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.
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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To investigate the sequence characteristics of uridine diphosphate glycosyltransferase in different flower color of Meconopsis and its relationship with the flower color of Meconopsis.The yellow-flowered Meconopsis integrifolia and the red-flowered M.punicea were used as study materials,and one UGT gene was screened from the full-length transcriptome data of each of the two flower colors of Meconopsis.Based on the results of KEGG annotations of the transcriptomes,these two genes were named MiUGT79B1 and MpUGT79B1,and they were cloned,and analyzed by bioinformatic and RT-qPCR.The results showed that the ORF length of MiUGT79B1 and MpUGT79B1 genes was 1 314,encoding 437 amino acids.Both MiUGT79B1 and MpUGT79B1 proteins were hydrophilic stabilizing proteins with their structures consisting mainly of α-helices and random coil.Phylogenetic tree and multiple sequence comparison analyses revealed that MiUGT79B1 and MpUGT79B1 were most closely related to Papaver somniferum,P.californicum, P.nudicaule,and Macleaya cordata.RT-qPCR analysis showed that MiUGT79B1 and MpUGT79B1 were highly expressed in the bud stage,MiUGT79B1 expression tended to decrease first and then increase,and MpUGT79B1 expression tended to decrease first and then remain stable.There was a significant positive correlation between the expression level of UGT gene and the accumulation pattern of total flavonoids.The results of this study provide a theoretical basis for further in-depth research on the function of MiUGT79B1 and MpUGT79B1 genes in the petal coloration of Meconopsis and its molecular breeding of flower color.
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Magnolia wufengensis is a rare ornamental tree species endemic to China.Its flower color variation types are extremely rich.There are a series of flower color variation varieties such as deep red,pink and white.However,the molecular mechanism of flower color regulation is still unclear.In order to explore the molecular mechanism of flower color regulation of Magnolia wufengensis,MawuCPC,a key candidate gene for flower color regulation of Magnolia wufengensis,was cloned from Magnolia wufengensis.Phylogenetic analysis,amino acid sequence structure analysis,subcellular localization experiment,yeast two-hybrid experiment and luciferase experiment were carried out.The expression level of MawuCPC in different flower color varieties was compared and analyzed by Real-time Quantitative PCR.The results showed that the coding region of MawuCPC gene was 258 bp,encoding 85 amino acids.Phylogenetic analysis showed that MawuCPC was clustered with CPC members of Arabidopsis thaliana and other species in the same evolutionary branch,which further supported that MawuCPC belonged to the CPC family of Magnolia wufengensis.Sequence structure analysis further showed that MawuCPC protein had only a conserved R3 domain and a bHLH binding motif.The result of subcellular localization experiment showed that MawuCPC protein was specifically localized in the nucleus.Yeast two-hybrid and luciferase assays showed that MawuCPC and MawuPAP1 could interact with the bHLH family protein MawubHLH1 of Magnolia wufengensis,and the binding of MawuCPC to MawubHLH1 could seriously inhibit the enhancement expression of MawuPAP1 and MawubHLH1 on the promoter of the structural gene MawuDFR gene.Real-time Quantitative PCR results showed that the expression level of MawuCPC was the highest in the white flower variety JB,followed by the pink flower variety Jiaoyan,and the lowest in the red flower variety Jiaohong 1.The above experimental results indicate that in Magnolia wufengensis,MawuCPC may also play its role in flower color regulation by competing with PAP1/2 protein to bind bHLH protein,and maybe the difference in the expression level of this gene promotes Magnolia wufengensis to produce extremely rich flower color variation types.
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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.
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In order to investigate how to alleviate the effects of insufficient light in potato production,the potato variety Dongnong 310 was used as the test material,and three shading treatments were set up:normal light(Z0,blackout rate is 0),single-layer blackout screen covering treatment(Z1),double-layer blackout screen covering treatment(Z2).Different concentrations of epbrassinolide(0 mg/L,denoted as CK)were sprayed in the potato tuber formation stage(0.02 mg/L,denoted as E1;0.10 mg/L,labeled E2).Three measuring periods were set,the plant height,chlorophyll a,b content,photosynthetic parameters,fluorescence kinetic parameters,and yield per plant were measured and analyzed.The relative expression levels of LHcb genes in 12 leaves were measured.The results showed that Z1 conditions were more suitable for the growth of Dongnong 310,while Z0 conditions produced photoinhibition,and the indexes were lower than Z1.Under the condition of Z2,the illumination was insufficient,and the indexes were lower than that of Z1.EBR spray could effectively increase the yield per plant,and E2 treatment under Z0 condition significantly increased by 32.62% compared with CK,E1 and E2 treatment under Z1 condition significantly increased by 48.38% and 24.24% compared with CK,respectively,E2 treatment under Z2 condition significantly increased by 18.45% compared with CK.In terms of weight of dry matter per tuber,E2 treatment under Z0 condition increased by 27.24% compared with CK,and E1 and E2 treatment under Z1 condition increased by 40.09% and 26.56% compared with CK,and the above differences were significant.The relative expression of 12 LHcb genes could be increased by increasing the shading degree and spraying EBR,which was conducive to improving the light energy capture and utilization in leaves.
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In order to explore the molecular mechanism of low nitrogen response in quinoa, low nitrogen response genes were screened to reveal the adaptive changes in quinoa response to low nitrogen.Based on the seedling growth observation and chlorophyll synthesis detection,we analyzed the transcriptome changes of quinoa after 5 d and 30 d under nitrogen deficiency conditions.The results showed that roots were preferentially developed under nitrogen starvation condition.Older leaves turned yellow or dropped down under both low nitrogen and nitrogen starvation conditions,therefore younger leaves could maintain green.Higher NUE was shown in both low and nitrogen starvation conditions.GO enrichment analysis indicated that significantly differential expressed genes were mainly involved in integral component of membrane,membrane,oxidation-reduction process,metabolic process,ATP binding,and metal ion binding.After 5 d of low or nitrogen starvation supply,KEGG enrichment analysis showed that phenylpropyl biosynthesis and glutathione metabolism were the most significant metabolic pathways compared with high nitrogen.After 30 d of treatment,the most significant metabolic pathway was the carbon metabolic pathway.The key genes in response to low nitrogen in quinoa were further explored.The results showed that peroxidase,glutathione S-transferase and ascorbate peroxidase genes were up-regulated and their expressions were higher after 5 days of low nitrogen and nitrogen deficiency treatment.The genes of phosphoglycerate kinase,cysteine synthetase,glyceraldehyde 3-phosphate dehydrogenase (NADP)and phosphoenolpyruvate carboxylase were up-regulated and the expression levels were higher after 30 days of low and low nitrogen treatment.The results of qRT-PCR agreed with the RNA-Seq.
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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.
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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.
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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.
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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.
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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 F2:3 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 F2:3 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.
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To explore the transcriptome changes after biocontrol bacteria treat Camellia oleifera in different ways,and understand the molecular regulatory mechanism of biocontrol bacteria enhancing C.oleifera resistance.The 2-year-old C.oleifera seedlings were selected as experimental materials,and the biocontrol bacterium Bacillus tequilensis was inoculated onto seedlings by leaf spraying and root irrigation,as well as an equal amount of sterile water was used as a control,and the C.oleifera leaves were taken after 15 days of treatment for transcriptome sequencing analysis.The results indicated that a total of 167 118 913 reads were obtained by transcriptome sequencing,with 19 164 differentially expressed genes(DEGs).Among which there were 4 722 DEGs in sample CK-vs-DZY 6715 and 14 442 DEGs in sample CKG-vs-DZY 6715G,and the number of DEGs common to the two groups of samples was 2 573,with the highest number of unique DEGs to sample CKG-vs-DZY 6715G(11 869),followed by sample CK-vs-DZY 6715(2 149).GO enrichment analysis showed that the DEGs of the two groups of samples were annotated into biological processes,molecular functions,and cellular components,mainly involving metabolic process,cellular process,single biological process,cell,cell part,membrane,binding,catalytic activity,transport activity,etc.KEGG pathway analysis showed that the DEGs of the two groups of samples were enriched into metabolic pathway and biosynthesis of secondary metabolites,while DEGs were also mainly enriched to plant hormone signaling transduction in sample CK-vs-DZY 6715,and the DEGs of sample CKG-vs-DZY 6715G were mainly enriched in the plant pathogen interaction.The main DEGs related to disease resistance were up-regulated expression,including pckA,PGAM,ACOX1,FATB,asnB,ansA,FLS,LAR,AUX1,GID1,ETR,STPK,CDPK,pfkA.However,there were differences in gene expression levels between spray and root irrigation methods,the expression levels of genes related to the biosynthesis of secondary metabolites and plant hormone signal transduction were higher in spray,while those related to metabolic pathway and plant pathogen interactions were higher in root irrigation.Thus,this paper concluded that the biocontrol bacteria Bacillus tequilensis could enhance disease resistance in C.oleifera after treatment with leaf spray or root irrigation,only that the main mechanism of action of the two treatment methods was slightly different.
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A lysozyme gene,MseLYS,was cloned from oriental armyworm,and then,its prokaryotic expression vector was constructed by the prokaryotic expression system,and finally,the expression pattern of this gene was detected in different stages and tissues of oriental armyworm,to provide theoretical reference for exploring the function and structure of the MseLYS and lay a foundation for further study of the antibacterial function and physiological mechanism.We cloned the sequence of the lysozyme gene MseLYS from the midgut of Mythimna separata by reverse transcription PCR(RT-PCR)and rapid amplification of cDNA ends(RACE).The open reading frame(ORF)sequence with the signal peptide removed was ligated to the expression vector pET-30a(+),and the inducible expression was carried out by IPTG.Quantitative PCR was used to detect the spatiotemporal expression pattern of this gene.Sequence analysis showed that the full length cDNA of MseLYS was 729 bp,and its ORF was 426 bp,encoding a total of 141 amino acid residues.Meanwhile,the 5'non-coding region and 3'non-coding region included 75,228 bp,respectively.The isoelectric point and molecular weight of the protein encoded by MseLYS were 7.72 and 16.13 ku,respectively.Phylogenetic tree demonstrated that MseLYS was clustered with other species' C-type lysozyme,and was highly consistent with other insect amino acid sequences,suggesting that MseLYS was a C-type lysozyme.SDS-PAGE showed that the expressed protein was consistent with the expected size,which was about 20 ku,indicating that MseLYS can be expressed efficiently in BL21(DE3).Instar expression profiling analysis illustrated that there were significant differences in the expression levels of MseLYS gene among different developmental stages of larvae,males,and females.The expression levels of MseLYS gene were higher in the late larval and pupal stages of oriental armyworm,while the expression levels were lower in other developmental stages.The tissue expression analysis results indicated that there were significant differences in the expression levels of this gene among different tissues of male adults,with higher expression levels in the fat body and thorax;this gene expression also showed significant differences among different tissues of female adults,with higher expression levels in antennae,wings,and cuticle.To sum up,the full-length sequence of MseLYS is cloned,its prokaryotic expression vector is successfully constructed,which could efficiently express the target protein,and its expression pattern is clarified in different tissues and ages.
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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.
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Epidermal growth factor receptor(EGFR)is excessively expressed in numerous solid tumours,establishing a stable over-expressing canine EGFR protein MDCK cell line will prove beneficial in providing an exceptional cellular model and research foundation for the study of canine or human tumour diseases.Firstly,the target fragment was prepared for construction of an overexpression vector of the EGFR gene.It was co-transfected with a lentivirus packaging vector into 293T cells.After 48 hours of transfection,the supernatant was extracted and purified to obtain a lentivirus containing the overexpressed EGFR gene.This virus was then transduced into MDCK cells,following puromycin selection and single-cell cloning.The infectivity efficiency was observed,expressing gene and protein levels were detected through RT-qPCR,Western Blot,and indirect immunofluorescence assays,and an EGFR protein overexpression MDCK cell line was established.Using flow cytometry,apoptosis and cell cycle of overexpression and control cell lines were separately determined.The canine EGFR gene was located on chromosome 18 and consisted of 30 exons encoding a transmembrane glycoprotein.Through the reaction system,PCR products were ligated into linearized expression vectors,generating 8 positive transformants with a size of 738 bp.The obtained lentivirus titer of EGFR gene overexpression was 3.5×108 TU/mL.Transfected MDCK cells exhibited superior fluorescent effect was remarkable under fluorescent microscopy,resulting in a significant increase in the expression level of the EGFR gene and its protein within the cells.Indirect immunofluorescence assay revealed a substantial enhancement in the expression of EGFR in cells.In contrast,early apoptosis and late apoptosis rates among the transfected cells increased notably,with a clear arrest occurring at the G2/M phase.This research successfully developed a stable canine EGFR overexpressing MDCK cell line.Overexpression of canine EGFR stimulated cell division,elevated DNA replication,and concurrently triggered programmed cell death.
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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.
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Bimonthly, Started in 1962
CN 13-1101/S
ISSN 1000-7091
CODEN: HHUOA6
Responsible Institution: Hebei Academy of Agriculture and Forestry Sciences
Sponsored by: the Academy of Agricultural Sciences and Agricultural Association of Hebei, Beijing, Tianjin, Shanxi, Henan and Inner Mongolia.
Editor-in-chief: Qiang Zhang
Edited and Published by: Editorial Department of Acta Agriculurae Boreali-Sinica
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