In order to preliminarily explore the key regulatory networks and genes involved in maize cold resistance,identify the key regulatory pathways and genes in response to low temperature stress,which laying a foundation for further research on the molecular mechanism of cold stress resistance.Here,a cold tolerance variety Mintian 6855 was employed to determine the gene expression pattern at 24,48 and 72 hours post low temperature of 5 ℃ stress by using transcriptome technique.The PCA analysis revealed that the repeated samples were well clustered together and significantly separated from CK samples.The results of difference analysis showed that about 4 000—7 000 difference genes expressed after cold stress treatments,while,only about 100—2 000 showed difference expressing among low temperature treated samples,indicating that low temperature was the main factor results in genes difference expressing,and the difference expression genes were mainly responded in the early stage.Meanwhile,KEGG annotation analysis results revealed that the differentially expressed genes were enriched in pathways of plant hormone signal transduction and MAPK,suggesting that these two signaling pathways actively respond to cold stress.In addition,different express genes were also enriched in plant-pathogen interaction as well as circadian rhythm plant,strongly implied that there were overlapping or common regulatory pathways in biological and abiotic stress pathways,while,genes that regulate circadian rhythms also playing a key role in plant adaptation to low temperatures.

This study intended to investigate the dynamic shoot characteristics and differences of several super rice cultivars and provide theoretical guidance for super rice breeding and cultivation in the South China rice area.Four super hybrid rice combinations,two super conventional rice varieties,and two high-quality super-high-yielding traditional rice were selected as test materials and applied.We investigated the plant leaf morphology,dry matter weight per unit area,yield at the seedling stage,active tillering stage,secondary panicle branch differentiation stage,initial heading stage and mature stage.The results showed that five dynamic shoot indexes,such as plant height,average number of tillers per plant,leaf morphology,dry matter weight per unit area and yield components,had significant seasonal ecological characteristics.Moreover,except the leave tillering angles from the early and middle stages of growth and development,other key indicators displayed that the super hybrid rice combination was significantly higher than the super conventional rice varieties.In details,the number of effective spikes per unit area,total grains per spike,thousand grains weight,economic coefficient and yield per unit area were significantly higher in the hybrid rice combinations than in the conventional rice varieties;while the spike length and seed setting rate of the conventional rice varieties were extremely significantly higher than those of the hybrid rice combinations.So,the construction of wide adaptation super rice breeding demands early and fast seedling development,high biological yield at mid and late growth stage,and high harvest index at maturity.Meanwhile,conventional rice varieties need to cultivate early and fast development characteristics,improve biological yield,and increase thousand grains weight and total number of grains per spike appropriately as well as maintaining high seed setting rate.Hybrid rice combinations requires to improve seed setting rate.

In order to explore the effect of low temperature during the flowering period in Brassica napus,Brassica napus L.GZhui(strong resistance to winter cold)and 10B(weak resistance to winter cold)were used as test materials,low-temperature stress (14 h, 12 ℃ at daytime; 10 h, 2 ℃ at nighttime) to these materials for 1, 2, 3, 4, 5 d, and normal environment(14 h,22 ℃ at daytime;10 h,18 ℃ at nighttime)as control,determine the changes of physiological indexes in bolting stem leaves and lower leaves under low-temperature stress,as well as pollen viability and stigma receptivity after the opening of flower buds of different length.The research results showed that the leaves of the two materials were slightly wilted after low-temperature treatment,and there was no obvious damage to the shape of the plants in all treatments,the changes of various physiological indexes of leaves after low-temperature treatment were more complicated,and the three antioxidant enzymes were sensitive to peroxidase(POD),and most of the antioxidant was significantly increased,lower leaves more than bolting stem leaves.The content of soluble sugar(SS)in osmotic adjustment substances changed significantly,GZhui increased significantly.Malondialdehyde(MDA)content increased significantly,10B more than GZhui,and the lower leaves and bolting stem leaves showed different performances.For flower buds larger than 6.0 mm,the pollen viability of the two materials was little affected by low-temperature treatment for 4 days.For less than 3.0 mm of flower buds,the development stopped and eventually died for more than 4 days of cold stress,and the pollen viability decreased significantly after 2—3 days of stress.After the flower buds of all levels,less than 6.0 mm were under low-temperature stress,the pollen viability of GZhui was higher than that of 10B,and the difference was obvious with the flower buds of 3.0—6.0 mm,so it was considered that the 3.0—6.0 mm flower buds could be used as an indicator for identifying different varieties of low-temperature tolerance during flowering.The performance of stigma receptivity was consistent with the trend of pollen vigor.The stigma receptivity of flower buds larger than 3.0 mm was not affected within 3 days of low-temperature treatment,and the receptivity of stigma more than 4 days of treatment was reduced;the flower buds smaller than 3.0 mm,the receptivity of stigma decreased to varying degrees within 3 days of low-temperature treatment.These results indicated that flower buds(smaller than 3.0 mm)were more sensitive to low temperature,resulting in reduced pollen vitality and stigma receptivity,and even abortion.

Protein kinases are important factors in plant defense system,protein kinase SnRK2 is a serine/threonine protein kinase,can play an important role in the plant stress signal transduction pathway through phosphorylation.We analyzed the expression pattern of SnRK2.4 under abiotic stresses,aiming to reveal its role in the regulation of adversity.The SnRK2.4 gene of high-quality cold-season turfgrass Poa pratensis L. was cloned using RT-PCR,the SnRK2.4 gene contained an ORF of 1 092 bp encoding a 363-amino acid protein,and its molecular characteristics were analyzed by bioinformatics.In addition,the expression pattern of this gene in different tissue parts under different abiotic stresses was observed using qPCR.The results showed that the Poa pratensis L.SnRK2.4 gene belonged to the SRK/SAPK superfamily,contained typical STKc_SnRK2 domain,tyrosine kinase phosphorylation site,casein kinase Ⅱ phosphorylation site and serine/threonine-protein kinase activity site,which had the highest homology with Brachypodium distachyon.The qPCR results demonstrated that the Poa pratensis L.SnRK2.4 gene was tissue-specific,highly expressed in panicles,and had no significant differential expression in roots,stems and leaves.Moreover,the Poa pratensis L.SnRK2.4 gene could respond positively to abiotic stresses such as drought,salt,low nitrogen,low phosphorus,ABA and BR.

In order to study the effects of straw returning on soil active organic carbon components and carbon cycle related enzyme activities in rice-rape rotation farmland of Chengdu Plain,we carried out 3-year straw returning field experiment(2017—2020),including no straw(CK),chemical fertilizer alone(NPK),50% straw returning and chemical fertilizer(SR1),100% straw returning and chemical fertilizer(SR2),200% straw returning and chemical fertilizer(SR3).We measured soil physical-chemical properties,soil organic carbon content(SOC),soil labile organic C fractions,carbon cycle related enzymes,and their correlations.The results showed that straw returning could effectively improve soil physical-chemical properties,soil available nitrogen,phosphorus and potassium contents.Compared with CK treatment,straw returning treatments significantly increased SOC,ROC,DOC,and MBC contents by 5.05%—8.55%,18.40%—36.80%,35.76%—66.93% and 27.20%—52.10%,respectively.In general,higher returning dosage resulted in higher C content.On the other side,compared with CK and NPK,straw returning treatments significantly increased soil cellulase,β-glucosidase,catalase,polyphenol oxidase.The activities of soil cellulase,β-glucosidase,and polyphenol oxidase under SR2 treatment were the highest,which were significantly higher than SR1 treatment by 16.25%,8.49%,and 14.69%,respectively.The catalase activity of SR3 treatment was the highest,which was significantly higher than that of SR1 treatment by 25.10%(P<0.05).There were significant positive linear correlations among soil SOC,labile SOC fractions,and carbon cycle related enzyme activities.Consequently,full straw returning has been proved of the most efficient way of improving active organic carbon components,carbon cycle related enzyme activities,and promoting the improvement of soil quality in rice-rape rotation farmland of Chengdu Plain.

In order to improve the blast resistance of Shuijing 3,an excellent food-flavor rice variety,CRISPR/Cas9 gene editing technology combined with gene chip technology were used to pyramid the R gene Pigm and the non-R gene bsr-d1 into Shuijing 3.Firstly,Bsr-d1 was selected as the target gene to construct a recombinant expression vector using the CRISPR/Cas9 gene editing system,and transformed into the excellent food-flavor rice Shuijing 3 by Agrobacterium-mediated method.The homozygous bsr-d1 mutant lines without T-DNA elements,including five mutation types as T insertion,G insertion,GA deletion,CGCA deletion and CGCAGA deletion,were screened out.The japonica line Jinyu 1 containing a broad-spectrum blast resistance gene Pigm was used as the gene donor parent to cross with the homozygous bsr-d1 mutant lines without transgenic components.The Pigm gene was introduced into bsr-d1 mutant lines by cross,backcross and self-cross combing molecular breeding chip to simultaneously perform Pigm gene and background-assisted selection.The improved lines SJ3-G1,SJ3-G2,SJ3-G3,SJ3-G4,SJ3-G5,which were homozygous for the disease resistance genes(carrying both bsr-d1 and Pigm genes)and whose background recovery rates were all above 96%,were finally obtained.The improved strains of Shuijing 3 displayed enhanced leaf blast resistance compared with the wild type in inoculated identification test using Magnaporthe grisea strain GUY11.After inoculation with M.oryzae,the POD activities in the improved strains of Shuijing 3 were significantly lower than that of the wild-type control,while the H₂O₂ contents were significantly higher than that of the wild-type control.The improved Shuijing 3 lines with blast resistance carrying both bsr-d1 and Pigm genes are obtained by CRISPR/Cas9 gene editing technology combined with gene chip technology.

In order to explore the role of melatonin in the interaction between wheat and leaf rust,we studied the incompatible combination between wheat variety Lovrin 10(hereinafter referred to as L10)and physiological race 260 of leaf rust. In this experiment,reactive oxygen species were induced by exogenous injection of Methyl viologen (Methy lviologen(MV)as an oxidant induces the production of superoxide anion,which can effectively increase the content of reactive oxygen species),and the best concentration of melatonin was determined by using the ability of melatonin to scavenge reactive oxygen species;then,melatonin was injected into the leaves of wheat seedlings and inoculated with leaf rust race 260,and the changes of H₂O₂ content were observed by DAB staining. The HR area was detected by Rohringer staining. The effects of exogenous melatonin injection on the antioxidant capacity of wheat were investigated by measuring the peroxidase(POD)and catalase(CAT)activities of wheat;through the above research,the role of melatonin in the interaction between wheat and leaf rust was clarified. The results showed that the reactive oxygen species was increased by exogenous injection of methyl viologen,and the optimum injection concentration of melatonin was 10 μmol/L. The results of DAB staining of incompatible combinations showed that after injection of 10 μmol/L melatonin,the accumulation of H₂O₂ induced by leaf rust infection in wheat leaves was less than that in control group,which indicated that melatonin participated in H₂O₂ scavenging. Rohringer staining showed that the area of wheat HR cells decreased after exogenous melatonin treatment,which effectively enhanced the resistance to leaf rust. In addition,exogenous injection of melatonin increased the activities of antioxidant enzymes POD and CAT,which indicated that the antioxidant capacity of wheat was significantly improved. The results showed that exogenous injection of melatonin was involved in the removal of reactive oxygen species in the process of wheat resistance to leaf rust,and improved the disease resistance of wheat.

The Pineal-hypothalamus-pituitary-thyroid axis(PHPTA)could significantly regulate reproductive activities of animals.In order to investigate the expression of Kiss1/GPR54 system on PHPT axis and its regulation on reproductive activities of Tibetan sheep,24 healthy and non-pregnant Tibetan sheep in estrus cycle were selected as experimental group and 6 non-breeding season Tibetan sheep as control group.The dynamic changes of plasma Kisspeptin were detected by ELISA,and the mRNA and protein expression of Kiss1 and GPR54 in optic nerve,pineal body,hypothalamus,pituitary body and thyroid gland were detected by Real-time quantitative PCR,Western Blot and immunohistochemistry.The results showed that the plasma Kisspeptin secretion in late estrus was significantly higher than that in other periods,and that in each period of breeding season was significantly higher than that in non-breeding season.The mRNA and protein of Kiss1 and GPR54 were expressed in optic nerve and PHPT axis.The relative expression of mRNA and protein of Kiss1 and GPR54 in optic nerve were significantly higher in early estrus than in other periods(P<0.05).The mRNA and its protein of Kiss1 of the pineal body reached its maximum value in estrus,significantly higher than that in reproductive cycle.The mRNA relative expression of Kiss1 and GPR54 of hypothalamus and thyroid gland were significantly increased in late estrus(P<0.05).The mRNA relative expression levels of the two in pituitary body were significantly higher in estrus than in other periods.Immunohistochemical results showed that Kisspeptin and GPR54 were mainly distributed in the glial nucleus and cytoplasm,respectively.Both were strongly positive in pineal cell cytoplasm.Kisspeptin was expressed in neuroendocrine small cells and glial cells in hypothalamus,GPR54 was expressed in cytoplasm of neuroendocrine small cells.Kisspeptin and GPR54 were mainly expressed in the cytoplasm of basophils in pituitary body.In thyroid gland,they were mainly expressed in the cytoplasm of follicular cells.The dynamic changes of plasma Kisspeptin and the differential expressions of Kiss1 and GPR54 in different tissues during estrus cycle of Ganjia Tibetan sheep indicated that Kiss1/GPR54 system was involved in regulating reproductive physiological activities of Ganjia Tibetan sheep.

This study aims to use high-throughput sequencing technology to perform mRNA sequencing and differential analysis of longissimus dorsi muscle tissue samples from Songliao black pigs and Landrace pigs,and screen out key genes that affect pig muscle growth,meat quality and fat deposition,so as to provide pork quality research provides new reference information.The longissimus dorsi muscle tissue samples of 6 Songliao black pigs and 6 Landrace pigs were collected,their RNA was extracted,and the mRNA was sequenced by Illumina HiSeq 2500 high-throughput sequencing technology,and the obtained reads were compared,annotated and differentiated.For expression analysis,NOISeq was used to screen out differentially expressed genes and perform enrichment analysis of related biological functions.The results showed that 664 differentially expressed genes were screened from the two pig species,of which 364 genes were highly expressed in Songliao black pigs and 300 genes were highly expressed in Landrace pigs.Through the biological function analysis of differentially expressed genes,LPIN1,FADS1,FADS2,PLIN2,PPARGC1A,PRKAG2 and ACSL1 were screened to participate in the regulation of lipid metabolism and muscle development.The related pathways were fatty acid metabolism,PPAR signaling pathway,AMPK signaling pathway,insulin signaling pathway and adipocytokine signaling pathway and so on.

NRL(NPH3/RPT2-Like)is a type of light-responsive protein unique to plants and plays a vital role in the phototropic signal pathway. To reveal the NRL gene maize genome's characteristics and expression,we analyzed them using bioinformatics methods combined with qRT-PCR technology. The property,structure,evolution of their encoded proteins,and growth period tissue expression and stress expression were analyzed. 31 ZmNRL genes identified were located in nine maize chromosomes,encoding protein amino acids 464-749 aa,which predicted to have chloroplast,nuclear and cytoplasmic locations. According to protein conservation,ZmNRL family was divided into four categories. Their gene structure also presented certain conservation,the most contained four exons. Analysis of the cis-elements of gene promoters revealed a large number of abscisic acids,jasmonic acid,light response,and anti-oxidation elements,among which G-box and Sp1 were two types of light-related elements. The expression of ZmNRL family genes in tissues during the growth period showed a temporal and spatial specificity,and the majority expression level was not high. Only ZmNRL2,ZmNRL4,ZmNRL24,and ZmNRL29 highly expressed. Furthermore,the characteristic modules were produced based on the data of the tissue co-expression genes. And the GO enrichment analysis of a particular leaf growth module containing six ZmNRL genes,mainly associated with the plastid organization biological processes and rRNA binding molecular functions. The expression of ZmNRL5,ZmNRL7,ZmNRL12,and ZmNRL19 genes were analyzed by qRT-PCR under salt,drought,high temperature,and Rhizoctonia solani inoculation treatments. The results showed that ZmNRL12 was significantly up-regulated in maize seedlings treated with high temperature,while ZmNRL5,ZmNRL7 and ZmNRL19 genes were down-regulated in drought,salt and pathogen treatments. In summary,31 ZmNRL genes were identified in the maize genome. They not only had apparent specific tissue expression but also participated in biotic and abiotic stress responses.

The SiPRR73 gene was cloned from Yangu 11 using RT-PCR technology,and through analyzing tissue-specific expression,responsive features of SiPRR73 to different photoperiods,photo-thermal combinational treatments and five abiotic stress treatments,the regulation mode of photoperiod and temperature on SiPRR73,and the responsive pattern of SiPRR73 to abiotic stresses in foxtail millet were explored. The results showed that totally 2 928 bp cDNA sequence of SiPRR73 was obtained from Yangu 11,which included 2 283 bp CDS region,encoding 760 amino acids. The SiPRR73 proteins of C4 crops including Panicum miliaceum,Panicum hallii,Sorghum bicolor and Zea mays showed relatively close relationship with SiPRR73. The second parietal leaf was the highest expression tissue of SiPRR73,but the expression level at root,stem and panicle tissues was relatively lower. The expression level of SiPRR73 was higher at light period than that at dark period under both short-day and long-day conditions,and during the whole vegetative growth phase,SiPRR73 showed higher expression level under long-day compared to short-day,which indicated that the expression of SiPRR73 was induced by light and controlled by photoperiod. The temperature determined expression peak number of SiPRR73 and the photoperiod determined occurrence time of expression peaks,so temperature and photoperiod participated in regulating of SiPRR73 expression mutually. PEG and low temperature stresses induced SiPRR73 expression totally,NaCl induced SiPRR73 expression at early stress stage,but inhibited it at later stress stage. Fe stress inhibited SiPRR73 expression at early stage,but induced it at later stage. ABA stress caused the close responsive feature of SiPRR73 to NaCl. This study indicated that SiPRR73 showed light-dependent expression feature,and photoperiod and temperature regulated SiPRR73 by interaction pattern,suggesting that SiPRR73 participated in adaptability regulation process to different photo-thermal conditions and might play a certain role in coping with drought,low temperature,ABA,NaCl and Fe stresses in foxtail millet.

The objectives of this study were to investigate the effects of nitrogen application rate on the grain yield,nitrogen accumulation and use efficiency of spring maize under different film mulching methods,and to provide a theoretical basis for efficient nitrogen application management of spring maize film mulching in high altitude area of Guizhou.Field trials were carried out from 2018 to 2019.The plots were arranged in a split-plot design.The main plots were assigned to the film mulching methods(wide film and narrow film) to be studied and divided into subplots according to the five nitrogen application levels(0,80,160,240 and 320 kg/ha).To study the effects of different film mulching methods and nitrogen application rates on yield,nitrogen accumulation,transport characteristics and utilization efficiency in different organs of spring maize.Film mulching,nitrogen application rate and their interaction significantly increased the yield of spring maize.The results showed that compared to narrow film mulching,wide film mulching not only increased the yield of spring maize by 17.8%,but also significantly enhanced the nitrogen accumulation and the retransfer of nitrogen accumulated before silking,then significantly increased the grain nitrogen accumulation.Moreover,compared to narrow film mulching,wide film mulching achieved significantly higher NUTE,NUPE,AEN,NPFP and NUE values showing increases of 4.9%,21.4%,23.5%,12.2% and 4.23 percentage points,respectively.Nitrogen application realized the synergistic growth of grain yield and plant nitrogen accumulation of spring maize,and could significantly affect nitrogen absorption,accumulation and transport.As the N application rate increased,the N translocation and the contribution rate of nitrogen transfer from vegetative parts to grain after silking could be effectively promoted,as well as enhanced.However,the nitrogen use efficiency,including the NDGPE,NHI,NUTE,NUPE,NUE,AEN,NPFP of spring maize,significantly decreased by increasing of N application rate.The regression analysis had shown that the optimal yield and nitrogen application rate for wide film mulching were reduced nitrogen application by 55 kg/ha and increased yield by 12.3% compared with narrow film mulching.The combination of wide film mulching and suitable nitrogen application rate was beneficial to the plant accumulation and absorption of nitrogen,so as to achieve high yield and nitrogen productivity,and then achieve the purpose of saving fertilizer and increasing yield.Considering the grain yield,N accumulation,transport and N use efficiency of spring maize,the reasonable N application rate of wide film mulching for spring maize in high altitude and other similar ecological zones in Guizhou was 160 kg/ha,which yield could reach 11 404.3 kg/ha.