone treatment.Excel,MEGA and DnaSP were used to analyze the variation sites and haplotypes of the SiMKKs gene related to rust resistance in 70 re-sequenced foxtail millet varieties,and the excellent rust-resistant haplotypes were identified based on phenotype analysis.The results showed that a total of 10 SiMKKs were identified in foxtail millet,which were distributed on 5 chromosomes.The number of exons ranged from 1 to 11,and the encoded protein contained 331-523 amino acids.The SiMKKs were divided into 4 groups.Groups A and B contained S/T-X₅-S/T motif,while SiMKKs in groups C and D did not have this motif.Conserved Motif 1-Motif 6 existed in all SiMKK proteins.The promoter region of each SiMKK gene contained 1 to 3 biotic stress-related cis-acting elements,such as defense and stress response,methyl jasmonate(MeJA) response,salicylic acid(SA) response and elicitor activation.Except SiMKK10-1 and SiMKK10-3,the other 8 SiMKK genes were expressed with different degrees in different tissues,and under rust infection,SA and MeJA treatments.The highest expression of SiMKK4,SiMKK5 and SiMKK10-2 were in roots at booting stage,and the highest expression of SiMKK6-1 and SiMKK6-2 were in stems at booting stage.The expression of SiMKK4 was up-regulated in the resistant response and down-regulated in the susceptible response within 24 h after inoculation,and its expression was related to disease resistance.The expression of SiMKK4 was up-regulated within 16 h and then down-regulatedafter SA and MeJA treatments,and showed continuous changes during SA treatment.In addition,the expression patterns of the remaining 7 SiMKK genes in SA and MeJA treatments were also consistent.The coding region of SiMKK4 gene contained 7 haplotypes and Hap_1 was the dominant haplotype,and no key variation sites related to disease resistance were found.In summary,the expression of SiMKK4 is identified to be associated with resistance to rust disease in foxtail millet,and SiMKK4 may participate in the early disease resistance response of foxtail millet through SA and MeJA signaling pathways.

To investigate the effect of exogenous melatonin (MT) on salt tolerance of soybean seedlings,and to screen the appropriate application concentration under different salt stress.The soybean variety Tianyou-2986 was used as the test material,and 3 salt concentrations (low salt S₃:3 g/L,medium salt S₅:5 g/L,high salt S₇:7 g/L ) and 6 MT concentrations were set(M₀:0 μmol/L,M₁:25 μmol/L,M₂:50 μmol/L,M₃:75 μmol/L,M₄:100 μmol/L,M₅:150 μmol/L),the morphological parameters,biomass,antioxidant enzyme activity and osmoregulatory substance content of soybean seedlings were analyzed.With the increase of salt stress,the root morphological parameters,biomass,root-shoot ratio,antioxidant enzyme activity and osmoregulatory substance content of soybean seedlings decreased,while the malondialdehyde content increased.Under low salt (S₃) and medium salt (S₅) stress,The number of total length,lateral roots,SOD and POD of 50 μmol/L MT were increased by 52.30%,19.98%,74.10%,40.03% (low salt) and 68.52%,19.24%,81.72% and 37.42% (medium salt),respectively.Under high stress (S₇),75 μmol/L MT increased by 71.17%,19.11%,80.79% and 27.01%,respectively.Under salt stress,exogenous 25—100 μmol/L MT promoted soybean seedling growth and improved salt tolerance to different degrees.The overall evaluation of the affiliation function showed that 50 μmol/L MT was the most effective in alleviating salt damage under low and medium salt stress,and the suitable concentration of MT was 75 μmol/L under high salt stress,and the main reason for the alleviation of salt damage in soybean seedlings was that MT increased the activities of antioxidant enzymes and osmoregulatory substance content,and reduced the content of malondialdehyde,which could alleviate oxidative and osmotic stresses of soybeans under salt stress.

To improve the applicability of biochar in saline-alkali agroecosystem,the effect and microbial mechanism of modified biochar were studied.In a 2-year field experiment,common biochar(4.5 t/ha),nitrogen-rich modified biochar(7.5 t/ha)and phosphorus-rich modified biochar(15.0 t/ha)were added to investigate their impact on crop grain yield,soil physicochemical properties and soil microbial diversity.It had been observed that the addition of biochar enhanced the quality of saline-alkali soil,with nitrogen-rich modified biochar and phosphorus-rich modified biochar demonstrating more notable effects.Biochar could boost crop yield,improve soil structure and reduce soil bulk density in saline-alkali land.The effects of the three biochar types were not consistent.Among them,the application of 15.0 t/ha phosphorus-rich modified biochar showed favorable responses,with grain yield of(8.92±0.12)t/ha,representing a 110% increase compared to the control group.Biochar affected soil microbial diversity.Common biochar increased soil microbial diversity,whereas phosphorus-rich modified biochar decreased it.With the continuous addition of biochar,soil physical and chemical properties could affect the relationship between soil microorganisms and plant structure,weakening their relationships.In this study,the application of 15.0 t/ha phosphorus-rich modified biochar was recommended to improve saline-alkali agroecosystem.

Salt-alkali stress has become one of the important factors restricting agricultural production in my country.Exploring the molecular mechanism of salt tolerance of crops has important theoretical and practical value for crop breeding.The purpose of this study is to clone the ZmMPI gene in corn and transform corn plants.First,qRT-PCR was used to analyze the ZmMPI expression changes in plants treated with saline-alkali solutions.Then DNAMAN software was used to perform multiple comparison analysis of the ZmMPI protein sequence.MEGA 7.0 software was used to construct a phylogenetic tree,and a series of software were used to analyze the ZmMPI protein sequence.ZmMPI performed bioinformatics analysis.Finally,molecular cloning technology was used to successfully clone the coding sequence of the ZmMPI gene,construct a plant overexpression vector,and use Agrobacterium-mediated genetic transformation method to transform the corn inbred line B104.The overexpression transgenic plants were transformed at the genome level,transcription level and protein level.Identify and analyze changes in expression levels.The results showed that the expression level of the ZmMPI gene showed an overall trend of first increasing and then decreasing after being subjected to salt-alkali stress;the ZmMPI protein sequence comparison result showed a similarity rate of 64.15%,and the phylogenetic tree showed that ZmMPI had the highest homology with Zea mays subsp.parviglumis ABA34115.1.The protein contained a protein domain Potato_inhibit,which had an α-helix,a random coil and a β-turn.It was relatively hydrophobic and had 10 predicted Potential phosphorylation sites;the identification results of the 49 transformation events obtained showed that the ZmMPI gene in 13 over-expressed transgenic lines could be expressed normally at the genome level,and the ZmMPI gene in 10 over-expressed transgenic lines could be transcribed and translated normally.Finally,10 overexpression transgenic lines capable of normal transcription and translation were obtained,laying the foundation for further exploring the molecular mechanism of ZmMPI gene in response to salt-alkali stress.

To investigate the regulatory effect of DEAD-box helicase 21(DDX21)on the replication of Transmissible gastroenteritis virus(TGEV).Firstly,Western Blot was utilized to analyze the effect of TGEV infection on DDX21 expression.Furthermore,we constructed eukaryotic expression plasmids of porcine DDX21 and established knockdown stable cell lines.RT-qPCR,Western Blot,Indirect immunofluorescence(IFA)and TCID₅₀ assays were used to investigate the regulatory effect of DDX21 on TGEV replication in vitro.Western Blot analysis showed that the protein levels of DDX21 were significantly up-regulated in PK-15 cells at the early stage of TGEV infection.RT-qPCR,Western Blot,IFA and TCID₅₀ experiments showed that over-expression of DDX21 significantly increased the mRNA level and protein of TGEV N in a dose-dependent manner.And the amino acids 601—784 aa of DDX21 were critical for promoting TGEV replication.Otherwise,the titer of TGEV was significantly down-regulated in DDX21 knockdown cell lines,whereas the titer of TGEV in DDX21 knockdown cell lines was reversed under the rescue experiment.This study revealed for the first time that DDX21 promotes the proliferation of TGEV and identified the key domain of DDX21 in regulating TGEV replication,which provided a theoretical a basis for future research on the function of DDX21 protein and the pathogenesis of TGEV.

Auxin response factor(ARF)is a class of transcription factors with B3 domain,which is a direct molecule regulating auxin response and controlling gene expression.A gene,ZmARF10,which encoded ARF protein and actively participated in drought-rehydration stress response,was previously screened in maize by analyzed transcriptome data.To further research the molecular mechanism of ZmARF10 regulating drought resistance of maize,and also provide a new idea for molecular breeding of drought resistance,the gene was firstly analyzed by bioinformatics software.Secondly,Quantitative Real-time polymerase chain reaction(qRT-PCR)was used to detect the expression pattern of ZmARF10 in different tissues,under high temperature,drought,high salt,ABA and restoring treatments,and in different inbred lines.Finally,the function of ZmARF10 was analyzed using CRISPR/Cas9 technology.The results showed that ZmARF10 was located on chromosome 3 of maize,with a total length of 2 127 bp,and encoded 708 amino acids with a typical B3 domain.The upstream 2 kb region of ATG of this gene contained response elements related to methyl jasmonate,auxin,abscisic acid and low temperature.The phylogenetic tree showed that the protein encoded by the ZmARF10 gene was closely related to sorghum.qRT-PCR results showed that ZmARF10 was a constitutive expression gene,and the expression level of ZmARF10 was the highest in mature corn roots of maize.Under high temperature,drought,high salt and ABA treatments,the expression of this gene was significantly up-regulated,and the up-regulation ratio was up to 8.2 times after drought stress.After drought stress,the expression level of ZmARF10 gene was significantly higher in the drought-resistant inbred line Zheng 36 than that in the drought-sensitive inbred line B73.Investigation of Arabidopsis wild type and ARF10-deficient mutants showed that,compared with wild type,the mutant plants showed leaf wilting and even dry death,roots curled,root branch number decreased,and lateral root growth and development were hindered under drought conditions.Determination of physiological and biochemical indexes showed that the relative water content,chlorophyll content and net photosynthetic rate of the deficient mutants were significantly lower than those of the wild-type plants after drought stress,indicating that the drought resistance of Arabidopsis decreased after ARF10 gene was knocked out.

To clarify the effects of drought stress and cultivars on the net photosynthetic rate(Pn),the activities of photosynthetic enzymes in flag leaf in the afternoon(FLA)during grain filling stage and grain yield of winter wheat,a pond culture experiment with four water levels and two winter wheat cultivars was conducted under the condition of rainproof pond cultivation in 2019—2021.The four water treatments included severe drought(W1),moderate drought(W2),mild drought(W3),and suitable water supply(W4).The Pn,and the activities of Rubisco,RCA,PEPC,ATPase and PPDK in flag leaf from 14:00 to 16:00 during the early(EGFS)and medium(MGFS)grain filling stage,and the grain yield at maturity of strong drought resistance cultivar Jinmai 47(JM47)and weak drought resistance cultivar Yanzhan 4110(YZ4110)were investigated.Drought stress decreased the FLA Pn and activities of most photosynthetic enzymes,and the grain yield of wheat.The decrease of these indexes increased with drought stress degree,but the effects were different among varieties and years.Compared with W4,the FLA Pn of JM47 under W1,W2 and W3 decreased by 33.6%—40.6%,12.0%—30.5% and 5.0%—13.5%,as well as YZ4110 decreased by 44.0%—52.0%,22.5%—38.1% and 11.5%—20.5%,respectively.Compared with W4,the FLA Rubisco activity decreased during EGFS but increased during MGFS for JM47,while it decreased by 13.3%—25.6%,7.1%—14.0% and 11.2%—11.6% for YZ4110,respectively,under W1,W2 and W3.Compared with W4,the FLA RCA activity significantly decreased under most drought treatments during EGFS,while increased under W2 and W3 for JM47 and decreased under W1 and W2 for YZ4110 during MGFS.Compared with W4,the FLA ATPase activity of JM47decreased under W1 but increased under W3,while that of YZ4110 decreased under W1,W2 and W3 by 19.3%—48.7%,7.2%—24.2% and 0.1%—8.9%,respectively.The FLA PEPC activity under different treatments varied with growing seasons and varieties.Compared with W4,the FLA PPDK activities of JM47 and YZ4110 under W1 were decreased by 12.4%—18.8% and 16.7%—18.2%.Compared with YZ4110,in most conditions,the FLA Pn and photosynthetic enzyme activities of JM47 had no significant difference under suitable water supply(W4),but increased under drought treatments(W1,W2 and W3).The results of correlation analysis showed that yield,FLA Pn were significantly positively correlated to FLA ATPase activity during EGFS and MGFS,as well as FLA PEPC activity during EGFS.Thus,increasing the FLA ATPase and PEPC activities during the grain filling period is conducive to the increase of FLA Pn and grain yield of wheat.

In order to investigate the physiological regulation mechanism of exogenous H₂O₂ on the cotton seedlings under NaCl stress,the cotton variety Xinluzao 48 was used as the test material in an outdoor potting method.Two-factor random combinations of salt stress (NaCl,concentration gradients of 0,100,200 mmol/L) and H₂O₂ (concentration gradients of 0,0.005,0.010,0.020,0.050 mmol/L) were set,to study the change rule of fresh weight,dry weight,chlorophyll content,chlorophyll fluorescence parameters,photosynthetic gas parameters,antioxidant enzyme activities and osmotic regulation system of cotton seedlings.The results showed that exogenous H₂O₂ effectively alleviated the inhibitory effect of salt stress on the growth of cotton seedlings,increased chlorophyll content,photosynthetic gas parameters and chlorophyll fluorescence parameters of cotton seedlings,maintained the normal operation of photosynthesis of cotton seedlings and ensured the accumulation of dry matter.Meanwhile,exogenous H₂O₂ could increase the activity of antioxidant enzymes (POD,APX,CAT),accelerated the removal of ROS from cotton seedlings.Exogenous H₂O₂ reduced the electrolyte leakage rate,MDA content,the content of osmoregulation substances such as pro,free amino acid and SS,and improved the salt resistance of cotton seedlings.Among all treatment,0.020 mmol/L exogenous H₂O₂ had the best effect in alleviating the salt stress suffered by cotton seedlings.In summary,exogenous H₂O₂ improves the adaptation of cotton seedlings to salt stress by improving photosynthetic performance,keeping stable photosynthesis in cotton seedlings,and maintaining the dynamic balance between ROS production and elimination in cotton seedlings.

Investigating the predominant genotypes of Bovine viral diarrhea virus(BVDV)infecting cattle in Tongliao,Inner Mongolia,to provide reference for the BVDV epidemiology and prevention and control.In the preliminary phase of the experiment,fecal samples from diarrheic calves were collected at a cattle farm in Tongliao,Inner Mongolia.These samples were tested using PCR to detect BVDV positivity.Positive fecal samples were then inoculated into madin-darby bovine kidney cells(MDBK)for isolation.The isolated strains were identified using RT-PCR and indirect immunofluorescence staining.Subsequently,the full-length genome of the isolates was sequenced,followed by genetic evolution analysis and genotype determination based on sequences of the 5'UTR,N^pro,and E2 genes.The results indicated that this experiment successfully isolated a strain of BVDV,designated as NM-21.Inoculation of NM-21 into MDBK did not induce cytopathic effects,indicating it was a non-cytopathic strain(NCP).Both RT-PCR and indirect immunofluorescence staining confirmed its positivity,with a virus titer of 10^-3 TCID₅₀/mL.Based on the full-length genomic sequence,and homology and genetic evolution analysis of the 5'UTR,N^pro,and E2 gene sequences,the isolate NM-21 showed the highest nucleotide homology with the BVDV-1c subtype strain NM2103(GenBank accession number ON337882.1)from Inner Mongolia,China.

In order to investigate the expression pattern and biological function of lncRNA TCONS_00143126 in E.coli type mastitis of cows in depth.This study used cDNA from bovine mammary epithelial cells as a template,and confirmed the presence of lncRNA TCONS_00143126 using PCR cloning and sequencing techniques.Subcellular localization analysis of lncRNA was performed,and potential target miRNAs and genes were predicted.The potential mechanism of its action in bovine mastitis was explored through KEGG pathway enrichment analysis.In addition,LPS was used to induce bMECs to construct an in vitro model of bovine mastitis,and the expression of lncRNA TCONS_00143126 in LPS-induced bMECs at 6,12 and 24 h was detected by RT-qPCR.The results showed that lncRNA TCONS_00143126 was real,and its expression was significantly up-regulated in LPS-induced bMECs,and it was mainly distributed in the nucleus.The results of target gene prediction and KEGG enrichment analysis showed that lncRNA TCONS_00143126 might regulate inflammatory signaling pathways such as JAK-STAT,mTOR and MAPK by targeting miRNAs(bta-miR-133a,bta-miR-193a-5p and bta-miR-375,etc.)and target genes(IFNE,SLC2A10,MEX3B),and then play a role in the inflammation of bovine mammary epithelial cells.

To study the structure and function of phosphotyrosine interaction domain 1 (PID1) gene in yak,and to explore its expression in various tissues.The CDS region of Sangsang yak PID1 gene was cloned using Sangsang yak adipose tissue cDNA as template,and the sequence was analyzed bioinformatically.Meanwhile,the relative expression level of PID1 gene was detected in seven tissues of yak namely heart,liver,spleen,lung,kidney,longissimus dorsi muscle and adipose tissue by Real-time Fluorescence Quantitative PCR(RT-qPCR).The results showed that the PID1 gene in yaks had a coding region length of 654 bp,which encoded 217 amino acids.Homology comparison showed that yak and wild yak were closely related,and the similarity reached 100%.The molecular weight of yak PID1 protein was about 24.84 ku and the theoretical isoelectric point was 6.30.According to the calculation results of instability coefficient,the instability of the protein was high (47.96),and it belonged to an unstable protein.The protein had one N-glycosylation site and 23 phosphorylation sites,with no signal peptide or transmembrane structure.The results of RT-qPCR showed that the expression of PID1 gene could be detected in all tissues,with the highest expression in the lung.The yak PID1 gene was cloned and its protein structure was analyzed.The expression of PID1 gene in yak tissue was also studied.Further study on the role of PID1 gene in yak fat deposition provided preliminary data.

To clarify the effects of sulfur application at different stages on the yield and photosynthetic characteristics of strong-gluten wheat,and to determine the appropriate time for sulfur fertilizer application,from 2019 to 2021 during two wheat growth seasons,using strong-gluten wheat Gaoyou 2018 as the experimental material,three sulfur application periods were set:before sowing (S60-b),topdressing at jointing stage (S60-j),and topdressing at anthesis stage (S60-a),with a sulfur application rate of 60 kg/ha,and no sulfur application (CK) as the control.The effects of leaf area index (LAI),relative chlorophyll content of flag leaf (SPAD),soluble protein content,soluble sugar content,yield and components of strong-gluten wheat under different sulfur application treatments were studied systematically.The results showed that compared to CK,S60-b,S60-j and S60-a could all significantly increase the thousand-grain weight (TGW),maximum grain filling rate and yield of Gaoyou 2018.In the two growing seasons,the average increase of TGW was 6.23%,4.27% and 7.04%,respectively;the increase of TGW was the highest at 35 days post anthesis,which was 5.51%,3.17%and 6.12%,respectively.The average increase of maximum grain filling rate was 2.84%,1.76% and 3.49% and the average increase in yield was 9.20%,2.73% and 5.71%,respectively.However,sulfur application had no significant effect on the number of ears and grains per unit area of Gaoyou 2018.Sulfur application at different stages had significant effects on the photosynthetic characteristics of strong-gluten wheat.The LAI,SPAD value of flag leaves in the middle and late stages of growth,the soluble protein content of flag leaves at 22 and 29 days post anthesis and soluble sugar content in flag leaves at 35 days post anthesis were significantly increased by S60-b and S60-a treatments.The five above indexes increased by more than 26.16%,7.38%,16.90%,55.29% and 81.11%,respectively.According to the results of two years,before sowing and topdressing at the anthesis stage with sulfur fertilizer could significantly increase the LAI of flag leaves in the middle and late growth stage,maintain high SPAD value,delay flag leaf senescence,increase the contents of soluble protein and soluble sugar,TGW and filling rate at the end of grain filling,showing higher yield.In summary,sulfur application have a positive regulatory effect on the yield of strong-gluten wheat,and before sowing and topdressing at the anthesis stage have a good regulatory effect.

Bioinformatics and expression patterns of Dicer-like(DCL),Argonaute(AGO)and RNA-dependent RNA polymerase(RDR)gene families in the whole genome of Solanum habrochaites were analyzed,so as to provide references for further study on the functions of DCL,AGO and RDR gene families in the response of S.habrochaites to abiotic and viral infection.Using Arabidopsis thaliana DCL,AGO and RDR genes as reference sequences,the genome of S.habrochaites LA1777 was searched by local perl language and software such as Pfam and SMART,and the members of ShDCL,ShAGO and ShRDR gene families were determined.Bioinformatics analysis of DCL,AGO and RDR family genes in S.habrochaites was carried out by means of ExPASy,GSDS 2.0,MEGA,Tbtools and SWISS-MODEL.According to abiotic stress treatment,Tomato chlorosis virus(ToCV)treatment and Real-time Fluorescence Quantitative PCR technology,the expression patterns of these genes were analyzed.Seven ShDCL,15 ShAGO and 6 ShRDR genes were identified from S.habrochaites,which were distributed on chromosome 5,7 and 6 respectively.The encoded proteins were similar in structure to DCL,AGO and RDR in other plants,and all of them contained conserved domains unique to this family.Phylogenetic analysis showed that these genes were divided into 4 subgroups,and there were high structural and functional similarities between S.habrochaites and S.lycopersicum.ShDCL2a,ShDCL2c,ShDCL3,ShDCL4,ShAGO1b,ShAGO3,ShAGO4b,ShAGO5,ShAGO7,ShAGO10a,ShAGO10b,ShRDR1,ShRDR2,ShRDR3a,ShRDR6a and ShRDR6b were significantly up-regulated after various abiotic stresses and ToCV infection.It is speculated that these genes play important roles in abiotic stress and virus infection.

Luchuan pig myoblast determining gene 1(MyoD1)was cloned,bioinformatics analysis was performed,and its expression was analyzed in different tissues of Luchuan pigs,to explore the role of MyoD1 gene in the growth and development of skeletal muscle.Published MyoD1 gene of wild boar sequence on NCBI as the template,designed specific primer,cloned the CDS region of MyoD1,and compared the similarity with wild boar,cattle,sheep,human,mouse and rat gene sequence,constructed system tree for bioinformatics analysis by online software.Then,the relative expression of MyoD1 gene in different tissues of Luchuan pigs by RT-PCR.The CDS of MyoD1 gene in Luchuan pig was 960 bp,encoding 319 amino acids,five base mutations.The similarities of MyoD1 genes between Luchuan pig and wild boar,cattle,sheep,human,mouse and rats were 99.2%,93.0%,92.3%,90.0%,84.3% and 84.0%,respectively.Total number of MyoD1 gene atoms in Luchuan pigs was 4 680,molecular mass of the protein was 33.99 ku,molecular formula was C₁₄₅₇H₂₂₉₆N₄₄₂O₄₇₁S₁₄;the instability coefficient was 63.87,indicating a lack of stability;isoelectric point was 5.63,which belongs to the acidic protein;no transmembrane structure existed,found 35 phosphorylation sites and 1 glycosylation site;protein secondary structure was dominated by an irregular coil,which accounting for 60.69%.The MyoD1 gene in Luchuan pigs had the highest expression in the longest dorsal muscle,which was significantly higher than in other tissues,while kidney had the lowest expression.MyoD1 gene was expressed in all tissues of Luchuan pigs and mainly in the longest dorsal muscle,we speculated that MyoD1 gene might play a crucial role in the muscle growth and development of Luchuan pigs.

In order to explore the expression of uncoupling protein 3(UCP3)in different tissues of Min pig and its effect on the expression of mitochondrial-related genes and ATP synthase-related genes in Min pig preadipocytes,the back adipose tissue of 1-month-old Min pig was collected as the research material.The preadipocytes were isolated by enzymatic digestion and cultured in vitro.At the same time,axillary fat,chest fat,back fat,inguinal fat,perirenal fat and intramuscular fat were collected to construct tissue expression profiles.Real-time Quantitative PCR was used to detect the expression of MCU family genes(MCU,MICU1,MICU2),ATP synthase(ATP5B,ATP5E)and 1,4,5-trisphosphate inositol receptor type 1(ITPR1)genes and the expression of UCP3 gene in different adipose tissues by in vitro transfection of UCP3 gene overexpression vector or interference fragment.The results showed that the enzymatic digestion method could quickly obtain sufficient preadipocytes in the back adipose tissue.The cell edge had good refractive index,clear boundary,and similar morphology to fibroblasts.UCP3 gene was expressed in different adipose tissues,with the highest expression in dorsal adipose tissue and the lowest expression in abdominal fat.After overexpression of UCP3 gene,the expression levels of MCU family genes and ITPR1 genes were significantly increased,and the expression level of ATP5B gene was significantly decreased.After UCP3 gene interference,the expression levels of MCU family genes and ITPR1 genes were significantly decreased,and the expression levels of ATP synthase genes were significantly increased.The results showed that UCP3 gene was differentially expressed in different adipose tissues,which could promote the expression of MCU family genes and ITPR1 genes and inhibit the expression of ATP5B gene.

In order to investigate the sequence characteristics of bovine Bta-miR-494 and its tissue expression,bioinformatics methods were used to analyze the characteristics of CpG islands,promoters and transcription factors of its upstream sequence and their conservatism among different species;then predicted and functionally enriched the target genes of bovine Bta-miR-494;and finally detected its spatial and temporal expression characteristics among different tissues and the same tissue at different periods of time of the bovine Bta-miR-494 by qRT-PCR method.The results showed that Bta-miR-494 was highly conserved among different species and had the closest affinity to goat,with no CpG island upstream and two transcriptional start sites;the target gene enrichment analysis showed that the target genes of Bta-miR-494 were significantly enriched in the signaling pathways related to muscle growth and metabolism,such as PI3K-Akt,p53 and MAPK; the predictive target gene enrichment analysis showed that the target genes of Bta-miR-494 were significantly enriched in the signaling pathways related to muscle growth and metabolism.qRT-PCR results showed that the expression of Bta-miR-494 was highest in the liver and lowest in the testis of adult cows;and the expression of Bta-miR-494 was significantly higher in the biceps femoris of adult cows than that of newborn calves,whereas the expression of Bta-miR-494 in the cardiac muscle and longitudinal muscle of dorsal cows was significantly lower than that of newborn calves,while the expression of Bta-miR-494 in the cardiac muscle and the longissimus dorsi muscle was significantly lower than that of newborn calves.This inter-tissue differential expression suggests that Bta-miR-494 may dominate different myogenic expression patterns during the development of muscle tissues at different growth stages in cattle.

Clarifying the effect of spring limited irrigation on the root development and grain yield of winter wheat in Haihe Plain is of great significance to reduce irrigation and improve water use efficiency.This study used Shimai 22 as the test material,irrigation treatments were traditional irrigation twice at jointing and anthesis stage(W2),no irrigation(W0),and single irrigation(W1)with four irrigation-time treatments(3L,4L,5L,and 6L)based on the number of leaves unfolded in spring.The results showed that compared with W2,W0 and W1 yield decreased by 54.6% and 24.4% respectively,the irrigation yield was highest at 4L in W1,and the effect of yield composition reduction was not significant.Limited irrigation reduced the total root weight density and root length density of winter wheat.During the anthesis period, the total root weight density of W1 decreased significantly by 17.2%, while the total root weight density and root length density of W0 decreased significantly by 47.5% and 35.1%, respectively. And under W1 condition, 4L has the highest total root weight density and root length density. The vertical distribution of roots showed that reducing the frequency of irrigation increased the distribution of roots in the soil layer below 40 cm,however,with the postponement of irrigation time,the root distribution of W1 deep soil decreased and root vigor increased.Among them, during the anthesis period, 4L was significantly higher than 6L by 28.8%, 14.2%, and 36.5% in the 120—160 cm, 160—200 cm, and 200—240 cm soil layers, respectively. Correlation and path analysis showed that total root weight density and root length density at joint—anthesis period had a positive effect on yield.The direct contribution of total root length density in 3L and 4L irrigation was the largest.Generally speaking,the root mass of 4L treatment was higher at jointing-anthesis period,the deep root distribution and root activity of 40—240 cm were increased,resulting in higher spike number and kernel number,which was beneficial to alleviate the decrease of winter wheat yield at limited irrigation,it can be used as an effective way of limited irrigation for winter wheat in Haihe Plain.

In order to study the structure and function of DJ-1 gene in yaks,this experiment used Meiren yak adipose tissue cDNA as a template,cloned the coding sequence (CDS) of yak DJ-1 gene using RT-PCR,and performed bioinformatics analysis,including domain prediction and protein physicochemical properties prediction of the gene sequence.Then,tissue expression analysis was performed using RT-qPCR.The results showed that the CDS region of the DJ-1 gene in Meiren yak had a total length of 570 bp and encoded 189 amino acids;the prediction of the DJ-1 domain in yaks showed that there was one PfpI domain containing the Pfam in positions 29—168 of the DJ-1 amino acid sequence.By predicting structure and function of the DJ-1 protein,the results showed no transmembrane structure and no signal peptide region;molecular weight was 20.035 31 ku,total number of atoms was 2 870,theoretical isoelectric point was 6.84,instability coefficient was 28.37,and expressed as a stable protein;the fat coefficient was 101.11,with a total average hydrophilicity coefficient of -0.004.It was a hydrophilic protein with a total of 11 phosphorylation sites.Meanwhile,the results of subcellular localization prediction indicated that the DJ-1 protein of Meiren yak was mainly distributed in the cytoplasm and mitochondria.The phylogenetic tree showed that Meiren yak had the closest genetic relationship with wild yaks and European cattle,and the farthest genetic relationship with Arctic foxes and wide snouted manatees;the RT-qPCR results showed that it was expressed in the heart,liver,spleen,lungs,muscle,fat,and testicular tissues of adult Meiren yaks,with the highest expression level in the heart and muscle tissues,and the lowest expression level in the liver and testicular tissues.

This study aims to obtain the MFSD4A gene sequence in Maiwa yak and elucidate its expression patterns in different tissues,analyze the mRNA expression levels of the interacting proteins of MFSD4A,and determine the subcellular localization of the MFSD4A protein,and provide a theoretical basis for further research on regulating MFSD4A in the growth and development of yak testis.RT-PCR was used to obtain the MFSD4A gene CDS sequence in Maiwa yak and perform bioinformatics analysis.RT-qPCR was used to detect the expression differences of the MFSD4A gene in 10 tissues,including heart,liver,spleen,lung,kidney,gluteal muscle,back muscle,fat,testis,and lymph,and the correlation between the interacting proteins and MFSD4A.pEGFP-MFSD4A fusion plasmid was transfected into Madin-Darby bovine kidney cells(MDBK)to investigate the subcellular localization of the MFSD4A protein.The results indicated that the entire length of the MFSD4A gene CDS region in the macaque yak was 1 530 bp,encoding 509 amino acids with a theoretical isoelectric point(PI)of 8.66.It had 12 transmembrane domains and belonged to the alkaline transmembrane protein.RT-qPCR results showed differential expression of MFSD4A mRNA in various tissues of the macaque yak,with the highest expression in the testis.It was also highly positively correlated with the expression levels of MFSD9,CBY1,INHBA,UNC93A,SVOP,and ID1 in the testis,suggesting that the MFSD4A gene might be related to the regulation of growth and development of yak testis.Fluorescence localization of the pEGFP-MFSD4A fusion vector transfected into Madin-Darby bovine kidney cells(MDBK)showed that the protein was mainly located in the nucleus.

The domain of unknown function 668(DUF668)family is a family of plants whose function is unknown.To unveil the function and characteristics of maize DUF668(ZmDUF668)gene family,we identified ZmDUF668 gene family by bioinformatics method.The results showed that there were 19 ZmDUF668 genes in maize,distributing on 8 chromosomes,named ZmDUF668-1—ZmDUF668-19;the most of proteins encoded by ZmDUF668 were alkaline,and most of the family members were localized in the nucleus,cytoplasm and chloroplast.ZmDUF668 family proteins can be divided into two subfamilies according to the multispecies phylogenetic tree.Ten Motifs were identified from 19 members of ZmDUF668,all of which contained Motif 1 and Motif 5.Through the analysis of protein conserved domain,it was found that 14 of the 19 members contained not only DUF668 domain but also DUF3475 domain.Gene structure analysis showed that members of the same subfamilies had similar gene structure.Synteny analysis showed that there were 21 collinear relationships between 13 ZmDUF668 genes and 10 OsDUF668 genes.The analysis of cis-acting elements revealed that the promoters of ZmDUF668 genes widely contained cis-elements that light-response,plant hormones and abiotic stress.The prediction results of protein-protein interaction network(PPI)indicated that only ZmDUF668-10 of the ZmDUF668 family proteins had interaction with other proteins.Analysis of RNA-Seq revealed that the gene expression level of some ZmDUF668 gene family members changed significantly under cold,heat,salt stress and ultraviolet treatment.The response of ZmDUF668 family genes to cold and heat stress was verified by RT-qPCR.Bioinformatics was applied to the analysis of the ZmDUF668 gene family,unveiling the characteristics of the members of the ZmDUF668 gene family,and providing theoretical basis for the subsequent study of the molecular biological function of the ZmDUF668 gene family.

The EXORDIUM(EXO) gene was identified in Arabidopsis thaliana as an brassinosteroid(BR)-responsive gene that promotes plant growth by mediating cell expansion.In order to investigate the function of EXO gene in Brassica napus and its expression pattern in different tissues at flowering stage,we used B.napus Zhongshuang 6 as the material,cloned the sequence of the coding region of the EXO gene named BnEXO,and carried out bioinformatics analysis,and used Real-time Fluorescence Quantification to determine the relative expression of BnEXO gene in B.napus in roots,stems,leaves,petals,buds,and pericarps at flowering stage.The results showed that the CDS sequence of BnEXO gene was 945 bp,BnEXO was a stable hydrophilic non-transmembrane protein,which belonged to secreted proteins and was expressed extracellularly,and the secondary structure of the protein was dominated by the random coil.The results of expression analysis in different tissues showed that the expression of BnEXO gene in different tissues was in the order of petals,pericarps,buds,stems,roots and leaves,and the highest expression was found in petals.In addition,20 B.napus EXO genes(BnaEXO),11 B.rapa EXO genes (BraEXO),and 11 B.oleracea EXO genes (BoEXO)were identified in this study based on the protein sequences of eight EXO gene family members in A.thaliana.Most proteins of gene family members were stable proteins,localized extracellularly,with amino acid lengths ranging from 271 to 411 aa,isoelectric point predictions ranging from 5.76 to 9.60,and molecular masses ranging from 28.76 to 46.21 ku.Phylogenetic analysis classified the EXO genes into five subgroups,EXOA,EXOB,EXOC,EXOD,and EXOE,with the least number of members in the EXOB subgroup.Gene structure analyses showed that most members contained only one exon and no intron,and the sequences of EXO gene family members were highly conserved.The results of cis-element analysis of the promoter region of the members in B.napus indicated that the BnaEXO genes play important roles in plant growth and development and in adversity stress.

As a key regulating factor in response to various abiotic stresses,plant heat shock transcription factor (Hsf) has a big family,and diverse structure,characteristics and functions.Hsf not only directly regulates Hsp and other relative gene expression and participates in the processes of response and adaption to various abiotic stresses,but also mediates many life activities regulation.Since the first Hsf was cloned from yeast in the 1980s,more and more Hsfs from other species have been identified and studied.In the previous reports,the identification of the Hsf family in plants was performed only in model species such as Arabidopsis and tomato.Furthermore,the studies is mainly focused on the HsfA subfamily,with few studies on the HsfB subfamily.And,the precise function of HsfC family is also largely unknown.With global climate change,the frequent occurrence of extremely high temperature events has seriously threatened the yield and quality of wheat,maize and other crops.To deal with the threat posed by heat stress,unraveling the mechanism of thermotolerance,identifying functional the targeted Hsfs and improving stress tolerance of crop through biotechnology methods is important.The number of Hsf family in field crops is various,the genome is complex,and the related research started lately compared with model species.To this end,our laboratory began to study the Hsf family of crops in 2009.Based on the latest genomic information,we confirmed the number of members,the modular structure and the spatio-temporal expression pattern of Hsf family.At the same time,with the help of transgenic wheat and mutant by genetic transformation and the CRISPR/Cas9 clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein mediated genome editing technology,several Hsfs were cloned and their regulatory functions of thermotolerance were identified,and some mechanism of thermotolerance was clarified.Our research not only enriched the theoretical basis of thermotolerance,but also provided new germplasm for biological breeding.At present,many studies have reported on functional identification and transcriptional regulation of Hsfs,however,evidences lack on which upstream component mediate Hsf's participation in regulation of thermotolerance,and the related mechanism is still unknown.Based on previous research results about wheat and maize Hsf families of lab,and many relative reports published in public,we reviewed the roles and mechanisms of plant Hsf in regulating process reported in recent years,aiming to promote research in illustrating the extensive and special roles and regulation network of plant Hsf family further,and dig useful genes and selective QTLs for biological breeding for plant thermotolerance.

The WRKY transcription factor family plays an important role in regulating abiotic stress.To systematically analyze the sequence characteristics and expression patterns of NtWRKY11 gene in tobacco,and explore the response mechanism of NtWRKY11 under abiotic stress such as low temperature and drought,the full-length cDNA sequence of NtWRKY11 gene was amplified by PCR using common tobacco cDNA as a template.The basic properties of NtWRKY11 protein were analyzed by bioinformatics software and the subcellular localization of NtWRKY11 protein was studied by constructing a plant expression vector.The expression of NtWRKY11 gene was detected by qRT-PCR technology in different tissues at the full flowering stage and under different abiotic stresses.The results showed that the full-length cDNA of NtWRKY11 gene in tobacco was 999 bp,encoding 332 amino acids,and it shared 54.23% similarity with ramie BnWRKY11.Its promoter region contained three kinds of cis-acting elements(one MBS,one MYB and three ARBE),which probably worked together to enhance the drought resistance of plants.It also contained three salicylic acid-responsive cis-acting elements(TCA-element),which could improve the low temperature tolerance of plants.Subcellular localization results indicated that the NtWRKY11 protein was located in the nucleus.The expression analysis of NtWRKY11 in different tissues at the full flowering stage showed that NtWRKY11 was highly expressed in old leaves,significantly higher than in roots.However,the expression in flowers was significantly lower than that in roots,and there was no significant difference between stems,new leaves and roots. The expression analysis under abiotic stress showed that the relative expression of the gene was significantly higher than that of normal (CK).The relative expression level under high salt stress was not significantly different from CK,while the relative expression level under high temperature stress was significantly lower than CK.All in all,NtWRKY11 is highly expressed in old leaves,and its expression level is enhanced under abiotic stresses such as drought stress and low temperature stress,indicating that this gene acts as a forward transcription factor to regulate drought and low temperature stress.

The folding mode of Chinese cabbage leaf ball is the main character that determines the appearance shape,taste and stress resistance of commercial organs.In order to explore the internal molecular mechanism of the formation of the folding mode of Chinese cabbage,we cloned the full length sequence of the transcription factor BrPIF5 gene from overlaping and outward-curling Chinese cabbage as experimental materials,and conducted bioinformatics analysis,constructed the plant overexpression vector,and used Agrobacterium to mediate the transformation into tobacco to obtain positive transformation plants.The expression level of BrPIF5 gene in tobacco was detected by qRT-PCR.The results showed that the protein encoded by BrPIF5 gene was a hydrophilic protein with a continuous and complete open reading frame of 634 bp,containing 210 amino acids.The protein was composed of more α-helical structure and random curl,including an AP2/ERF domain.BrPIF5 protein and the other 9 gene family members contained a conserved motif 1,and the position was different from that of other gene family members,which was located in the front of the protein sequence.Phylogenetic tree showed that BrPIF5 gene had close evolutionary relationship with SoPIF15,BhPIF1,BoPIF4,AtPIF4 and BrPIF4 family members.The tobacco strain with overexpression of BrPIF5 was obtained by Agrobacterum-mediated genetic transformation,and the leaves of the tobacco positive transformation strain showed inward curling.qRT-PCR showed that the expression level of BrPIF5 gene in the overlaping Chinese cabbage was higher than that in the outward-curling Chinese cabbage,and the gene expression level in the positive tobacco plants was higher than that in the control.It was further proved that BrPIF5 gene controlled the inward curling of Chinese cabbage leaves,thus promoting the formation of leaf ball folding type.

In order to identify the key genes controlling rind hardness and breed crack-resistant watermelon varieties.An F₂ segregating population was created using the high-firmness line 901 and the low-firmness line BSH.Both BSA-seq and RNA-seq approaches were utilized to map the genes responsible for rind hardness.The results of BSA-seq revealed an interval region of 2.14 Mb on chromosome 10,spanning from 1 620 000 to 3 760 000,where the intersection of SNPs and InDels identified 150 candidate genes.Among these,two genes showed non-synonymous mutations,and one gene exhibited a frameshift mutation.Correlation analysis between BSA-seq and RNA-seq identified 6 correlated genes,including Cla97C10G187120, Cla97C10G187020,Cla97C10G187430,Cla97C10G187510,Cla97C10G187280,and Cla97C10G186540.Through bioinformatics analysis,the candidate gene Cla97C10G187120 was identified.The result of qRT-PCR indicated that the transcriptome data was reliable.And the relative expression of the candidate gene Cla97C10G187120 was lower in the line 901 than the line BSH.This study lays a crucial foundation for understanding the molecular mechanisms underlying watermelon rind hardness.

The aim of this study was to investigate the correlation of single nucleotide polymorphisms of the carboxyl ester lipase gene (CEL) with eye muscle area in Hu sheep.The 1 049 Hu sheep with accurate phenotypic records and in good health were selected,their eye muscle area was determined after slaughter,and blood was collected for genomic DNA extraction.Amplification of target fragments and genotyping of CEL gene by PCR and KASPar SNP typing,and association analysis with eye muscle area of Hu sheep,RT-qPCR was used to detect the expression of CEL gene in 10 tissues of Hu sheep.The results showed that the Hu sheep CEL gene was expressed in different tissues and was highest in the duodenum.There was a synonymous mutation of g.4039718 C>T in this gene,which was moderately polymorphic.Trait association analysis showed that this polymorphic locus was significantly associated with eye muscle area in Hu sheep,with TT-type individuals having significantly higher eye muscle area than CC-type individuals.Descriptive statistics showed that the coefficient of variation for eye muscle area was 12.29%,which had high selection potential.Correlation analysis showed that eye muscle area was significantly and positively correlated with growth and slaughter traits such as body weight,body height,breast circumference,slaughter rate,carcass weight and live weight before slaughter.The results indicated that the g.4039718 C>T polymorphic locus could be used as a candidate molecular marker for genetic improvement of eye muscle area traits in Hu sheep.

This study aimed to investigate the function of the anthocyanidin glycosyltransferase gene(UFGT)in the anthocyanin biosynthesis in common buckwheat.Guihong 1 was used as the experimental material.A UFGT gene,named FeUFGT1,was obtained by homologous alignment analysis,and cloned followed by performing on bioinformatics analysis.Overexpression vector of this gene was constructed,then transferred into Arabidopsis thaliana anthocyanin 3-O-glycosyltransferase mutant atugt78d2 to assess its function.The results revealed that the open reading frame of FeUFGT1 was 1 404 bp and encoded 467 amino acid residues with speculated molecular weight of 51.46 ku and theoretical isoelectric point of 4.97.FeUFGT1 protein contained GT-B type glycosyltransferase family domain and a PSPG-box,specific one of the plant UGT family at the C-terminus.The phylogenetic analysis showed that FeUFGT1 protein was closely related to the 3-O-glucosyltransferase of Siraitia grosvenorii UGT74AC1.The expression level of FeUFGT1 in the white-flower petals of Fengtian 1 was 3.7-fold as high as that in the red-flower petals of Guihong 1 with a significant difference.Mutant recovery experiment showed that FeUFGT1 could restore the phenotype of the Arabidopsis mutant which lacked anthocyanin accumulation.

In order to further explore the primary QTL loci for grain-related traits in wheat and explore the genetic relationships among grain traits,124 DH populations constructed from wheat varieties AN859 and WN988 with large differences in grain traits were utilized as research materials,The phenotypic values of grain length,grain width,and thousand grains weight were measured in seven environments over two years,respectively,to carry out the multiple regression analysis of grain traits,and QTL detection of grain-related traits was performed based on the 55K microarray data of the DH populations.The results showed that grain width contributed most to thousand grains weight in the multiple regression analysis.QTL localization for grain traits by complete interval mapping,a total of 69 QTLs related to grain traits were detected on 19 chromosomes except chromosome 6D and 7B,including 24 QTLs for thousand grains weight,28 QTLs for grain length,and 17 QTLs for grain width,with phenotypic interpretations of individual QTLs ranging from 6.87% to 27.74%.Among them,grain length-associated Qgl.ahau-7A.1 on chromosome 7A was detected under seven environments and BLUP,with a phenotypic interpretation rate of 9.48%—22.26%,an additive effect of 0.11—0.21 mm,and a physical interval of 4.91 Mb(AX-110430243—AX-110442528),for the new primary effector QTL.Therefore,the Qgl.ahau-7A.1 locus can be used as a region of focus for subsequent fine localization and molecular marker-assisted breeding.

To explore the effects of different farming modes on the growth and development,photosynthesis,leaf structure and yield of maize,and to provide a theoretical basis for optimizing the cultivation measures and creating efficient planting patterns of maize in Hexi oasis irrigation area.Two tillage methods,no-tillage(NT)and conventional tillage(CT),and three planting patterns,wheat-maize intercropping(W/M),winter rapeseed-maize rotation after wheat(W-G→M),and wheat-maize rotation(W-M),were set up in the experiment,with a total of 6 treatments.The results showed that compared with CT,the plant height,stem diameter and leaf area of NT maize increased by 6.83%,4.10% and 3.97%,respectively.The dry matter quality of intercropping maize was higher than that of rotation,but the difference was not significant.The leaf pigments increased first and then decreased with the growth period,which showed that NT chlorophyll a,b and carotenoids were 11.93%,22.41% and 13.43% higher than CT,respectively,and the difference was significant.The net photosynthetic rate(Pn)and intercellular CO₂ concentration(Ci)of NT leaves were 9.17% and 3.81% higher than those of CT.The stomatal conductance(Gs)and transpiration rate(Tr)of CT treatment were 9.95% and 1.48% higher than those of NT.The leaf structure of NT maize was better,the mesophyll cells were more and arranged in order,the vascular bundles were clearly visible,the garland structure was larger,the palisade tissue and sponge tissue were rich,and the leaf thickness of NT was 2.51% thicker than that of CT,and the difference was significant.The yield of NT maize increased by 8.02% compared with CT,and the yield benefit of intercropping was greater than that of rotation(LER>1).This study found that the growth and development,leaf structure and yield of no-tillage maize were better than those of traditional tillage,and wheat intercropping maize could be promoted as the main farming mode in this area.

In order to investigate the function of watermelon calcium-dependent protein kinase (CDPK) in grafted seedlings and abiotic stress environments, this study used RT-PCR technology to clone the ClCDPK(Cla97C01G019720) gene from watermelon grafted seedlings and performed bioinformatics analysis on it. Further designed specific primers with Kpn Ⅰ and Sal Ⅰ enzyme cleavage sites based on the ClCDPK sequence,conducted amplification and double enzyme cleavage, and connected with pCAMBIA1300 to successfully construct the expression vector pCAMBIA1300-35S-ClCDPK for the target gene.Using RT-qPCR technology, the gene expression levels of ClCDPK were measured in self rooted seedlings (ZG) and grafted seedlings (JJ) after being subjected to salt and drought stress, respectively.The results showed that the ORF of ClCDPK gene was 1 647 bp, encoding 548 amino acids. Its protein contained STKc_CAMK and FRQ1 functional domains, and was a hydrophilic protein. Subcellular localization prediction showed that the protein was located in the nucleus. Evolutionary tree analysis of ClCDPK with CDPK from six other plants revealed that it was closely related to CDPK from Cucurbitaceae melons and pumpkins, with protein sequence homology alignment exceeding 92.64%, indicating high homology.The RT-qPCR expression results showed that the expression level of ClCDPK in grafted seedlings was significantly higher than that in self rooted seedlings. With the duration of stress, the expression levels of ClCDPK in grafted and self rooted seedlings first increased and then decreased, and under the same stress treatment, the expression level of ClCDPK in grafted seedlings was higher than that in self rooted seedlings.This study indicated that ClCDPK responded positively to salt and drought stress, and the ability of grafted seedlings to resist stress was higher than that of self rooted seedlings. It is speculated that ClCDPK is one of the key factors in watermelon's response to grafting, thereby improving the salt and drought resistance of watermelon grafted seedlings.

Germination control is an important basis for promoting rapid emergence and uniform seedlings of Brassica napus.In order to explore the effect of exogenous sucrose on B.napus seed germination and seedling development,it selected the early-maturing rapeseed variety-Xiangyou 420 to conduct germination tests under different conditions and detected the expression changes of moisture,abscisic acid,endogenous sugars and key genes.The results showed that exogenous sucrose delayed the germination initiation process of Xiangyou 420 seeds,improved the uniformity of germination,promoted the cotyledons to turn green and formed true leaves after the cotyledons expanded,inhibited the excessive elongation of the main root and promoted the development of lateral roots.Exogenous sugar first inhibited and then promoted water absorption during the germination process of Xiangyou 420 seeds.It inhibited water absorption within three hours after sowing and released the inhibition of water absorption within six hours after sowing.The ABA content in seeds continued to decrease during the germination process,and exogenous sucrose promoted the decrease in ABA content within three hours after sowing.The total soluble sugar content decreased rapidly within 15 to 18 hours after sowing,and the reducing sugar content increased with the germination process in the early stages of germination.Exogenous sucrose had no significant effect on the changes in total soluble sugar content but inhibited the increase in reducing sugar content within nine hours after sowing and promoted the increase in reducing sugar content 15 hours after sowing.During the germination process,the expression of two sucrose phosphate synthase (BnaSPS) genes decreased significantly during the radicle germination stage and then increased after the cotyledons turned green.Exogenous sucrose inhibited the expression of the two BnaSPS genes before radicle germination, and significantly induced,the expression of BnaC09g37470D gene after the cotyledons turned green.In summary,it preliminarily reveals that exogenous sucrose regulates water absorption at the initial stage of imbibition through osmotic effects,and delays the initiation of B.napus seed germination by regulating endogenous sugar utilization in seeds during the germination stage and sugar conversion in the early stages of seedling development through sugar metabolism,promotes the seed germination process after imbibition,improves the uniformity of seed germination,and promotes the development of seedlings.

By screening differentially expressed genes and metabolites in the green and purple leaves of Brassica napus,it lays a theoretical foundation for the analysis of mechanism of anthocyanin synthesis.Phenotypic observation of GL and PL seedlings showed that PL exhibited a mottled distribution of purple when two true leaves were grown(3 weeks),as the purple gradually deepened during development,it gradually became lighter and eventually disappeared after the bolting stage(>16 weeks),while GL leaves remained green throughout the entire period;transcriptome analysis identified 2 523 co-differentially expressed genes at weeks 6,13,and 16,and these differentially expressed genes were significantly enriched in the anthocyanin synthesis pathway.Twenty-four genes related to anthocyanin synthesis were significantly differentially expressed,including 3 MYBL2,1 C4H,1 F3H,2 F3'H,2 TT8,3 DFR,4 ANS,5 UGT,and 3 TT19.The expression levels of these genes in PL leaves were higher than those in GL at 6 and 13 weeks.Eight differentially expressed genes were selected for qRT-PCR validation and the results were consistent with the trend of transcriptome analysis data.A total of 50 anthocyanins were detected in the anthocyanin-targeted metabolome,of which 29 showed significant difference in accumulation;compared to GL,PL had 16 types of anthocyanin accumulation upregulated and 13 types of anthocyanin accumulation downregulated.The up-regulation of TT8 and its target genes(DFR,ANS,UGT,TT19)in early development(6—13 weeks)promoted the accumulation of cyanidin-based anthocyanins and the decrease of petunian-based anthocyanins in PL.The expression levels of differentially expressed genes and metabolites in the anthocyanin synthesis pathway reached significant levels.

Modern soybean cultivars typically display yellow to brown pods,while their wild ancestral specie,Glycine soja,possesses black pods.Pod color is an important domestication trait and phenotypic characteristic,which is strongly related to pod blasting habit and avoidance of predation.Two alleles were certified to control the pod color in soybean,among which the brown pod L2 gene has not been identified.In order to identify L2 gene on the soybean genome,and provide a theoretical basis for functional analysis and breeding application of brown pod related genes in soybean.The cultivated varieties Zhonglongyou 203(yellow pod)and wild varieties FF1235(black pod)were used as parents to generate an F₂ segregating population for genetic analysis in this study.The BSA-seq was performed using two gene pools which were constructed by brown pod and yellow pod individuals from the F₂ population,respectively.On this basis,recombinant exchange individuals were analysed.The results showed that brown pod was a quality trait controlled by a pair of alleles in soybean.The brown pod L2 gene was located in the 0—0.75 Mb region of Chromosome 3.By further use of 7 polymorphic InDel markers in fine mapping,the candidate interval was finally delimited between Indel-L2-3 and Indel-L2-6 with 344 kb physical distance.There were 32 candidate genes in the interval,among which Glyma.03G005700 gene was annotated as isopropylmalate polymerase.Glyma.03G005700 gene is highly homologous to the discovered black pod gene L1 (Glyma.19G120400),which may be responsible for converting 4-hydroxypyruvate into eucomic acid and piscidic acid,and may be a key gene in the regulating the formation of brown pod in soybean.

Calmodulin-like proteins(CMLs),one of the Ca²⁺ receptors in plants,are involved in the process of plant growth and development,as well as adaptation to environmental changes.To understand the sequence characteristics of garlic CMLs and their responses to osmotic stress,AsCML15 and AsCML42 genes from garlic variety Cangshan siliuban were cloned,and their expression patterns under drought and salt stress conditions were determined.The results showed that the open reading frame of AsCML15 and AsCML42 genes were 498 and 543 bp in length,respectively,encoding 165 and 180 amino acid residues.AsCML15 and AsCML42 harbored four and three EF-hand domains,respectively.AsCML42 was closer to Arabidopsis AtCML42 and AtCML43 in evolutionary relationship,whereas AsCML15 was more closely related to Arabidopsis AtCML15 and AtCML16.Real-time Quantitative PCR technology showed that AsCML15 and AsCML42 were expressed in bulbs,leaves,and roots,and these two genes can be induced by 200 mmol/L NaCl and 15% PEG6000.The AsCML15 and AsCML42 genes may be involved in the process of garlic resisting salt and drought stress,and their biological functions can be further identified.

This study explored the effects of intercropping and rotation on the growth,yield and quality of continuous cropping peanut,to provide theoretical basis for achieving high yield in peanut production.From 2022 to 2023,sweet potato-peanut rotation system(PSP)and maize-peanut intercropping and rotation system(PMP)were set up in the experimental farm of Henan University of Science and Technology on the basis of continuous cropping peanut for 2 years and 11 years respectively,with continuous cropping peanut as control(CCP1 and CCP2,respectively).The effects of PSP and PMP on photosynthetic characteristics,root characteristics,dry matter accumulation and distribution and yield of peanut were studied.The results showed that compared with CCP1,the leaf area index(LAI)of rotating peanut in PSP system(SRP)was significantly increased by 35.08%—53.68% and 24.32%—33.52% at pod-setting stage(PSS)and full pod maturity stage(PMS),respectively.The SPAD value at PSS and pod bulking stage(PBS)increased by 11.93%—18.55% and 5.95%—9.63%,respectively.Compared with CCP2,the LAI of rotating peanut in PMP system(MRP)increased by 46.81%—57.96% and 27.00%—61.78% at PSS and PMS,respectively.At PSS and PBS,compared with CCP2,the SPAD value of MRP and intercropping peanut(MIP)increased by 3.32%—3.69%,7.50%—8.64% and 5.47%—18.37%,15.73%—31.11%,respectively.At PSS and PBS,compared with CCP1,the net photosynthetic rate of SRP increased by 23.68%—41.31% and 26.52%—32.55%,and compared with CCP2,MRP increased by 12.77%—17.81% and 16.88%—62.07%,respectively.They both significantly improved the root length and root tip number,and promoted the dry matter accumulation and the distribution to pods during PMS,and the yields increased by 31.42%—47.36% and 54.12%—75.09%,respectively.Compared with CCP2,MIP reduced the LAI,net photosynthetic rate,root length,root tip number,as well as dry matter accumulation and yield of peanut under the influence of maize shading.At the same time,the content of peanut oleic acid and oleic acid-linoleic acid ratio was significantly increased after rotation.Among them,SRP increased by 1.63—1.65 percentage point and 6.59%—10.52%,respectively,compared with CCP1,and MRP increased by 1.95—2.82 percentage point and 9.75%—14.16% compared with CCP2,respectively.In summary,sweet potato-peanut rotation and maize-peanut rotation increased the peanut yield compared with continuous cropping peanut,the reason was that sweet potato-peanut and maize-peanut rotation promoted peanut root growth,delayed the leaf senescence,and increased photosynthetic rate,especially the photosynthetic rate during late growth period,which promoted the dry matter accumulation and distribution to seeds.Besides that,they could improve the quality of peanut to a certain extent.

In order to study the correlation between stem traits and the formation of elasticity and density tolerance of maize,and further reveal the mechanism of lodging resistance,6 maize varieties with different lodging resistance were selected as materials,and 3 planting densities of 6.0×10⁴,7.5×10⁴ and 9.0×10⁴ plants/ha were set.The stem pull lodging angle in the field was used as the evaluation index of elasticity,and the morphological characteristics between plants and basal nodes.The internode anatomical structure,material accumulation and mechanical characteristics were analyzed.The results showed that plant height,ear height,basal internode length,thickness,epidermal thickness,crusty tissue thickness,total vascular bundle,sheath area of small vascular bundle,fresh weight per unit length,dry weight per unit length and content of each component,puncture and folding strength had significant effects on the stem pull lodging angle,among which internode diameter(r=0.521^**)and dry weight per unit length(r=0.562^**)had the greatest effects.The greater the planting density,the smaller the internode diameter,the thickness of the hard skin tissue,the total number of vascular bundles,the fresh weight per unit length,the dry weight,the content of each component,the lodging resistance,the worse the elasticity of the stem.There were significant differences in stem traits among different varieties.The elasticity related traits of Lishou 1,Chuangyu 107,Jingnongke 728 and MC278 were better than those of other varieties,and the variation amplitude was smaller with the increase of density,so the elasticity and density tolerance were stronger.Traits such as internode diameter and dry weight per unit length had significant effects on stem pull lodging angle, that was stalk elasticity,and the variation amplitude of these traits after densification determined the density tolerance of stalk.

To explore the effects of fertilizer reduction and organic fertilizer application on physicochemical properties and microbial community structure of tobacco-planting soil,and provide theoretical reference for fertilizer reduction and rational application of organic fertilizer in tobacco production.Using conventional fertilization without organic fertilizer as control(CK),Illumina high-throughput sequencing technology combined with bioinformatics,the changes of soil physical and chemical properties and microbial community structure under different treatments such as 10% reduction of fertilizer(T1),10% reduction of fertilizer+sesame cake fertilizer(T2),10% reduction of fertilizer + humic acid fertilizer(T3),and 10% reduction of fertilizer + sesame cake fertilizer + humic acid fertilizer(T4)were analyzed.The results showed that compared with CK,soil nutrient content and soil enzyme activity decreased under T1 treatment,and soil physical properties were slightly improved.Combined with organic fertilizer,soil nutrient and physical properties were further improved,bulk density decreased,moisture content and porosity increased,and the contents of available phosphorus and available potassium under T2 and T4 treatment were significantly higher than those under CK and T1 treatment.The enzyme activity of soil treated with organic fertilizer increased significantly.Combined application of organic fertilizer increased the bacteria and fungi in tobacco-growing soil,among which the dominant bacteria were Proteobacteria and Actinobacteriota.Followed by Firmicutes,Bacteroidota,Chloroflexi and Acidobacteriota.The dominant fungal groups are Ascomycota,Anthophyta,Mortierellomycota,Chlorophyta,Ciliophora and Basidiomycota.The Alpha diversity index showed that the reduction of fertilizer decreased the richness of microbial community,but the combined application of organic fertilizer increased the diversity index of bacterial and fungal communities,and the bacterial community richness increased more significantly.RDA analysis showed that the important soil physicochemical factors affecting soil microbial community structure and diversity included organic matter,available potassium,alkali-hydrolyzed nitrogen,available phosphorus and soil physical properties,etc.Soil physicochemical factors had a greater impact on bacterial community structure.In conclusion,under the condition of fertilizer reduction,organic fertilizer combined with organic fertilizer can significantly improve soil nutrients,physical properties,soil enzyme activity and microflora structure,especially when combined with sesame cake fertilizer and humic acid fertilizer treatment.

In order to determine whether Tomato brown rugose fruit virus(ToBRFV)occurs in Zhangjiakou and Yinchuan tomato producing areas,and to explore the genetic information and evolution of ToBRFV,so as to provide an important scientific basis for the diagnosis and control of ToBRFV and the genetic engineering of tomato resistance to viral diseases.Molecular detection was carried out in Zhangjiakou and Yinchuan suspected diseased fruits,and sequence analysis and genome-wide phylogenetic analysis of the virus gene were carried out by relevant molecular biology software.The results showed that the genome structure similarity between the virus isolates from Zhangjiakou and Yinchuan fruit was more than 99% similar to most of the ToBRFV isolates in GenBank,and the tomato fruit virus in the two places was determined to be ToBRFV.ToBRFV isolates were highly regional,and ToBRFV viruses from different regions of China were associated with many countries and regions from Europe and Asia,and the Zhangjiakou isolate was most closely related to the Chinese isolate(MT018320.1),and the Yinchuan isolate was clustered with the Peruvian isolate,indicating that the Yinchuan isolate may have originated from South America.The similarity of the four ORF amino acids of the Zhangjiakou isolate from China(MT018320.1)was the highest,while the similarity between the Yinchuan isolate and the Zhangjiakou isolate was low.In addition,this study found for the first time that the 444th base of the Yinchuan isolate CP protein changed from A to G,resulting in a meaningful mutation of the 131st amino acid of the CP protein from V(valine)to A(alanine).In summary,ToBRFV disease occurred in Zhangjiakou and Yinchuan tomato producing areas,and the virus came from different places in the two regions.

To evaluate the potential of SpCas9-NG in maize genome editing,the key gene for chlorophyll synthesis,ZmSCD,was selected as the target.The absence of this gene causes seedlings to exhibit albinism,providing a visual assessment of editing efficiency.Based on the PAM sequence recognition rules of SpCas9 (5'-NGG-3') and SpCas9-NG (5'-NG-3'),target sites were designed on the second and third exons of ZmSCD.These target sequences were successfully constructed into SpCas9 and SpCas9-NG knockout vectors,which were then introduced into the maize inbred line KN5585 using Agrobacterium-mediated genetic transformation.The callus tissues were cultured until leaf tissues differentiated,and the albinism rate was recorded to determine the editing efficiency of the different editors.The genomes of albino seedlings were then sequenced.Through three rounds of genetic transformation,SpCas9 produced 76,125 and 28 callus tissues,while SpCas9-NG produced 100,69 and 30 callus tissues.The results showed that the gene editing efficiency of SpCas9 across the three transformations was 14.47%,13.60%,and 10.71%,respectively,while SpCas9-NG editing efficiency was 12.00%,10.14% and 13.33%.Sequencing results of the albino seedlings revealed overlapping peaks near the target sites for both editors.The study demonstrates that SpCas9-NG had comparable editing efficiency to traditional SpCas9 in maize,showing similar gene editing capabilities.In contrast,SpCas9-NG has a broader PAM sequence adaptability,allowing for more flexible target design.This flexibility enables more precise and diverse editing within the maize genome.

To explore the expression patterns of potato genes and provide theoretical basis for the subsequent study of gene resistance to potato diseases.Three endogenous small G protein genes StRac5,StRac7 and StRac13 were cloned from potato,and their bioinformatics and expression patterns were analyzed.The results showed that StRac5,StRac7 and StRac13 contained similar conserved sequences to the small G proteins in Arabidopsis and rice,all belonging to ROP proteins.The protein domains of StRac5 and StRac13 were identical with AtRop1,AtRop3,AtRop5,AtRop6 of Arabidopsis and rice OsRac5,OsRac6,OsRac7,and StRac7 was identical with AtRop9.Phylogenetic tree analysis showed that StRac7 belonged to class Ⅱ and was closest to Arabidopsis AtRop9.StRac13 belonged to class Ⅲ and was the closest genetic relative to Arabidopsis AtRop7.StRac5 belonged to class IV and was the closest genetic relative to rice OsRac5 and OsRac6.The relative expression levels of StRac5,StRac7 and StRac13 genes in different tissues of potato were leaf>root>stem.Compared with the expression in stems,the expression of StRac5,StRac7 and StRac13 in leaves were increased by 1 222.4%,2 531.3% and 468.2%,respectively.After inoculation,the relative expression of StRac5,StRac7 and StRac13 genes increased first and then decreased,and the relative expression of StRac5 and StRac13 genes increased by 62.2%,40.4%,respectively,compared with 0 h after inoculation 24 h.The relative expression of StRac7 gene was increased by 827.8% at 72 h after inoculation compared with 0 h.After ABA,SA,GA and 6-BA treatment,the relative expression of StRac5,StRac7 and StRac13 genes showed a downward trend.After JA and IAA treatment,the relative expression of StRac5,StRac7 and StRac13 genes showed an up-regulated trend.Therefore,StRac5,StRac7 and StRac13 genes can respond to potato late blight infection,and their expression patterns are different in different tissues and under different hormone treatments.

Photoperiod is one of important environmental factors affecting plant bolting and flowering which are regulated by plant phytochrome proteins.The signal network mediated by PHYB has an important inhibitory effect on plant bolting and flowering.Previous studies revealed that there were large segment insertion/deletion differences between the PHYB gene promoter of the Chinese cabbage late-bolting line 06-247 and the easy-bolting line He102.In order to further investigate the impact of promoter mutation on PHYB and the key genes of its downstream pathways,this study was conducted.Based on bioinformatics methods,the redundancy characteristics of phytochrome genes in genome Chinese cabbage were firstly analyzed.It was found that the Chinese cabbage genome contained six phytochrome genes,of which PHYA had two copies,and PHYB,PHYC,PHYD,and PHYE all had only one copy.Then amino acid sequence alignment was used for screening of the specific sequence of PHYB.Antigenic determination clusters were designed based on the specific sequence and the antibodies against PHYB was prepared.The fluorescence quantitative RT-PCR technology and Western Blot technology were used to study the relative content of PHYB in 06-247 and He102.At the same time,the dynamic changes of key regulatory genes such as CCA1,FLC,CO and FT which in the downstream of PHYB pathway were also compared.The results showed that the promoter mutation caused significant differences both in level of mRNA in 06-247 and then significantly increased the protein level of PHYB.At the same time,the downstream regulatory genes such as CCA1,FLC,CO and FT were highly expressed in 06-247,which had an important impact on bolting resistance of 06-247.

Silk color is an important agronomic trait for determining distinctiveness and uniformity of maize variety.In order to analyze the genetic mechanism of silk coloring characteristics of anthocyanins in maize,a doubled haploid(DH)population with 213 lines derived from green silk inbred line WL134 and purple silk inbred line D7 was used for QTL mapping analysis under the environment of 2022 and 2023,respectively.The results showed that there were significant differences in silk coloring characteristics of anthocyanins among different lines and years,the heritability was 0.864.A total of 9 QTLs were detected in two years.These QTLs with phenotypic variation explained(PVE)ranging from 4.83% to 9.26% were detected on chromosomes 2,3,5,6,8,and 9 of maize.A stable and repeatable site qSC5 located between 19.15 Mb and 19.80 Mb was detected on chromosome 5 in two-year data.The LOD scores of the major QTLs were 4.65 and 5.76 in 2022 and 2023 respectively,with phenotypic variation explained(PVE)of 7.22% and 7.17%.It was a new site for regulating silk coloring characteristics of anthocyanins compared with previous studies in maize.Based on SNP markers on both sides of qSC5,90.91% of the genotypes in extreme purple silk DH lines were CCCC,while only 44.00% of the genotypes in green silk DH lines.This marker was significantly correlated with silk color in maize and might link to key genes regulating silk color characteristics of anthocyanins.

Light-harvesting chlorophyll a/b binding proteins are important in plant photosynthesis and abiotic stress response.To study the characteristics of GhLhcb2A1 and its expression patterns and functions in low temperature and drought response in upland cotton,full-length CDS of GhLhcb2A1 gene was cloned from the leaf cDNA of Jimian 262 by PCR.Bioinformatic analysis was conducted to learn the basic characteristics of the gene.The expression patterns and functions in low temperature and drought response were evaluated by qRT-PCR and virus-induced gene silencing.It was shown that the length of GhLhcb2A1 CDS was 798 bp,encoding 265 amino acids.GhLhcb2A1 was highly expressed in leaves and was significantly up-regulated in leaves and roots under low temperature and drought treatment.Compared with the control,its expression maximized at 3 h under low temperature and drought in leaves with 17.42 and 30.03 folds increase respectively,whereas maximized at 6 h under low temperature and 12 h under drought in roots with 11.65 and 65.04 folds respectively.Subcellular localization assay verified that GhLhcb2A1 was expressed in the chloroplasts of cells.Compared with the control plants,GhLhcb2A1 silenced plants showed a more severe phenotype of water loss and dryness under both low temperatures and drought.The accumulated malondialdehyde content in the leaves of the silenced plants was significantly higher than that of the control,while the proline content and superoxide dismutase activity were significantly lower than those of the controls,suggesting that GhLhcb2A1 silenced plant reduced the resistance to low temperature and drought.The above results implied that this gene played a positive role in regulating low temperature and drought response.

In order to promote the cultivation of lodging resistant varieties of Brassica napus and explore the genetic resources of lodging resistance in the stem of Brassica napus,this study used four lodging resistant and three easily lodging resistant germplasm as materials,and measured the content of five components,including cellulose,hemicellulose,total pectin,protopectin,and lignin,in the lower stem during the flowering period.Transcriptome sequencing analysis was performed on the lower stem of YLS0084(lodging resistant)and YLS1691(easily lodging prone).The results showed that the average content of cellulose and total pectin in the anti toppling material,significantly higher than materials prone to lodging;transcriptome analysis revealed a total of 7 397 differentially expressed genes with upregulation and 9 438 downregulation,which were enriched in pathways such as carbohydrate metabolism,translation,amino acid metabolism,and signal transduction;nine genes related to lodging resistance in rapeseed(BnaA01G0071800ZS,BnaA01G0175700ZS,BnaA01G0205800ZS,BnaA03G0404800ZS,BnaA03G0517200ZS,BnaA05G0431400ZS,BnaA07G0056300ZS,BnaA09G0031300ZS,and BnaC05G0128400ZS)were validated through qRT-PCR,and were significantly upregulated in YLS0084.The results of this study demonstrates that the content of cellulose and total pectin in the stem of Brassica napus has a positive effect on lodging resistance and provides important genetic resources for lodging resistance in Brassica napus.

MADS-box transcription factors are widely found in plants and play important roles in plant growth and development and secondary metabolism.To investigate the expression of MADS-box transcription factor family in different accumulation periods of capsaicin.MADS-box transcription factor family was identified by using transcriptome data from different accumulation periods of capsaicin,and their functions were preliminarily analyzed.Subcellular localization,conserved motifs,phylogenetic tree and chromosomal localization of members of the MADS-box transcription factor family of chili peppers were carried out.The results showed that a total of 95 MADS-box transcription factors were identified in the Capsicum annuum L.transcriptome data;containing 105-395 amino acids;with molecular weight ranging from 11.55-44.46 ku;with theoretical isoelectric points ranging from 5.16-10.01;mainly expressed in the nucleus,and all of them contained MADS conserved structural domains,and phylogenetic analysis showed that MADS proteins could be divided into eight subfamilies.There were 73 CaMADS family members localized to 12 chromosomes.There were 26 differentially expressed MADS-box genes,six of which were up-regulated during C1 vs C2 and down-regulated during C2 vs C3.Based on KEGG enrichment and protein interactions,it was predicted that CaMADS13 might be involved in lignin synthesis in chili peppers.CaMADS24 might be involved in the synthesis of capsaicin and lignin synthesis precursor,coumaroyl-coenzyme A.Bioinformatics analysis was used to identify the MADS-box family of transcription factors in chili peppers,which provides data reference and theoretical basis for in-depth study of the molecular mechanism of action in the secondary metabolism of capsaicin.

In order to screen out suitable water and nitrogen combinations for watermelons in Yellow River irrigation area of Ningxia, different water and nitrogen treatments were designed to study the effects of water and nitrogen interaction on SPAD value of watermelon leaves, fruit quality,yield and nitrogen uptake and utilization. The results showed that SPAD values were higher by W1N4,W2N3,W2N4,W3N3 and W3N4 treatment,the quality was better under nitrogenous fertilizer amounts at N2 and N3.The yield was the highest under W3N4 treatment,reaching 76 565.36 kg/ha and increased by 8.34% to 37.57% compared with other treatments significantly.Followed by W3N2 and W3N3 treatment.Compared with other levels,when the irrigation water level was W1,the water use efficiency of facility watermelon irrigation was higher.Among them,the irrigation water use efficiency of W1N3 and W1N4 treatment was higher,reaching 43.91,45.32 kg/ha respectively,while it was significantly increased by 14.00% to 56.40% from other treatments.Fruit nitrogen accumulation and total nitrogen accumulation under W3N4 treatment were all the highest compared with other treatments significantly,increasing by 22.75% to 192.36% and 17.00% to 123.39% respectively compared with the other treatments.Partial factor productivity of nitrogen and nitrogen fertilizer utilization rate under W3N2 treatment were all the highest compared with other treatments significantly.Partial factor productivity of nitrogen increased by 11.00% to 343.68%separately compared with the other treatments and nitrogen fertilizer utilization rate increased by 3.34 to 10.02 percentage points compared with other treatments.The correlation analysis showed that SPAD,the center of soluble solids,Vc,yield,irrigation water use efficiency and nitrogen accumulation,were all significantly positively correlated with each other,and they were significantly negatively correlated with partial factor productivity of nitrogen and nitrogen use efficiency,the edge of soluble solids was positively correlated with nitrogen accumulation of the plants,and negatively correlated with partial factor productivity of nitrogen and nitrogen use efficiency.To sum up,the watermelon had better quality when nitrogenous fertilizer amounts were N2(80 kg/ha) and N3(160 kg/ha),the yield-increasing effect was the best under the combination of water amount for W3(2 200 m³/ha)and nitrogenous fertilizer amount for N4(240 kg/ha).The interaction between high amount of irrigation water and nitrogenous fertilizer application is beneficial to the nitrogen absorption in watermelon,and the interaction between low nitrogen application amount and high nitrogenous fertilizer amount is conducive to utilization of nitrogen fertilizer.

In order to study the problem of strong winter/spring Brassica napus seed germination and flowering period under different winter sowing dates.Two strong winter rapeseeds and two spring rapeseeds provided by Gansu Agricultural University were used as materials.The experiment was carried out in the experimental field of Gansu Agricultural University from October 2022 to August 2023.The winter rapeseeds was carried out on October 11,2022.The winter/spring rapeseeds was sown every 20 days from December 10,2022,and the sowing ended on February 8,2023.The flowering period was recorded,and the germination seeds of winter rapeseed were sampled every 20 days to determine their physiological and biochemical characteristics and analyze the expression characteristics of vernalization genes(FLC,VRN2,FRI,FT).The results showed that the flowering period of winter/spring rape seeds was different by 22—34 days.The difference of flowering time between autumn sowing and spring sowing was 4—7 days.The flowering time of winter rapeseed in autumn sowing(October 11 th)was close to that of spring rapeseed under different winter sowing dates(December 10th,December 30th,January 19th,February 8th),and the flowering overlap time was as long as 15—20 days.With the delay of the sowing date,the relative expression levels of FLC,FRI and FT genes in germinating seeds of winter sowing were down-regulated.The relative expression of VRN2 gene was down-regulated in the early vernalization and up-regulated in the late vernalization.The activities of superoxide dismutase(SOD),peroxidase(POD),catalase(CAT)and the contents of soluble protein(SP),gibberellin(GA3)and salicylic acid(SA)in germinating seeds were increased in the early vernalization,but those were decreased in the late vernalization.The contents of malondialdehyde(MDA)and abscisic acid(ABA)were increased in rapeseed germinating with the increase of vernalization time.

In order to explore the effects of different irrigation methods and nitrogen fertilizer management on late rice growth,development,yield formation,and nitrogen use efficiency,Y Liangyou 911 was used as test material.Two irrigation methods(W1.flooded irrigation;W2.moistening irrigation) and three nitrogen fertilizer managenments base,tiller,ear and grain fertilizer ratios(N1:5∶3∶2,N2:3∶4∶3 and N3:3∶4∶2∶1) were designed.No fertilization served as the control (CK1.flooded irrigation;CK2.moistening irrigation).Leaf area index,SPAD value of rice leaves,dry matter mass,yield formation and nitrogen use efficiency were determined under each treatment combination.The results showed that compared with W1,the LAI of rice treated with W2 was lower in the early growth stage and higher in the middle and late growth stages.The SPAD value of W1 treatment was higher,but there was no significant difference in SPAD value in the late growth period.Under the same irrigation conditions,compared with N1,N2 and N3 treatments it could delay the decline of LAI and SPAD values in the late growth period of rice.W2 treatment could significantly increase rice dry matter accumulation by 6.61%-16.37% compared with W1 treatment.Under nitrogen fertilizer treatment,the dry matter mass was higher in the early and middle stages of growth with N1,and the dry matter mass was higher in the later stages with N1 and N3.The yield increase of W2 mode was 7.59%-10.47% compared with W1 mode.The yield of W2N3 treatment was 3.24%-14.53% higher than that of other treatment.Although the effective panicle number was lower in W2N3 treatment,other yield components were increased,which resulted in higher yield.During the two years,W2N2 and W2N3 had higher values of total nitrogen accumulation and nitrogen absorption efficiency,W2N3 had higher values of nitrogen agronomic utilization,nitrogen partial productivity and nitrogen harvest index,and W1N3 had the higher value of physiological nitrogen utilization.In conclusion,irrigation methods and nitrogen management significantly affect rice yield and nitrogen uptake and utilization.The W2 (moistening irrigation) coupled with N3 (base fertilizer∶tillering fertilizer∶ear fertilizer∶grain fertilizer=3∶4∶2∶1) nitrogen management method is more conducive to rice dry matter accumulation,yield improvement and efficient use of nitrogen fertilizer,which can not only meet high yield,but also play a role in water saving.It is the best coupling method of water and fertilizer.

In order to further explore the genetic rule of grain and stalk yield of sorghum,grain sorghum Xinliang 52 and Sudan grass TS 185 were hybridized as parents to obtain F₂ and F_2∶3 populations,115 pairs of polymorphic primers were used to construct the genetic linkage map of 430 F₂ progeny populations by interval mapping.A total of 86 QTLs were detected for 12 agronomic traits,including number of tillers, number of blades, stem diameter, panicle length, plant height, stem fresh weight, whole plant fresh weight, shelling rate, panicle weigh, thousand kernels weight, grain weight per panicle and grain number per panicle, with LOD value as the threshold value.The QTL of stem fresh weight was located between sam17164-sam15397 on chromosome 1.On chromosome 2,the QTL of number of tillers was located in Xcup64-Xcup26 region,leaf was located in Xtxp019-sam01138 region,and stem fresh weight was located in Xcup26-Xtxp080 region.Panicle length QTL was located between sam44791 and sam33751 on chromosome 3.The QTL of shelling rate was located at sam39622-sam43980 on chromosome 7.The QTL for whole plant fresh weight located in sam10491-sam17740 on chromosome 8 and the QTL for number of tillers located in sam710901b-sam59778 on chromosome 10 were all newly detected loci.

Tiller-related traits are important characteristics of wheat plant type,which determine plant structure and affect grain yield.In order to understand the inheritance and drought resistance of tiller-related traits in wheat under different water conditions,and to excavate the loci related to tiller-related traits,240 wheat varieties (lines) were selected as the subjects of this study,based on the phenotypic identification of tiller angle,effective tiller number and yield per unit area under normal irrigation (NI) and drought stress (DS) conditions,and the comprehensive evaluation of drought resistance,combined with 90K gene chip,genome-wide association study (GWAS) was to identify genetic loci for tiller-related traits and to screen for superior germplasm.The tiller angle,effective tiller number and yield per unit area showed significant difference,and the coefficient of variation ranged from 0.07 to 0.33.According to D-value,the drought resistance of Zhongyou 206 was the best.A total of 54 stable genetic loci significantly associated with tiller angle and other traits were detected, distributed on all chromosomes except 3D, 4D and 5D. Three identical stable loci were commonly detected under both treatments, located on chromosomes 2B, 4B, and 6B. Additionally, four pleiotropic loci were commonly detected in different traits, located on chromosomes 2B, 2D, and 5B.At the same time,the haplotype analysis of Ra_c491_902 (R²=5.45%—17.91%),which was significantly correlated with tiller angle on chromosome 2B,showed that there were three haplotypes:TA-Hap1,TA-Hap2 and TA-Hap3,the haplotypes (lines) containing TA-Hap1 were mainly derived from Huanghuai winter wheat regain.Five candidate genes related to tiller angle were screened by screening the stable genetic loci detected under different treatments.Gene annotation of the genes selected on Ra_c491_902 showed that the genes encoding cytochrome P450 family protein can be used as important genes such as regulating tillering angle,plant drought resistance and defense,to explore the association between genes and phenotype,and lay the foundation for the genetic improvement of tiller-related traits in wheat.

In plants,the PIP5K2 gene of the phosphatidylinositol 4,5-bisphosphate PIP5K gene family plays a key role in regulating plant growth.To investigate the function of the PIP5K2 gene in castor gene cloning,bioinformatics analysis,and expression analysis of the castor PIP5K2 gene were conducted.The results showed that a gene fragment of 2 136 bp in length was obtained through PCR using castor cDNA as the template.Bioinformatics analysis of the protein sequence encoded by this gene determined that the PIP5K2 gene encoded a protein consisting of 672 amino acids,with a pI value of 6.74 and a molecular weight of 76.47 ku.The protein's average hydrophilicity was -0.636,classifying it as a hydrophilic protein.Its secondary structure included α-helices,β-turns,extended strands,and random coils.Based on the prediction results,the tertiary structure of the PIP5K2 protein was consistent with its secondary structure,and shared a high degree of homology with the Jatropha curcas.The relative expression levels of the PIP5K2 gene were generally high in the five-leaf stage of the female,marker female and bisexual inflorescence types,and were generally low in the main stem panicle flowering stage.The highest relative expression level was observed in the five-leaf stage of the marker female inflorescence,while the lowest relative expression level was observed in the main stem panicle flowering stage of the bisexual inflorescence.The highest relative expression level was approximately 80 times higher than the lowest relative expression level.Among the three inflorescence types,the relative expression level of the PIP5K2 gene was similar in the four-leaf stage.However,compared to different growth stages within each inflorescence type,the expression level of the PIP5K2 gene was significantly higher in the four-leaf stage than in the flowering stage.Based on these results,it can be inferred that the PIP5K2 gene may regulate the mid-stage growth of castor plants and have some correlation with plant dwarfing.

In order to investigate the effects of density on individual and population structure characteristics and yield of winter wheat under rainfed and limited water supply conditions,a field experiment was carried out at Gaocheng Experimental Station,Shijiazhuang City during 2022-2023 season using JM22 wheat cultivar under four densities:which were 1.8×10⁶ (D180),3.0×10⁶ (D300),4.2×10⁶ (D420) and 5.4×10⁶ (D540) plants/ha.Two irrigation treatments for each density,which were no irrigation during the whole growth period (W0) and irrigated once at the jointing stage (W1).The influence of planting density and irrigation treatments on leaf area (flag leaf,top 2 nd leaf,top 3 nd leaf,top 4 th leaf),non-leaf green organs (ear,awns,stem sheath) area,leaf area index,non-leaf green organ area index,dry matter accumulation,photosynthetic active radiation interception rate,water use efficiency (WUE) and yield of winter wheat were studied.The results showed that the leaf area of each leaf layer and non-leaf green organs decreased with the increase of density.D300 treatment got the highest leaf area index and non-leaf green organs index,and significantly higher than that in D540.The contribution of post-anthesis dry matter to grain yield was more than 70%.Compared with D540 treatment,reducing plant density decreased the transfer of dry matter before anthesis,but increased the accumulation of dry matter after anthesis and its contribution to grain yield.With the increase of density,WUE increased first and then decreased,D300 treatment achieved the highest WUE.Specifically,the WUE of D300 was 1.2%-14.4% and 2.5%-12.7% higher than that of other densities under W0 and W1,respectively.Compared with W0,W1 treatment increased the area of leaf and non-leaf green organs of different densities,delayed leaf senescence,and improved the photosynthetic active radiation interception rate of canopy.Ultimately,the grain yield increased by 28.1%-39.7%.Under the conditions of this experiment,D300 treatment increased the leaf and non-leaf green organ area,leaf and non-leaf green organ area index,canopy photosynthetically active radiation interception rate,post-anthesis dry matter accumulation and its contribution to final grain yield.The yield of D300 was 2.4%-6.6% and 0.3%-9.7% higher than that of the other densities under W0 and W1,respectively,which was the optimal density in this study.

To clone the PpLAC7 gene sequence of Jinxiu yellow peach,investigating its gene sequence information and its role in flesh browning.The cDNA sequence of PpLAC7 was cloned from Jinxiu yellow peach by homologous cloning.The promoter region elements,physicochemical properties,secondary and tertiary structure,phylogenetic tree and amino acid sequence alignment of PpLAC7 gene were performed bioinformatics by using bioinformatics software.In addition,it also investigated the subcellular localization and the expression pattern of PpLAC7 during the process of fresh browning in Jinxiu yellow peach.The results of bioinformatics analysis indicated that the promoter region included light responsiveness,MeJA-responsiveness,drought-inducibility,seed-specific regulation elements,etc.The full length of PpLAC7 gene was 1 692 bp,encoding 563 amino acids,molecular weight was 61.867 ku,total number of atoms was 8 621,GRAVY was -0.028,theoretical pI was 5.95,instability index was 36.48,aliphatic index was 87.26.The secondary structure of PpLAC7 protein mainly contained α-helix of 14.74%,extended strand of 28.77%,β-turn of 6.22% and random coil of 50.27%.Phylogenetic analysis showed that PpLAC7 had high similarity to MdLAC7.Deduced amino acid sequence indicated that PpLAC7 protein,the same as other species,also contained three typical copper ion domains.Subcellular localization analysis revealed that PpLAC7 was located in endoplasmic reticulum.The expression of PpLAC7 gene was up-regulated during the storage of Jinxiu yellow peach fruit.Combining the relationship of flesh browning index and PpLAC7 expression,it indicated that PpLAC7 may play an important role in the browning process of Jinxiu yellow peach fruit.

In order to study the differences in yield and nitrogen utilization,transport and accumulation of different winter wheat cultivar(line)types in the Northern Huanghuai,the yield traits and nitrogen utilization efficiency of 29 and 26 winter wheat cultivars(lines)for testing in the Northern Huanghuai wheat area were investigated and analyzed in 2021—2022 and 2022—2023,respectively.Through cluster analysis,wheat was classified into four types:high-yielding,medium-high-yielding,medium-yielding and low-yielding.Nitrogen content of wheat stems,leaves,spikes,and grains at maturity were measured at anthesis and maturity,respectively,to analyze the relationship between parameters related to nitrogen utilization traits and yield.The results showed that the average yields among high-yielding,medium-high-yielding,medium-yielding and low-yielding varieties(lines)differed significantly from 2021 to 2023,with the number of spikes per hectare of high-yielding varieties significantly higher than that of other types.At the anthesis stage,the nitrogen accumulation and distribution rate of each organ were stems and sheaths>leaves>spikes;at the maturity stage,the nitrogen accumulation of each organ was grains>stems and sheaths>spikes>leaves.The post-anthesis nitrogen accumulation,pre-anthesis nitrogen transport and its contribution to the grain of different varieties of wheat were higher in high-yielding varieties(lines),and the pre-anthesis nitrogen transport and its contribution to the grain were greater than the post-anthesis nitrogen accumulation and its contribution to the grain.Nitrogen utilization efficiency,nitrogen harvest index,nitrogen accumulation at anthesis and nitrogen accumulation at maturity were significantly and positively correlated with wheat grain yield.Therefore,water and fertilizer management can be carried out through the nitrogen uptake and transfer pattern and distribution characteristics of different varieties,or selecting and breeding wheat varieties with high nitrogen utilization efficiency to achieve high-yield and high-efficiency production of wheat.

This study revealed the changes of bacterial community structure and diversity in facility cucumber rhizosphere soil affected by different reactors,aiming at providing theoretical basis and practical basis for cucumber rhizosphere soil improvement and sustainable utilization of protected soil.This experiment was based on the V3-V4 region of the 16S rRNA gene,seven treatments namely,original greenhouse soil(CK),untreated cucumber rhizosphere soil for 100 days(CK1) and 200 days(CK2),corn straw bioreactor-treated cucumber rhizosphere soil for 100 days(S1)and 200 days (S2),and sheep manure bioreactor-treated cucumber rhizosphere soil for 100 days(M1)and 200 days(M2).High-throughput sequencing technology using Illumina Miseq was used to analyze the diversity,structure,and physical and chemical properties of the bacterial communities in the rhizosphere soils of different bioreactor treatments on facility cucumber.The results showed that 6 344 OTUs were obtained from soil samples after sequencing,which mainly belonged to 39 phyla,315 orders and 980 genera.M2 treatment could improve the bacterial richness in cucumber rhizosphere soil and significantly increase the diversity of bacterial community.At the phylum level,the dominant population structure of bacterial phylum in soil treated by corn straw bioreactor and sheep manure bioreactor was similar,among which Actinobacteriota and Proteobacteria were the dominant phylum.At the genus level,norank_f_JG30-KF-CM,Arthrobacter,norank_f_norank_o_Gaiellales,norank_f_67-14,Blastococcus,Gaiella and Marmoricola were significantly different among different treatments.According to the composition of bacterial community abundance,M2 and S2 treatments increased the relative abundance of some beneficial bacterial groups in cucumber rhizosphere to some extent.RDA analysis showed that the soil bacterial community was significantly affected by soil environmental factors,and the contents of ammonium nitrogen(P=0.015),total potassium(P=0.002)and available potassium(P=0.005)had significant effects on the bacterial community.Therefore,M2 treatment can improve the bacterial richness in facility cucumber rhizosphere soil,increase the diversity of bacterial community and change the bacterial community structure,which is beneficial to the improvement of facility cucumber rhizosphere soil.

Calcium and potassium are important mineral nutrient elements in wheat.It is significant to explore the related genetic mechanisms and effects on human nutritional health.To provide a theoretical basis for biofortification breeding of trace elements in wheat grains,we used 164 F₆ recombinant inbred lines(RILs)derived from Avocet/Chilero(AC)and 175 F₆ RILs derived from Avocet/Huites(AH).Our investigation focused on phenotypic variations in grain calcium(GCa)and grain potassium(GK)content in five environments.QTL mapping was conducted with diversity arrays technology(DArT)chip.Nineteen QTLs associated with grain calcium content were identified,distributed on chromosomes 1A,1D,2A,2B,3A,3D,4A,4B,4D,5A,5B,7A,7B,and 7D,explaining 3.23%—16.29% of phenotypic variation.Simultaneously,23 QTLs linked to grain potassium content were identified on chromosomes 1B,2A,2B,3A,3B,4A,4D,5A,6A,6B,and 7D,explaining 3.31%—24.66% of phenotypic variation.QGCa.haust-1A,QGCa.haust-AC-5A and QGK.haust-AC-2A.2 were located in multiple environments.QGCa.haust-1A and QGCa.haust-AC-5A explained 7.82%—12.72% and 9.68%—15.57% of phenotypic variation,and the physical intervals were 498.67—532.21 Mb and 461.52—486.26 Mb,respectively.QGK.haust-AC-2A.2 explained 8.15%—15.20% of phenotypic variation,with a physical range of 354.61—462.37 Mb.The genetic effect analysis of QGCa.haust-1A,QGCa.haust-AC-5A,and QGK.haust-AC-2A.2 showed that each locus effectively increased the calcium and potassium content in wheat grain.Aggregation effect analysis indicated that the lines with QGCa.haust-1A and QGCa.haust-AC-5A effect loci had highly significantly higher calcium content than those with only a single locus.In summary,three stable loci of grain calcium and potassium content are mapped on chromosomes 1A,2A,and 5A,which could significantly increase calcium and potassium content in wheat grain.

In order to explore the effects of high temperature stress on different heat-tolerant watermelon inbred lines,heat-sensitive (D27) and heat-tolerant (K53) watermelon seedlings were treated at 42 ℃ for 48 h,and their phenotype,tissue structure,photosynthetic characteristics,antioxidant enzyme activities and osmotic regulators were measured and analyzed every 12 h.The results showed that the leaf thickness,fence tissue thickness,sponge tissue thickness and tissue compactness of heat-tolerant K53 were larger than those of heat-sensitive D27 after high temperature stress.The proportion of sponge tissue in the palisade tissue of D27 decreased more than that of K53.With the increase of high temperature stress time,the net photosynthetic rate (Pn),transpiration rate (Tr) and stomatal conductance (Gs) of the two inbred lines decreased,and the intercellular CO₂ concentration (Ci) increased.And the change amplitude of D27 was greater than that of K53.Among the four photosynthetic pigment contents,the heat-tolerant type was higher than the heat-sensitive type under high temperature stress at different treatment times.The superoxide dismutase (SOD) and peroxidase (POD) of the two inbred lines increased first and then decreased with the increase of high temperature stress time,and the enzyme activity was the highest at 24 h,and the enzyme activity of K53 was significantly higher than that of D27.After high temperature stress,the relative conductivity of the two inbred lines increased,and the relative conductivity of K53 increased less than that of D27.The malondialdehyde (MDA) content of D27 was reduced;the MDA content of K53 decreased after an increase.With the increase of high temperature stress,the soluble protein content and proline content (Pro) of K53 were significantly higher than those of D27 at 24 h.In summary,the heat-tolerant type K53 had a stronger resistance to high temperature stress than the heat-sensitive type D27.

To improve seedling blast resistance of rice cytoplasmic male sterile(CMS)line Taonong1A and its maintainer Taonong1B,the Chinese local rice variety Gumei 4 was used as the donor parent harboring the broad-spectrum and durable blast resistance gene Pigm,and the Pigm-tightly-linked codominant marker T9E3 was developed and employed in marker-assisted selection(MAS)breeding practice.T9E3 showed clear and stable polymorphism between the donor and acceptor parents,amplifying a 926 bp DNA band from Gumei 4 genome,while the PCR product from either genome of Taonong1B or Taonong1A was about 2 000 bp.Greenhouse inoculation assay was performed using thirty-two Magnaporthe oryzae isolates collected from different rice areas,Gumei 4 and the improved rice lines showed much higher resistance frequency(90.63%)than that of two acceptor parents Taonong1B and Taonong1A(9.38%).Accordingly,Gumei 4 and the improved rice lines showed high-level seedling blast resistance(0 grade)but highly susceptible(8 grade)for two acceptor parents in natural nursery.Three blast resistant CMS pure lines(Taonong1A-Pigm-1—Taonong1A-Pigm-3)and the corresponding resistant maintainers(Taonong1B-Pigm-1—Taonong1B-Pigm-3)were preliminarily bred through MAS breeding,and one pair of them,Taonong1A-Pigm-2/Taonong1B-Pigm-2,was screened for potential application duo to the high-level blast resistance and elite sterility,agronomic and yield traits.Taonong1A-Pigm-2 showed complete male sterility with nearly 60% typical abortive pollens and 40% round abortive pollens,higher stigma exsertion rate(71.1%)and lower enclosed spikelet rate(33.6%),Taonong1B-Pigm-2 exhibited excellent agronomic and yield trait indexes,such as moderate sowing-heading duration(69 days),plant height(76.2 cm),main panicle length(28.1 cm),panicles per plant(15.8),spikelets per panicle(115.3)and 1000-grain weight(27.7 g)。New parent materials were created for the application of cytoplasmic male sterile(CMS)line hybrid rice breeding,and the comprehensive improvement of seedling blast resistance and characters of Taonong1A and Taonong1B were realized.

In order to clarify the sequence characteristics of the MYB62 transcription factor,expression changes after stress,and further explore the biological function of the MYB62 transcription factor,kohlrabis were used as plant materials.MYB62 transcription factors were obtained by homologous cloning method and bioinformatics was analyzed.Spatiotemporal expression analysis of MYB62 and its expression after stress were performed by Real-time PCR.Results of gene cloning showed that gDNA length of BocMYB62 gene was 1 353 bp,the length of CDS was 837 bp,which contained four exons and three introns and encoded 278 amino acids.The sequence structure analysis showed that BocMYB62 was a hydrophilic protein with two SANT-MYB domains,belonging to the R2R3-MYB type in the MYB transcription factor family.The spatial structure prediction revealed a typical α-helix structure.Phylogenetic analysis indicated that BocMYB62 was closely related to MYB62 in Brassica napus.The spatiotemporal expression results showed that BocMYB62 was consistently higher in green kohlrabi than that in purple with clear tissue specificity.The expression of BocMYB62 was significantly increased during drought stress with the highest expression under 12 h.The BocMYB62 expression was significantly lower than control and the lowest at 4 ℃ cold stress.These results inferred that BocMYB62 may be involved in the regulation of anthocyanin biosynthesis and the regulation of stress.It provides a theoretical basis for the subsequent functional identification of MYB62 transcription factors.

In order to study the mechanism of PSY1 gene in different ripe fruit colors of peppers, Y15016,Y15016-2,SP01,SP02 and Z1 were used as materials,and the functional properties of PSY protein and the expression of PSY1 gene in different fruit color materials of pepper were studied and analyzed by combining some bioinformatics methods.The results showed that the full-length PSY1 gene could be cloned in all five pepper varieties,and there was no difference in sequence.Gene structure analysis showed that the PSY1 gene contained six exons and five introns with a total length of 2 844 bp,and its CDS contained 1 260 bp and encoded 419 amino acids.Sequence alignment and phylogenetic tree analysis showed that the PSY protein of pepper was the closest to the homologous PSY protein of tomato and tobacco of the same family.The results of qRT-PCR analysis showed that the expression level of PSY1 gene in the five materials was lowest in the root tissue and highest in the leaf tissue.Besides,the PSY1 gene was expressed in all the tissues of these materials.The expression level of orange mutant Y15016-2 was higher than that of wild-type Y15016,while that of yellow mutant SP02 was significantly lower than that of wild-type SP01.At different stages of fruit development,the expression of PSY1 gene increased with the development of fruits,except for the decrease in stage Ⅲ,and reached the maximum value at the maturity stage(Ⅳ—Ⅴ)of different fruit color materials.The results of PSY1 gene promoter analysis showed that there was no difference in the sequences in the test materials.The results suggest that the differential expression of PSY1 gene may play an important role in the formation of peppers with different fruit colors.

To clear under the condition of slow release nitrogen fertilizer applying different content of maize development rule of the diurnal change of photosynthetic characteristics and growth,the light response curve was to fit,so as to farming-pastoral zone of spring maize cultivation and nitrogen utilization to provide theoretical basis.This study took Guangde 5 as the research object,and based on the long-term positioning experiment in 2018,the leaves ear position in spinning—tasseling stage of maize were mesured and analyzed under six nitrogen gradient of N 0(N0,CK), 120(N8), 180(N12), 240(N16), 300(N20),360(N24) kg/ha in 2019 and 2020. SPAD value, intercellular carbon dioxide concentration, stomatal conductivity, diurnal variation of net photosynthetic rate, transpiration rate, light response curve and dry matter accumulation rule.The results showed that with the increase of nitrogen fertilizer application, the changes of SPAD value, net photosynthetic rate, transpiration rate and stomatal conduction degree day of maize increased first and then decreased,and N16 treatment was the highest,and intercellular carbon dioxide concentration was the lowest.The changes of net photosynthetic rate,transpiration rate and stomatal conduction degree day showed unimodal curve.The light response curve fitting analysis showed that the maximum net photosynthetic rate of N16 treatment was the highest.In 2019 and 2020,N16 treatments were 37.48%,29.51%,31.85%,18.17%,37.32% and 80.04%,59.73%,50.30%,6.42%,62.51% higher than N0,N8,N12,N20 and N24 treatments,respectively.Dry matter mass accumulation in whole plant and ear were positively correlated with maximum net photosynthetic rate,apparent quantum efficiency and SPAD value.Comprehensive analysis showed that the most suitable nitrogen fertilizer application rate in western Inner Mongolia was 240 kg/ha.

In order to reveal the molecular mechanism of the response to exogenous selenium of sugar beet,seeds of the sugar beet line HD802 were used as test materials for hydroponic experiments.The concentrations of nano-selenium were set to 0(water spray,control),20,50,80,100,150,200 mg/L,respectively.When the plants grew to 8 true leaves stage,nano-selenium solution was evenly sprayed.After 24 h,the activities of superoxide dismutase(SOD),peroxidase(POD),catalase(CAT)and the content of malondialdehyde(MDA)were measured.After data analysis,50,150 mg/L nano-selenium treated leaves were selected for examination.RNA-Seq was performed to analyze the differentially expressed genes and significant enrichment pathways.The results showed that 50 mg/L nano-selenium treatment had promoting effects on the growth of sugar beet leaves,while 150 mg/L treatment had destructive effects on sugar beet leaves.A total of 9 161 DEGs were identified,of which 3 717 were up-regulated and 5 444 were down-regulated.GO functional enrichment was mainly enriched in processes such as signal transduction,cell communication, integral component of membrane, intrinsic component of membrane, primary metabolic process, organic substance metabolic process, and biological process. The metabolic pathways of KEGG mainly included plant pathogen interaction,plant hormone signal transduction,and other pathways.Transcription factor analysis involved 11 families,including AP2,zf-Dof,HLH,WRKY,HSF_DNA-bind,NAM,zf-BED and Homeobox.It found the optimal concentration for selenium treatment to promote the growth and development of sugar beet seedlings,and preliminarily screened the relevant genes responsive to exogenous selenium in sugar beet.

To investigate the nature and function of PlSPL1 (SPL)gene of Paeonia lactiflora,further elucidate the commonalities and characteristic differences of the PlSPL1 gene in different species,and explore the role of PlSPL1 in the degree of stem straightening in Paeonia lactiflora. The full-length sequence of the PlSPL1 gene was obtained by RACE technology using the stalk of herbaceous peony Hongfeng as the research material,and the structural,physicochemical properties,and phylogenetic relationships of PlSPL1 were analyzed and predicted using bioinformatics software,then,the expression level of PlSPL1 in different developmental stages of herbaceous peony stems was analyzed using qRT-PCR technology,and subcellular localization analysis of the protein was conducted using laser confocal microscopy technology.The results showed that the open reading frame of the PlSPL1 gene was 3 000 bp,encoding 999 amino acids.The protein had a molecular formula of C₄₈₆₉H₇₆₈₂N₁₄₀₆O₁₄₉₇S₄₃,a molecular weight of 111.25 ku,a theoretical isoelectric point of 6.26,encoding a hydrophilic unstable acidic protein,phosphorylation modification predominantly on serine,no signal peptide,with transmembrane structure,and had a secondary structure consisting mainly of random coil.Phylogenetic tree analysis revealed that the PlSPL1 protein was most closely related to peony,followed by a closer relationship with grape;protein sequence comparison analysis revealed that the PlSPL1 protein had a conserved SBP domain which was unique to SPL transcription factor family.The relative expression analysis found that PlSPL1 gradually showed a decreasing trend with stem development,indicating that PlSPL1 was negatively regulating the stem development of herbaceous peony and it was hypothesised that PlSPL1 played important roles in degree of stem straightness;and the subcellular localisation showed that the PlSPL1 protein was localised in the nucleus.The above results indicate that PlSPL1 is participated in the stalk development of herbaceous peony.

To study the effects of ridge height on soil heat status,root growth,and tobacco maturity tolerance in tobacco planting,a 2-year field experiment was conducted using the flue-cured tobacco variety Yueyan 97 as the test material from 2022 to 2023.Three treatments were set up,including a ridge height of 30 cm(CK),a ridge height of 38 cm(T1),and a ridge height of 46 cm(T2),to analyze the changes in soil temperature and heat flux,root appearance and growth indicators,root vitality,and tobacco maturity related indicators under different ridge heights.The results indicated that the daily temperature difference in soil varies with the depth of the soil layer,and the daily temperature difference in the surface layer of the soil was the largest.Increasing ridge height can increase the average temperature of tobacco planting soil by 0.4-1.8 ℃ and increase soil heat flux by 4-56 W/m².In the 2 a experiment,the maximum increase in root length for a ridge height of 38 cm compared to a ridge height of 30 cm was 27.26%,the maximum increase in root dry mass was 26.21%,and the maximum increase in root vitality was 14.97%,with significant differences.Compared with a ridge height of 30 cm,the soluble protein content,peroxidase activity,and cell membrane stability index of a ridge height of 38 cm increased by 17.99%,27.82%,and 9.05 percentage points(2022),respectively and by 10.23%,12.44%,and 8.16 percentage points(2023),respectively.The maximum decrease in malondialdehyde content was 24.84% and 44.43%,respectively.Correlation analysis showed that root activity was significantly negatively correlated with malondialdehyde content,peroxidase activity was extremely significantly positively correlated with root growth indexes, and soluble protein content was significantly positively correlated with root length.In summary,increasing ridge height is beneficial for improving the thermal status of tobacco planting soil,promoting root growth,enhancing root vitality,enhancing leaf antioxidant capacity,and enhancing tobacco maturity tolerance.A ridge height of 38 cm is an appropriate ridge height for promoting root growth and improving tobacco maturity tolerance in southern tobacco regions.

This study systematically investigates the effects of silicon-modified biochar (MSC) on the chemical properties of acidic soil,organic carbon and silicon fractions,and the growth of tomato plants.Silicon-modified biochar was prepared,with a focus on investigating its impacts on carbon and silicon chemical fractions,and the availability in acidic soils;tomato growth and soil microbial activity were also evaluated.The results showed that silicon-modified biochar significantly increased soil pH,cation exchange capacity,electrical conductivity,available phosphorus and potassium.MSC also raised the levels of water-soluble sodium and iron in the soil and enhanced the activities of hydrogen peroxidase and sucrase enzymes,thereby improving soil quality.Both biochar modification and unmodified biochar significantly increased the content of different carbon fractions in the soil.Compared with unmodified biochar,silicon-modified biochar significantly increased soil microbial biomass carbon(21.9%) and water-soluble organic carbon (898.3%).Furthermore,silicon-modified biochar significantly increased the contents of soil available silicon,water-soluble silicon,free silicon,active silicon,iron-manganese-bound silicon and amorphous silicon by 362.6%,158.9%,18.1%,34.9%,193.8%,and 74.1%,respectively.Meanwhile,the application of biochar promoted the growth of tomato plants and the absorption of silicon nutrients,with modified biochar showing more pronounced effects.The accumulation of plant dry matter,silicon content,and absorption rate increased by 82.0%,98.9%,and 261.5%,respectively.In summary,silicon-modified biochar significantly affected the carbon and silicon chemical forms and transformation in the soil,increased soil effectiveness and enzyme activity,thereby promoting nutrient absorption and growth of crops,demonstrating its good potential application in agricultural production.

In order to clarify the effect of fertilizer reduction on wheat yield and starch particle size distribution characteristics, seven fertilization treatments were set up with wheat varieties Longke 1109 and Yangmai 25 as materials.No nitrogen fertilizer (CK),farmer's customary nitrogen application rate (ternary compound fertilizer 750 kg/ha+ topdressing 150 kg/ha urea,CF),slow-release fertilizer 900 kg/ha one-time base application (SF900),slow-release fertilizer 750 kg/ha one-time base application (SF750),slow-release fertilizer 600 kg/ha one-time base application(SF600),slow-release fertilizer 750 kg/ha base application+topdressing 150 kg/ha urea (S750T),slow-release fertilizer 600 kg/ha base application + topdressing 150 kg/ha urea (S600T).The effects of slow-release fertilizers on grain yield and starch particle size distribution of wheat were analyzed.The results showed that under the condition of reducing fertilizer application,the spike number first increased and then decreased,the grain number per spike decreased,the 1000-grain weight increased,and the yield of wheat under SF750 treatment was the highest.The content of wet gluten and protein decreased and the content of starch increased in two stubble.The volume and proportion of surface area of B-type starch in strong and weak grains first increased and then decreased,while the proportion of the volume and surface area of A-type starch grains decreased first and then increased in dry stubble.In rice stubble,While the proportion of the volume and surface area of B-type starch grains increased,while those of A-type starch grains decreased.The gelatinization parameters of the two stubble decreased.In summary,the reduction of fertilizer application mainly affects the grain size distribution of endosperm starch,decreases indicators such as gelatinization parameters,and increases grain yield and starch content,which further decreases the content of wet gluten and protein.Compared with the customary nitrogen application rate of farmers,reducing fertilizer application increases wheat grain yield,increases wet gluten and protein content,and decreases starch content.

In order to explore the improvement effect of calcium fertilizer and ARC microbial agent on red earth dry land with low-calcium,the peanut variety Xianghua 522 was used as the experimental material,and two levels of calcium hydroxide fertilizer(0,750 kg/ha,code Ca₀ and Ca₅₀)and three levels of ARC microbial agent(0,30,60 kg/ha,code A₀,A₂ and A₄)were set to form six treatments to carry out pot experiment.It measured soil nutrient and soil enzyme activity in 0—20 cm arable layer soil at peanut seedling stage,flowering stage,pod setting stage and pod filling stage,and pod economic characters and yield were measured at harvest.The results showed that:single application of calcium fertilizer and the combined application of calcium fertilizer and ARC microbial agent could significantly improve the pH of soil at all growth stages,but ARC microbial agent had little effect on it.Compared with CK(Ca₀A₀),Ca₅₀A₂ and Ca₅₀A₄ significantly increased the content of hydrolyzable nitrogen in soil in the whole stage and the content of available phosphorus in soil in the first three growth stages;the content of available potassium in soil of Ca₅₀A₄ was higher than CK in general,and it was significant at seedling stage and pod setting stage;compared with CK,Ca₅₀A₀ significantly increased the content of exchangeable calcium in soil in four stages,with an increase of 23.78%—56.21%;the content of calcium ion in soil with calcium fertilizer application was significantly higher than that without calcium fertilizer application(the flowering stage was not significant),and it was little affected by ARC microbial agent;the content of organic matter in soil remained stable in the whole growth stage,but Ca₅₀A₄ and Ca₅₀A₂ were significantly higher than CK in each growth stage.Compared with CK,the soil sucrase activity of soil each treatment was significantly increased in four stages,and the increase was the largest in Ca₅₀A₄,ranging from 50.79% to 162.56%;the protease activity of soil was significantly increased by Ca₅₀A₂ in four stages with an increase of 26.58%—244.63%;the acid phosphatase activity of soil was significantly increased by Ca₅₀A₄ and Ca₀A₂ during the whole growth stage;the catalase activity of soil in all treatments showed a decreasing trend in general.All treatments could increase the yield of peanut pod in different degrees,and the effect of calcium fertilizer application was greater than that of ARC microbial agent,among which Ca₅₀A₄ had the best effect,with the pod weight per plant increasing by 12.29%,mainly increased the pod number per plant and the full pod number per plant.To sum up,the combined application of calcium fertilizer and ARC microbial agent has a good interaction effect on improving soil nutrient content,stimulating soil enzyme activity and increasing peanut yield,and the best effect is 750 kg/ha calcium fertilizer+60 kg/ha ARC microbial agent(Ca₅₀A₄),which can provide a theoretical basis for green and high yield cultivation of peanut.

Dihydroflavone 4-reductase(DFR)is a key enzyme in the anthocyanidin synthesis pathway.Its expression is closely related to the accumulation of anthocyanidin,which affects flower color.In order to explore the relationship between DFR gene and the formation of flower color in Meconopsis,red-flowered M.punicea and yellow-flowered M.integrifolia were selected as experimental materials.The DFR gene was successfully cloned from them using RT-PCR technology,and bioinformatics and RT-qPCR analysis were performed on them.The results showed that the cDNA full-length of the two DFR genes in Meconopsis was 1 131,1 125 bp,respectively,encoding 376 and 374 amino acids,named MpDFR and MiDFR. Bioinformatics analysis revealed that both proteins belonged to hydrophilic proteins and possessed the unique NADPH-binding domain and substrate-specific binding domain of DFR.The phylogenetic tree indicated that MpDFR and MiDFR proteins had the closest genetic relationships with Papaver somniferum,Cinnamomum micranthum,Epimedium sagittatum, Macadamia integrifolia and Telopea speciosissima. Motif analysis found that the DFR protein motif is relatively conserved in different plants.The RT-qPCR results showed that MpDFR and MiDFR were expressed in different tissues with tissue specificity.During the bud stage,the expression levels of MpDFR and MiDFR genes were the highest and significantly higher than the other two stages.In addition,the expression level of DFR in M.punicea is higher than that in M.integrifolia.Based on the research findings of Quyan's research team at Southwest Forestry University,it was found that the accumulation pattern of anthocyanin-like compounds in these two species of Meconopsis was consistent with the expression pattern of the DFR gene.Therefore,studying the function of DFR genes is crucial for gaining a deeper understanding of anthocyanin biosynthesis pathways and improving plant flower color through genetic engineering methods.

In order to reveal the physiological mechanism of winter rye in response to low temperature stress,the physiological changes of osmoregulatory substances and antioxidant enzyme activity in the leaves and tillering nodes of Wintergraze 70 and White BK-1 were compared during the cold hardening and rejuvenation periods.The results showed that during the cold hardening period,rye mainly improved its cold resistance by accumulating osmoregulatory substances in leaves and tillering nodes.During this period,the semi lethal temperature gradually decreased,and the semi lethal temperature of White BK-1 reached -9.93 ℃,which was 1.95 ℃ lower than Wintergraze 70 while the soil freezing.The increase of soluble sugar and soluble protein contents in the leaves and tillering nodes of White BK-1 was greater than those of Wintergraze 70,which meant these physiological pathways played an important role in improving the tolerance of White BK-1 to low temperatures in winter.Research on the rejuvenation period found that with the extension of low temperature stress time,the content of malondialdehyde first increased and then decreased.Rye resisted the low temperature of rejuvenation period by increasing the content of osmoregulatory substances,improving the activity of antioxidant enzymes in leaves and regulating the activity of antioxidant enzymes in tillering nodes.White BK-1 suffered less damage under the low temperature stress during rejuvenation period,and its malondialdehyde content was lower than Wintergraze 70 after recovery.In addition,after recovery,two varieties of rye accumulated higher contents of proline and soluble protein in the tillering nodes,which providing sufficient nutrients for growth and development after rejuvenation.

In order to further explore the function of TaMAPK5 in the interaction between wheat and Puccinia triticina,the CDS region of TaMAPK5 gene was cloned,and the prokaryotic expression vector pET28a-TaMAPK5 was constructed and transformed into E.coli BL21(DE3).The optimal concentration,induction time and induction temperature of isopropyl β-D-thiogalactoside (IPTG) induced expression of the target protein were explored,and the target protein was purified by Ni-NTA affinity chromatography.The purified recombinant protein was used to immunize New Zealand rabbits to prepare TaMAPK5 polyclonal antibody.The results showed that the full length of the CDS region of TaMAPK5 was 1 110 bp,and the TaMAPK5 recombinant protein was induced with IPTG at a final concentration of 0.050 mmol/L and incubated at 16 ℃ for 48 h.The recombinant protein was used as an antigen to immunize New Zealand rabbits,and a polyclonal antibody capable of specifically recognizing TaMAPK5 was successfully prepared.The antibody titer was 1∶51 200.The results of Western Blot showed that TaMAPK5 was induced by P.triticina infection in wheat and P.triticina incompatible combinations.The recombinant protein TaMAPK5 was successfully expressed and purified,and its polyclonal antibody was prepared.It was revealed that TaMAPK5 protein may positively regulate wheat resistance to leaf rust infection.

Abstract:To explore the effects of phosphorus fertilizer reduction ratio and phosphorus fertilizer management method on tomato yield,phosphorus utilization rate and soil fertility level under simultaneous reduction of chemical fertilizer nitrogen,phosphorus and potassium in high fertilizer application areas,an experiment was conducted in a greenhouse located in Dingxing County,Hebei Province.Overwintered long-season tomato was chosen as the experimental plant.Treatments included CF(N-P₂O₅-K₂O,1 009.5-774.0-1 458.0 kg/ha),P1(N-P₂O₅-K₂O,750.0-375.0-1 125.0 kg/ha),PB2(N-P₂O₅-K₂O,750.0-225.0-1 125.0 kg/ha),PT2(N-P₂O₅-K₂O,750.0-225.0-1 125.0 kg/ha),P3(N-P₂O₅-K₂O,750.0-75.0-1 125.0 kg/ha)and P4(N-P₂O₅-K₂O,750.0-0.0-1 125.0 kg/ha).Fertilizer phosphorus was applied basally in the PB2 treatment,and the other fertilizer-reduced treatment fertilizer phosphorus was applied in a "Basal dressing and topdressing" method.The result showed that compared to CF,tomato yield of PT2 treatment over the three-year period revealed an average increase of 12.0%,with the highest increase.After three years of fertilizer reduction,the root dry weight of P1,PB2 and PT2 significantly increased,along with improvements in the chemical phosphorus fertilizer utilization rate,phosphorus fertilizer agronomic utilization rate,and the chemical phosphorus fertilizer harvest index.Compared to CF,PT2 treatment resulted in an increase in root shoot ratio of 48.2%, phosphorus fertilizer recovery rate and phosphorus fertilizer harvest index increased by an average of 32.9 and 2.7 percent points, phosphorus fertilizer agronomic utilization rate was 9.02 times higher than that of CF.PT2 treatment was the highest among all fertilizer reduction treatments.Compared to the CF treatment,soil $NO_3^{-}$-N,Available P and Available K contents were reduced by an average of 8.2%—14.9%,4.4%—19.9%,and 7.3%—24.8%,respectively,over the three-year period.In conclusion,a 35.2% reduction in chemical fertilizer,which included a 70.9% decrease in chemical phosphorus fertilizer,did not have a negative impact on yield in greenhouses with excessive fertilizer use.Additionally,the combination of "Basal dressing and topdressing" method for phosphate management enhances tomato yield in comparison to basal dressing alone.This method also reduces available phosphorus content and increases the efficiency of chemical phosphorus fertilizer utilization.

EβF synthetase gene(EβFS)enables the production of(E)-β-farnesene in plants to control aphids.In order to identify the expression characteristics and the function of EβFS from Mentha canadensis,McβFS1 was isolated from mint leaves,and bioinformatics analysis and expression patterns were analyzed.McβFS1 over expression vector was constructed and then was transformed into wild-type Arabidopsis by Agrobacterium-mediated floral dip method.The transgenic Arabidopsis thaliana was subjected to aphids bioassays.The results showed that the coding region of McβFS1 was 1 650 bp,encoding 549 amino acids.The protein molecular weight was 63.85 ku,the isoelectric point was 5.23,and there was no transmembrane domain.The predicted secondary structure contained α-helix,β-turn,extended strand,and random coil,which had 69.58%,3.10%,4.01%,and 23.32%,respectively.Phylogenetic analysis revealed that McβFS1 had a close genetic relationship of 91.47% similarity with Mentha× piperita Q5W283.1.However,the sequence difference is significant,making it a new EβF synthase gene.The qRT-PCR results showed that McβFS1 was expressed in root,stem,leaf and flower,and the expression level in leaf was significantly higher than that in root and stem.The choice test suggested that transgenic Arabidopsis was higher resistant to aphids than wild type.

The hybridization of Lentinula edodes mononuclear mycelium is influenced by A and B two mating type factors.Based on whole genome sequencing,mating type identification of Lentinula edodes mononuclear hyphae can be carried out,but there are limitations in terms of technical complexity and high cost.In order to achieve rapid identification of unknown mating type genes in Lentinula edodes mononuclear mycelium,we used H31 and BJ4 strains as materials,and determined PCR amplification primers that can be used for identifying unknown mating type genes through specific amplification and resequencing of mating type site regions.The specific method was to use the hyphae hybridization results to determine the two mononuclear hyphae that could be hybridized,and record their mating types as A1B1 and A2B2,respectively.Based on the alignment analysis of the A and B mating type site gene sequences obtained from NCBI,primers were designed in the conserved region of their mating type sites to amplify the different mating type mononuclear hyphae A1B1 and A2B2.By comparing the resequencing information of the amplified products,we obtained nonconserved regions in gene sequences of different mating types A1 and A2,B1 and B2.Then,four mating type gene specific amplification primers to identify mating type genes in H31 and BJ4 Lentinula edodes mycelium were designed in this region.Based on the molecular level mating type identification results,hybridization validation was conducted on H31 and BJ4 Lentinula edodes mononuclear mycelium,and the results showed that the molecular identification results of mating type genes were consistent with the hybridization results of mononuclear mycelium.The mating type gene identification method established in this study no longer relies on whole genome sequencing.This method uses Lentinula edodes as experimental material,but it is also applicable to the identification of mating type genes in other edible mushrooms.Therefore,the results of this study have broad application value for genetic breeding of edible mushrooms.

Aluminum-activated malate transporter(ALMT)gene family,as a widely distributed gene family in plants,the encoded protein plays an important role in regulating plant root acid secretion and response to aluminum ions.In order to clarify the sequence characteristics and expression features of H. macrophylla Endless Summer(HmALMT11)under abiotic stress, further exploring the biological functions of H. macrophylla Endless Summer, and providing a theoretical basis for the subsequent identification of the function of HmALMT11.HmALMT11 gene was cloned from the leaves of Hydrangea macrophylla Endless Summer,and the bioinformatics,subcellular localization and aluminum stress response analysis of this gene were also carried out.HmALMT11 contained an open reading frame of 1 590 bp and encoded 529 amino acids,with a molecular weight of 59.1 ku and a theoretical isoelectric point of 9.10 was an unstable basic protein containing 6 transmembrane domains.The subcellular localization analysis revealed that the protein encoded by HmALMT11 was specifically targeted to the plasma membrane.The qRT-PCR analysis revealed that treatments with both low(100 μmol/L) and high(800 μmol/L) concentrations of Al₂(SO₄)₃ increased the expression level of HmALMT11 in the roots,stems,and leaves of Hydrangea macrophylla Endless Summer.Notably,the highest expression level was observed in the roots,demonstrating a clear tissue-specific expression pattern.These results suggested that HmALMT11 protein might play an important role during the regulation of aluminum tolerance in hydrangea.

Soil organic carbon and humus components are affected by soil quality,fertilization management measures and other factors.In order to clarify the regulation effect of long-term chemical fertilizer application on soil organic carbon(SOC)and soil humus components in different soil layers,a 43 a(2021)long-term fertilization experiment was conducted in Laiyang,Shandong Province.Six treatments were selected:low nitrogen fertilizer(N1),high nitrogen fertilizer(N2),high nitrogen fertilizer combined with phosphorus fertilizer(NP),high nitrogen fertilizer combined with potassium fertilizer(NK),high nitrogen combined with phosphorus and potassium fertilizer(NPK)and no fertilizer control(CK).The results showed that compared with CK,N1 could significantly increase the SOC content of 0—5 cm,with an increase of 22.84%.Single nitrogen fertilizer treatment could significantly increase the SOC content of 5—10 cm,with an increase of 20.94% and 28.60% in N1 and N2,respectively.N1 could significantly increase the SOC content of 10—20 cm,with an increase of 17.05%,while other treatments had no significant change.Compared with CK,N1 could significantly increase the content of humic acid(HA)in 10—20 cm and 20—30 cm soil layers,with an increase of 22.86% and 40.49%,respectively,while there was no significant change in 0—10 cm soil layer.NP could significantly increase the content of fulvic acid(FA)in 0—5 cm and 5—10 cm soil layers by 89.44% and 124.63%,respectively.NK could significantly increase the content of FA in 10—20 cm soil layer by 100.22%,and NPK could significantly increase the content of FA in 20—30 cm soil layer by 107.48%.N1 could significantly increase the content of humin(Hu)in 0—5 cm soil layer,with an increase of 69.34%.N2 could significantly increase the content of Hu in 5—10 cm soil layer,with an increase of 66.18%.N1 could significantly increase the content of Hu in 10—20 cm soil layer,with an increase of 79.50%,while there was no significant change in 20—30 cm soil layer.In summary,under the conditions of this experiment,long-term application of chemical fertilizers can effectively improve the fixation of soil organic carbon in non-calcareous fluvo-aquic soil and change the composition of soil humus,and the effects of different fertilization strategies are quite different.Among them,the effect of single application of nitrogen fertilizer on carbon sequestration is better.

Expansin (EXP) plays a crucial role in plant adaptation to environmental stress by regulating the relaxation of cell wall components and enhancing flexibility.The aim of this study was to investigate the characteristics of the EXP family members and their expression patterns under salt stress,and to explore the salt tolerance genes in Aquilegia coerulea.Genome-wide identification and analysis of the AcEXP family of Aquilegia coerulea was using bioinformatics methods and its expression pattern under salt stress was analysed using RNA-seq expression data and qRT-PCR.The results revealed that the entire genome of the Aquilegia coerulea contained 27 EXP genes,distributed across six chromosomes and one scaffold.The secondary structure of proteins was dominated by β-sheet and random coil.The majority of the proteins were hydrophobic,and all 27 EXPs were found in the cell wall.The phylogenetic tree showed that members of the AcEXP family were classified into four subfamilies:EXPA,EXPB,EXLA and EXLB.Members of the same subfamily possessed similar gene structure and protein conserved motifs.The AcEXP genes contained multiple elements that responded to phytohormones and stress.The transcriptome data was used to analyze the expression pattern of AcEXP under salt stress.Twenty-one EXPs responded to salt stress,but the expression patterns were different in roots and leaves.AcEXPA8/9/12,AcEXPB1,AcEXLB1 and AcEXLB2 were differentially expressed in leaves and roots.The qRT-PCR results further verified the expression pattern of AcEXP genes under salt stress.It comprehensively analyse the basic characteristics of the EXP gene family members of Aquilegia coerulea and expression changes under salt stress.The study preliminarily demonstrates that AcEXPA8/9/12,AcEXPB1,AcEXLB1 and AcEXLB2 are involved in the salt stress response process and responded positively.

The study aims to investigate the effect and mechanism of PPARA on intramuscular fat (IMF) deposition,and to search for molecular markers related to IMF deposition in sheep.The experiment was conducted using Small-tail Han sheep (STH) and a hybrid F₁ population between Suffolk sheep and Small-tail Han sheep(STH×SFK)as the research object.By measuring their meat quality,the histological structure and fat drop distribution of longissimus dorsi muscle of two sheep populations were compared by H&E staining and Oil Red O staining;qRT-PCR and WB techniques were used to detect the expression differences of PPARA mRNA and protein levels among different populations,and analyze the correlation between PPARA gene and IMF;Sanger sequencing technology was used to detect the PPARA gene SNP site to evaluate the possibility of using it as a genetic marker related to IMF deposition in sheep.The results indicated that the shear force and water loss rate of STH were significantly higher than those of STH×SFK,but pH was extremely significantly lower than that of STH×SFK,marble score of STH was lower than that of STH×SFK population;histological staining showed that STH muscle fibers were thick and tightly arranged,with IMF concentrated in the muscle fiber gaps,and muscle fibers were thin and loose in structure in STH×SFK,and IMF was evenly and widely distributed among muscle cell and muscle fiber spaces,with a higher content.The expression levels of PPARA mRNA in the STH were significantly higher than those in the STH×SFK population,and PPARA protein expressions of the STH were extremely significantly higher than those of the STH×SFK population.Correlation analysis showed that the expression level of PPARA gene was extremely significantly negatively correlated with sheep fat droplet area ratio and pH,extremely significantly positively correlated with shear force and water loss rate,and weakly correlated with marble score.Sanger sequencing results showed that base mutations T49885C,T50007C,G50013A,G50835A,G50942A and G51154C occurred in the second intron of PPARA gene in two sheep populations,but no C49885T mutation was detected in the STH population.The analysis of SNP genetic diversity showed that the homozygosity of SNP loci in the population was greater than the heterozygosity,and they were in a medium to low degree of polymorphism in the population;except for the G50835A mutation,all other SNPs were in line with the Hardy-Weinberg equilibrium,and with a stable genetic basis.It was believed that PPARA gene had an important impact on IMF deposition in sheep,and could be used as a potential molecular genetic marker for IMF trait selection in sheep.

Leaf color mutant is an ideal material for studying C4 photosynthesis pathway and chlorophyll metabolism mechanism.In order to study the molecular mechanism of yellow-green leaf mutation in millet,and lay a foundation for the functional study of yellow-green leaf genes and the molecular mechanism analysis of chlorophyll metabolism,a stable hereditary yellow-green leaf mutant ygl7 was identified in the ethyl methacrylate(EMS)mutant library of Changnong 35.Agronomic traits,photosynthetic pigment content and photosynthetic parameters,and chloroplast ultrastructure observation were carried out on the mutant and wild type.At the same time,genetic analysis of mutant leaf color was performed,primary mapping was performed by BSA method,fine mapping was performed by an F₂ population,and candidate genes were predicted according to functional annotation combined with RNA-Seq.The expression pattern was analyzed by qRT-PCR,and the protein interaction was verified by yeast two-hybrid experiment.The results showed that the leaves of ygl7 were obviously yellow-green at seedling stage and elongation stage,and gradually turned to light green at heading stage compared with the wild type.The chlorophyll content and photosynthetic rate of ygl7 during the whole growth stage were significantly lower than that of the wild type,and the chloroplast structure was abnormal.Genetic analysis showed that ygl7 yellow-green leaf phenotype was controlled by a pair of recessive single genes.Yellow-green leaf gene was located in the 434.9 kb region of chromosome Ⅶ.Candidate gene analysis predicted that Seita.7G290300 encoding protoporphyrin Ⅸ magnesium chelatase Ⅰ was the candidate gene for regulating yellow-green leaf.The results of qRT-PCR showed that Seita.7G290300 was highly expressed in leaves,and the expression of Seita.7G290300 in mutant was lower than that of wild type.The expression levels of genes related to chlorophyll synthesis pathway(CHLD,CHLI)and photosystem(LHCB1,LHCB6)were down-regulated in the mutants.The experiment of yeast two-hybrid showed that SiYGL7 interacted with MORF2.

The aim of this study was to investigate the effect of Ezrin on the maturation of mouse oocytes,fertilization,and early embryo development,as well as to reveal its mechanisms.Mouse germinal vesicle(GV)stage oocytes and pronuclear-stage embryos were injected with siRNA targeting Ezrin(si-Ez)or a negative control siRNA(si-NC),then the rates of oocyte maturation,fertilization,and early embryo development were calculated,the length and density of microvilli on the surface of the MⅡ stage oocytes were detected with scanning electron microscopy,the spindle localization and cortical granule migration were observed using immunofluorescence staining.The results showed that knocking down Ezrin during the GV stage significantly reduced the maturation rate of mouse oocytes and extremely significantly lowered the developmental rate of early-stage embryos after fertilization.While it decreased the fertilization rate of mature oocytes,the difference was not significant.Knocking down Ezrin during the pronuclear stage extremely significantly reduced the total cell number of mouse blastocysts.Although there was a downward trend in the rates of morula and blastocyst survival,there was no significant difference compared to the control group.Scanning electron microscopy revealed that knocking down Ezrin in GV stage mouse oocytes significantly decreased the length and density of microvilli on the surface of MⅡ stage oocytes.Immunofluorescence results showed that knocking down Ezrin in GV stage mouse oocytes affected the migration of the spindle and cortical granules.These results suggest that Ezrin,through its influence on microvilli formation,spindle localization,and cortical granule migration,blocks the maturation of mouse oocytes,consequently affecting fertilization and early embryo development.

The identification and expression pattern analysis of CmPIPs gene family in melon could provide theoretical basis and support for further exploring the function of CmPIPs gene family and the genetic improvement of melon.TBtools,MEME,MEGA X and Plant-CARE tools were used to analyze the bioinformatics of CmPIPs,and the expression level of CmPIP2;7 in the pericarp of melon at different stages after pollination,and the expression level of each member of CmPIPs in different tissues and different concentrations of plant hormone treated young leaves were visualized in the software GraphPad Prism 10.The results showed that CmPIP2;7 and CsPIP2;8 had nearest kinship;the 12 members of the CmPIPs family were mainly distributed on chromosomes 1,3,4,5,9,10 and 11;except that CmPIP2;8 had 3 CDS regions,the other members had 4 CDS regions.The promoter regions of each member of CmPIPs had multiple cis-acting elements,hormone responsive elements,such as auxin,gibberellin,and abscisic acid.The expression level of CmPIP2;7 was significantly up-regulated during rapid development and maturity of melon fruit.Members of various families of CmPIPs were expressed in different tissues of melon.After treatment with 40.0 μmol/L auxin,the expression level of CmPIP2;4 was significantly up-regulated,while the expression level of CmPIP1;1,CmPIP 2;1,CmPIP2;2 and CmPIP2;3 were extremely significantly down-regulated,and when the concentrations of abscisic acid were 0.4,4.0 and 40.0 μmol/L,the expression level of CmPIP1;1,CmPIP2;1,CmPIP2;3,CmPIP2;9 were significantly down-regulated.After treatment with 44.640 μmol/L methyl jasmonate,the expression level of CmPIP2;1 and CmPIP2;5 were significantly down-regulated,while the expression level of CmPIP2;2,CmPIP2;3,CmPIP2;7 and CmPIP2;9 were significantly up-regulated.When the concentration of ethylene glycol was 4.0 mmol/L,the expression level of each member of CmPIPs was significantly up-regulated.The gene structure,sequence characteristics,evolutionary relationship and collinearity of CmPIPs gene family members were clarified,and their expression pattern were analyzed.

WRKY is a unique class of transcription factors in plants,which plays an important role in plant abiotic stress response,seed dormancy and germination,growth and development,etc.In order to reveal the function and underlying molecular mechanism of GmWRKY44 gene in soybean WRKY transcription factor family,bioinformatics analysis and biology function verification of soybean Williams 82 GmWRKY44 were performed.GmWRKY44 gene was 1 077 bp in length and encoded 358 amino acids;the results of structural prediction and evolutionary analysis showed that,the secondary structure was composed of 23.46% α-helix,4.75% β-fold,58.94% irregular coil and 12.85% extended chain,and the tertiary structure was unified with the secondary structure;It contained a conserved WRKY domain, the zinc finger structure was of the C₂H₂ type, and it belonged to the WRKY IIc subfamily; GmWRKY44 is a homologous gene of Arabidopsis thaliana AtWRKY71 with a similarity of 35.56%, and the two genes had similar gene structures. RT-qPCR analysis showed that GmWRKY44 responded to salt stress and its expression level first decreased and then increased.Under salt stress,the germination rate and root length of wild-type(Col-0)and GmWRKY44 overexpressing Arabidopsis lines were inhibited to a certain extent,but GmWRKY44 overexpressing lines were significantly better than Col-0.In addition,under salt stress,the growth inhibition of GmWRKY44 overexpressing lines was lower than that of Col-0.Physiological index analysis revealed that under salt stress,the overexpression lines of GmWRKY44 exhibited significantly higher activities of superoxide dismutase(SOD),peroxidase(POD),and catalase(CAT)than Col-0,while the content of malondialdehyde(MDA)was significantly lower than Col-0.These data indicated that overexpression of GmWRKY44 could improve salt tolerance in transgenic Arabidopsis.

The aim of this study was to express the ORF3 protein of Avian hepatitis E virus (aHEV)and prepare its polyclonal antibodies,which can provide biomaterials for further study on the biological function of new vaccine of aHEV ORF3 protein.The ORF3 gene of CaHEV-GDSZ01 strain was amplified by RT-PCR,ligated with pET-32a(+)expression vector to construct recombinant prokaryotic expression plasmid,and transformed into BL21(DE3)competent cells to induce expression.The expression of ORF3 protein was analyzed by SDS-PAGE gel electrophoresis and immunization of rabbits to obtain rabbit anti-ORF3 polyclonal antibody.The titer of polyclonal antibody was determined by indirect ELISA,and its specificity was detected by Western Blot and IFA.The pET-32a-ORF3 recombinant prokaryotic expression plasmid was successfully constructed,and the 27 ku insoluble recombinant protein of aHEV ORF3 was successfully expressed by prokaryotic expression system.The results of Western Blot and IFA showed that the prepared polyclonal antibody could specifically recognize ORF3 protein,and the ELISA results showed that the titer of polyclonal antibody was 1∶51 200.In this study,the ORF3 protein of CaHEV-GDSZ01 strain was successfully expressed and its polyclonal antibody was prepared.

In order to investigate the effect of reducing nitrogen and increasing density on the yield formation characteristics of ratooning rice,hybrid rice variety Chuangliangyou 669 was used as the material to conduct two years of field experiments under conditions of three nitrogen application rates(N1:180 kg/ha;N2:153 kg/ha;N3:126 kg/ha)and two plant row spacing(M1:20.0 cm×16.7 cm;M2:16.7 cm×16.7 cm).The results showed that:reducing nitrogen reduced the leaf area index(LAI)of ratooning rice,but increasing density within a reasonable range of nitrogen reduction could increase the LAI of the main and ratooning seasons.The LAI of N1M2 and N2M2 was higher in the interaction treatments.Reducing nitrogen and increasing density both reduced the SPAD value of ratooning rice leaves,but the effect of density was not significant.Reducing nitrogen led to a decrease in dry matter weight,while increasing density could significantly increase dry matter weight.The interaction treatment of N1M2 and N2M2 had a higher dry matter weight.Reducing nitrogen reduced the yield of ratooning rice,but increasing density within a reasonable range of nitrogen reduction could increase yield.The interaction treatment of N1M2 and N2M2 had a higher yield.Reducing nitrogen significantly reduced the number of effective panicles in the main season,the total number of grains per panicle,and the regeneration rate and number of effective panicles in the ratooning season.However,increasing density had a compensatory effect on the number of panicles.Reasonable nitrogen reduction and density increase(N2M2)could coordinate the relationship among yield components and achieve higher yields.The correlation analysis showed that reasonable nitrogen reduction and density increase increased the effective number of panicles and total grains per panicle in the main season,as well as the effective number of panicles in ratooning season mainly by increasing LAI and dry matter weight of the main and ratooning season,and thereby improving the yield of ratooning rice.Overall,the nitrogen reduction and density increase treatment N2M2(nitrogen rate of 153 kg/ha,plant row spacing of 16.7 cm×16.7 cm)can save 15% nitrogen and achieve a higher yield.

In order to analyze the possible role of dTLR2 in duck innate immunity,we study the expression of dTLR2 mRNA in immune organs during 1 to 10 weeks,as well as the expression in blood after challenging with Escherichia coli and Salmonella enteritis by Fluorescence Quantitative PCR.We also prepared polyclonal antibodies of the extracellular domain of dTLR2 by recombinant protein expression for further studying the biological function of dTLR2.The results showed that dTLR2 mRNA was expressed in spleen,thymus and bursa at different weeks.The expression of dTLR2 mRNA was significantly different among different weeks in the spleen and thymus tissues,and there was no significant difference in bursa.After infection with E.coli and S.enteritis,the expression of dTLR2 mRNA was significantly higher than that of the control group in the first and second days,but there was no significant difference among groups in the third day.Analyzing the sequence of dTLR2 gene,we predicted the extracellular domain,designed primers for PCR assay,and built pET28a-TLR2 recombinant plasmid.The recombinant plasmid was translated into E.coli BL21 (DE3) for expression of dTLR2-His protein.The SDS-PAGE results showed that the fusion recombinant protein efficiently expressed in the supernatant,molecular mass was around 29 ku.The results of Western Blot showed that the antibody which harvested from immunizing rabbits could detect recombinant dTLR2 and endogenous dTLR2.The analysis of the expression of dTLR2 and successful preparation of polyclonal antibody provide favorable support for further studying of dTLR2 biology function.

This study aimed to detect the SNPs of CDKN2A gene in Wenshang Barred chicken,and perform the analysis of its bioinformatics and expressions in skin tissues.Based on the high quality chromosome-level genome of Wenshang Barred chicken assembled by the research group of Resource Conservation and Evaluation in Jiangsu Institute of Poultry Science,the sequences were blasted to chicken reference genome (GRCg7b) and the SNPs were acquired.These SNPs were detected and genotyped by MassARRAY nucleic acid mass spectrometry in the chicken population.Bioinformatics software was used to predict the regulatory sequence of CDKN2A gene.The physicochemical properties and structural characteristics of the encoded protein were analyzed.The expressions of this gene in the skin tissues of Wenshang Barred chicken at different periods were detected by transcriptome and Real-Time Fluorescence Quantitative PCR.The results showed that the total length of CDKN2A and its CDS were respectively 9.854 kb and 183 bp,encoding 60 amino acids.Forty-one SNPs were found in this gene and its upstream and downstream 1 kb,of which four SNPs were newly discovered loci.These four new SNPs were located in the downstream and the intron,respectively.The first exon had two SNPs,both of which were missense mutations:V9D and C10R.The detection rates of 34 SNPs among the 41 SNPs were all over 91% in the Wenshang Barred chicken population.These SNPs were almost homozygous mutants without polymorphism.Eight promoters and five CpG islands were predicted in the 2 kb upstream of CDKN2A in Wenshang Barred chicken.The promoter region contained a total of 12 transcription factor binding sites,including Sp1,MEB-1,YY1,C/EBPα,AP-2α.The protein encoded by this gene was a positively charged,unstable hydrophilic protein with a molecular mass of 7.30 ku and a theoretical isoelectric point of 12.82.The predictive subcellular localization of the protein was mainly in mitochondria.The protein had no signal peptide regions and signal peptide shear sites.It had eight potential phosphorylation sites and no glycosylation sites.The protein contained a typical TRP_2 (transient receptor potential ion channelⅡ) conserved domain and its secondary and tertiary structures were mainly α-helices and random coil.In addition,it had interactions with MDM2,NPM1,USP36 proteins.The expressions of CDKN2A gene in the back skins increased from 1 to 35 days old in Wenshang Barred chicken.

To investigate the characterization and function of LbaHY5 gene of Lycium barbarum,explore the role of LbaHY5 in the degree of stem straightening in Lycium barbarum.Using Ningqi No.1 as the test material,a preliminary analysis was conducted on the structure and function of the LbaHY5 gene using bioinformatics,subcellular localization,and qRT-PCR methods,the results indicated that the LbaHY5 gene cloned from Ningqi No.1 had a full-length open reading frame of 483 bp,encoding 160 amino acids,with a molecular weight of 17.52 ku and an isoelectric point of 9.69,classifying as a hydrophilic protein.Conserved domain and multiple sequence alignment analysis showed that LbaHY5 protein contained a type bZIP domain,which belonged to the bZIP gene family.Evolutionary tree analysis showed that the LbaHY5 protein in goji berries was highly homologous to the HY5 protein in Solanaceae plants such as tomatoes and potatoes.In addition,promoter analysis showed that the promoter region of this gene was rich in multiple functional elements,which were respectively related to processes such as light signaling,hormone signaling pathways,and abiotic stress response.The subcellular localization indicated that LbaHY5 was located on the nucleus.The transcript level of LbaHY5 gene was the highest in the flower and the lowest in the stem.During the fruit development period,the expression level of LbaHY5 gradually increased.The results speculated that LbaHY5 gene might play an important role in the growth and development of floral organs and fruits in Lycium barbarum.