Being effective only to the traits controlled by large-effect QTL,conventional marker-assisted selection(MAS)can hardly improve the quantitative traits that are controlled by many small-effect QTL.Later on, the genomic selection (GS) technology proposed in 2001 solves the problem of improving complex traits controlled by the minor polygenic effects due to its high prediction accuracy through estimating the breeding value of individual with high-density molecular markers. At present,it has been successfully applied to the genetic improvement of quantitative traits in animals and plants such as dairy cattle,pigs,sheep,maize,and wheat in the US,Canada,Australia,Germany,France and so on.As a breaking-through breeding technology and a hot spot in research and application now. In this review,we summarized the factors affecting predictive accuracy of GS and the research progress of GS in breeding maize,wheat,rice and rapeseed at home and abroad,and finally discussed the existing problems in its application. This review will provide some reference for GS of the crops in the future.

Wheat yellow dwarf disease,caused by Barley yellow dwarf viruses(BYDVs),is an important viral disease in wheat production.BYDV GAV has become the main pathogen causing wheat yellow dwarf disease.Till now the studies on the function of BYDV GAV encoded proteins P1,P2 and CP are lacking.We focued on the function of P1,P2 and CP,which could lay the foundation for the pathogenic mechanism of BYDV GAV.Phylogenetic analysis of BYDV GAV encoded P1,P2 and CP was conducted using Mega 7.0.We constructed the YFP expression vector of P1,P2 and CP,and then transformed them into GV3101 and infiltrated the leaves of Nicotiana benthamiana.The subcellular localization of P1,P2 and CP was observed using confocal laser scanning microscopy(CLSM).We constructed the biomolecular fluorescence complementation assay(BiFC)vectors of five coding proteins,and then transformed them into GV3101 and infiltrated the leaves of Nicotiana benthamiana.CLSM was used to observe the interaction of P1,P2 and CP and other viral proteins in vivo.Furthermore,we constructed the Potato virus X(PVX) expression vectors of P1,P2 and CP,transformed them into GV3101 and infiltrated the leaves of Nicotiana benthamiana.At 5 days post inoculation(dpi)the symptom formation of PVX infection was observed.The systemic leaves were collected for detection of viral accumulation to determine the effects of the pathogenicity of P1,P2 and CP.Results showed that BYDV GAV was most closely related to BYDV PAV at the nucleotide level.Subcellular localization of P1,P2 and CP was cytoplasm and nuclear.P1 interacted with itself in vivo using BiFC.In the pathogenicity assay,the systemic leaves of PVXCP infection showed chlorosis at 5 dpi,and PVX accumulation was detected,while PVX,PVXP1 and PVXP2 infection showed no symptoms in systemic leaves and PVX accumulation was undetectable,which was detected at 10 dpi,indicating that CP promoted the formation of PVX symptoms.In brief,P1 possibly involves in viral infection via self-interaction in vivo,and CP can promote viral infection.

In order to have a clearer understanding of the research progress on cold resistance of winter rapeseed in cold and arid regions of China,and to provide theoretical support for breeding and production.The research progress of cold-resistance and cold-resistance breeding of winter rapeseed were summarized respectively from six aspects such as selection index and method,genetic law,breeding method,screening effect,germplasm classification and molecular mechanism research,and four aspects such as breeding method,breeding varieties,cold resistance classification and adaptability screening of varieties.Based on the analysis of the existing problems,six measures were put forward,such as creating germplasm resources by multiple ways,screening out core germplasm by comprehensive evaluation,carrying out research on cold-resistance mechanism and construction of innovation system of molecular design breeding,constructing breeding system and carrying out cold-resistance breeding by multiple ways,improving the adaptability of varieties by comprehensive screening,promoting varieties yield level and improving benefits by carrying out matching cultivation techniques,in order to promote the development of rapeseed production in cold and arid regions of our country.

In order to further definite the genetic basis of black rot resistance in cabbage,and breed high-quality disease resistant cultivars,F₁ progenies were obtained by crossing the inbred line 4674(high resistance to race 1)as male parent,with the inbred line 4673(high susceptibility to race 1)as female parent.The F₂ population containing 152 individuals were obtained by F₁ self-crossing. The F₂ population was inoculated by spraying at seedling stage. After 12-14 days, the phenotype of F₂ population was identified according to the identification method of cabbage seedling stage.A total of 175 markers with good polymorphism and clear bands were selected from 404 pairs.Subsequently, the 175 molecular markers were used for genotyping of F₂ population and constructing a genetic linkage map. Finally, the QTL for black rot resistance of cabbage was mapped with phenotypic data and genetic map.The results showed that there were 154 molecular markers linked to nine chromosomes,including 110 pairs of InDel markers and 44 pairs of SSR markers and covering a length of 714.29 cM,and the average distance between markers was 4.64 cM.Seven QTLs were located,of which three major QTLs were qBR-7-2,qBR-7-3 and qBR-4-3,mapped on the genomic markers(CG842110-CG842482 and M29-M39)of chromosome 7,and on the genomic markers between CD838151 and BOE417 of chromosome 4,respectively.The explainable phenotypic variation of QTLs were 16.0%,9.2% and 10.0%,and their LOD values were 5.75,3.20 and 3.47,respectively.

In order to investigate the reasons for the differences in plant height between SSSL-B50 and HJX74,an F₂ population was constructed by backcrossing SSSL-B50 with HJX74 for genetic analysis and gene mapping of the plant.The genetic analysis showed that F₁ plants exhibited tall plant height.And the separation ratio within F₂ was 3:1(χ²=0.18<3.84)between the tall plant individuals and the semi-dwarf ones,which indicated that the tall plant height of SSSL-B50 was a dominant trait.Based on the linkage analysis between the marker genotypes and phenotypes of plant height in the F₂ population,the gene controlling tall plant height was mapped between markers S18 and X161 within the 38.38—39.07 Mb interval of the substituted segment on chromosome 1.Moreover,it was found that the known gene of "Green revolution",SD1,fell to the mapping interval.Through amplification and sequence alignment of SD1 gene in Oryza glumaepatula,SSSL-B50,and HJX74,it showed that a 280 bp deletion in CDS in HJX74 compared with Oryza glumaepatula and SSSL-B50.The qRT-PCR results showed that the expression level of SD1 was significantly higher in the second,third,and fourth sections of SSSL-B50 stem than in HJX74.The comparison of the SD1 gene sequence of Oryza glumaepatula with the previous reported results showed that the base substitutions occurred at two sites (E100G, Q339R) in the encoded amino acid.The results revealed that the tall plant height of SSSL-B50 was controlled by SD1.Furthermore,a new allelic type of SD1, SD1^Glu,was identified in Oryza glumaepatula.

It explored the ratio of organic fertilizer replacing nitrogen fertilizer in the piedmont plain of Hebei Province,in order to provide a basis for reducing the amount and increasing the efficiency of nitrogen in wheat in this area.Field experiments were carried out in Boyuan farm in Yongnian,Hebei Province for two consecutive years,and five organic and inorganic fertilizer combination treatments were set up.The results showed that organic fertilizer instead of 20% and 40% chemical fertilizer could significantly improve the number of grains per spike and yield.Compared with the high nitrogen and saving nitrogen treatment of single chemical fertilizer application,the yield increased by more than 4.0%,and the number of grains per spike increased by 3.6—5.6.Most of the grain quality indexes for organic fertilizer instead of 20% and 40% chemical fertilizer treatment,and saving nitrogen treatment were better,and the stabilization time increased by 2.2—2.7 min,the tensile area increased by 10.5—17.5 cm²,and the maximum tensile resistance increased by 28.0—75.5 EU.Various nitrogen efficiency indicators of treatment for organic fertilizer instead of 20% were higher.The nitrogen fertilizer efficiency,nitrogen utilization efficiency,and nitrogen harvest index increased 109.3%,9.3% and 11.3% respectively compared with high nitrogen treatment,and 6.9%,8.5% and 8.3% respectively compared with the saving nitrogen treatment.When organic fertilizer replaced chemical fertilizer in different proportions,nitrate nitrogen in 0—20 cm soil appeared "surface accumulation",and the content of nitrate nitrogen increased,which was more than 38.5% higher than that of the saving nitrogen treatment.The nitrate nitrogen in 20—40 cm soil was significantly higher for the saving nitrogen treatment and the high nitrogen application treatment.Organic fertilizer instead of 20% nitrogen fertilizer treatment had the best yield and grain quality,significantly improve the nitrate nitrogen content in 0—40 cm soil,improve the nitrogen absorption and utilization of wheat,and finally obtain higher environmental benefits.

Grain specific promoter with high transcription activity can regulate the specific and high-level expression of target genes in plant grains.In order to explore specific promoters of maize grain,the dominant expression gene GRMZM2G006585 was screened from the published data of maize expression profile microarray.The DNA sequence about 2 000 bp upstream of its coding region was cloned and named PZm2G006585.By using the online websites New PLACE and PlantCARE to analyze its promoter elements,we found that it contained multiple grain specific related elements such as E-box and P-box.It was preliminarily considered that the upstream sequence of the cloned coding region was a grain specific promoter from maize.In order to verify its function,the expression vector of GUS gene was constructed and transformed into plants.GUS histochemical staining results of transgenic rice showed that the expression pattern of exogenous genes driven by the promoter was grain specific and embryo dominant.The results of GUS activity detection in T₃ seeds of transgenic Arabidopsis thaliana single copy line showed that the GUS activity driven by PZm2G006585 was 909.52 nmol/(min·mg).The discovery and functional verification of the grain-specific promoter PZm2G006585 can provide candidate promoter resource for specific expression of target genes in maize,rice and other monocotyledons.

To investigate the potential functions of DUF760s in rice growth and development,the genome-wide identification,classification,promoter sequence and expression profile analysis of the DUF760 gene family were performed.This research identified 6 and 8 members of DUF760 gene family in rice and Arabidopsis respectively by bioinformatics analysis.Phylogenetic tree analysis divided these members into two subfamilies,there were also some characteristic differences between the two subfamilies in protein conserved motif and gene structure.Multiple cis-acting elements responding to stresses and phytohormones existed in the promoter regions of rice DUF760 family genes.ABRE(abscisic acid response)element was present in the promoter sequence of all DUF760 family members,the promoter region of OsDUF760-1 possessed 9 abscisic acid(ABA)related response elements.The transcriptional expression level of OsDUF760-1 in rice was significantly down-regulated after ABA treatment,while OsDUF760-3 was significantly up-regulated.The expression change patterns of these two genes in rice after drought stress treatment were consistent with that after ABA treatment,which indicated that these two genes may participate in rice drought stress response through ABA signaling pathway,and play different roles.In addition to strong responses to ABA and drought stress treatments,members of rice DUF760 family also showed relatively strong expression changes in response to JA(Jasmonic acid),low temperature and M.oryzae treatments.

Gymnocypris eckloni is an important cold-water indigenous fish from the upper reaches of the Yellow River.Because of its slow growth and development and late sexual maturity,its population has been declining for many years.MSTN(Muscle Growth Inhibitor) is a factor that negatively regulates muscle growth.This study cloned the MSTN-1 gene of G.eckloni and examined its expression characteristics to provide basic information on the molecular regulation mechanism of growth retardation in G.eckloni.The full-length cDNA sequence of MSTN-1 gene was obtained by PCR,5'-RACE and 3'-RACE.qPCR was used to detect the expression characteristics of MSTN-1 gene in different tissues of G.eckloni and in muscle tissues of different Cyprinidae fishes.The full length of the cDNA sequence of MSTN-1 gene was 2 190 bp,with an open reading frame of 1 128 bp,a 5'UTR of 96 bp,and a 3'UTR of 966 bp,encoding a total of 375 amino acids.MSTN-1 was an unstable hydrophilic secretory protein containing a signal peptide and no transmembrane structure,which was mainly distributed in the nucleus,mitochondria and cytoplasm.The secondary structure of MSTN-1 protein was dominated by random curls,and exists in two TGF-β structural domains:namely,the TGF-β prepeptide region (37—268 aa) and the TGF-β functional region (281—375 aa).MSTN-1 protein contained one the proteolytic site RIRR and nine conserved cysteinee in the TGF-β functional region.Amino acid sequence homology analysis showed that the amino acid sequence of MSTN-1 from G.eckloni had high similarity with other Cyprinidae fishes MSTN-1s and low similarity with mammalian and avian MSTN-1s.Phylogenetic analyses showed that the MSTN-1 of the G.eckloni clustered in the same evolutionary branch with other Cyprinidae fishes.The results of Real-time Fluorescence Quantitative PCR showed that MSTN-1 was expressed in brain,muscle,gill,heart,liver and kidney tissues of the G.eckloni,but the expression was higher in brain and muscle tissues.The gene was expressed in the muscle tissues of different Cyprinidae fishes,with the highest expression in the muscle tissues of the G.przewalskii,followed by the higher expression in the muscle tissues of the G.eckloni and the Cyprinuscaurpio.The full-length cDNA sequence of the MSTN-1 gene of G.eckloni was obtained by cloning,and bioinformatics analysis and qPCR assay were performed.The characteristic differences in the expression of MSTN-1 in the muscle tissues of these indigenous fishes can help to further understand the molecular mechanism of growth retardation in highland fishes.

WRKY transcription factor is a family of transcription factors unique to plants.Studies have demonstrated that WRKY transcription factor played an important role in plant growth and development and in plant response to biological and abiotic stress.In order to reveal the function of tomato WRKY gene,two inbred lines of tomato with high resistance to bacterial wilt Hm 2-2(R)and high susceptibility to bacterial wilt BY 1-2(S)were used as experimental materials based on the preliminary transcriptome data,and a WRKY transcription factor SlWRKY75 gene(Solyc05g015850.3)was cloned.The structure,expression pattern and function of the gene and its encoded protein were analyzed by bioinformatics analysis,multiple alignment of amino acid sequences,phylogenetic tree construction,Real-time Quantitative PCR(qRT-PCR)and virus induced gene silencing(VIGS).The results showed that the full length of the cDNA of this gene was 653 bp,its maximum open reading frame was 519 bp,encoding 172 amino acids,the relative molecular weight of the protein was 19.878 51 ku,the theoretical isoelectric point was 9.32.The protein belonged to the hydrophilic non-secreted protein,and there was no transmembrane structure.Meanwhile,the protein had a highly conserved WRKY domain and a CX₄CX₂₃HXH zinc finger motif,which belonged to the Class Ⅱ family.Phylogenetic tree analysis showed that SlWRKY75 was closely related to Solanum pennellii SpWRKY75 and clustered into a group with other Solanaceae,while it was far related to Hevea brasiliensis HbWRKY75 and Gossypium hirsutum GhWRKY75 and was in different branches in the phylogenetic tree.The results of qRT-PCR analysis showed that the expression of SlWRKY75 gene were tissue-specific and could be induced by Ralstonia solanacearum,salicylic acid and jasmonic acid.VIGS result showed that silencing SlWRKY75 reduced plant resistance to bacterial wilt,indicating that SlWRKY75 positively regulated tomato resistance to bacterial wilt.These results suggested that SlWRKY75 gene played an important role in regulating tomato resistance to bacterial wilt.

In order to study the effects of OsBAK1P,a precursor of Brassinosteroid insensitivity 1-associated receptor kinase 1,on the agronomic characteristics of rice.A target sequence fragment with a CDS fragment size of 651 bp was amplified from the rice panicle cDNA using the Zhonghua 11 japonica rice variety as the material according to the specific primers designed by the gene;the PTCK303-OsBAK1P overexpression vector and the PTCK303-OsBAK1P RNA interference vector were successfully constructed by restriction enzyme digestion and ligation methods;the Agrobacterium tumefaciens EHA105 strain was transformed into the correct expression vector plasmid,and the positive Agrobacterium tumefaciens clones were screened out by using CDS amplification primers;the callus of japonica rice cultivar Zhonghua 11 was infected by Agrobacterium tumefaciens genetic transformation to obtain transgenic plants;finally,compared to Zhonghua 11,two overexpressed and RNA-interfered T₁ transgenic plants with similar phenotypes were selected to measure and analyze the agronomic characteristics such as plant height,panicle length,and leaf angle,changes in root length and coleoptile length at the early stage of germination,and response to brassinolide(BL).The results showed that the plant height of OE-OsBAK1P transgenic rice was dwarfed,the panicle length was shorter,and the leaf angle decreased,meanwhile,the root length increased and the seedling length shortened after seed germination,and the leaves were not sensitive to BL.However,the plant height,panicle length and leaf angle of RNAi-OsBAK1P transgenic rice increased,meanwhile,root length shortened and seedling length increased after seed germination,and the leaves were sensitive to BL.To sum up,these results provide theoretical support for changing rice plant structure to increase grain yield and may provide a reference for further studies on other functions of OsBAK1 and its precursor OsBAK1P.

In order to provide a theoretical basis for rational application of organic fertilizers and ensure the healthy development of soil ecosystems in corn fields,the effects of soil fungal communities under different levels of organic fertilizer application for the composition,diversity,and functional characteristic were studied.In the meantime,a field experiment was used,with a total of 4 treatments were set up,and the high-throughput sequencing and FUNGuild method were used to analyze the soil fungal diversity,community structure and functional groups in different fertilization rates.The results showed that the increased application of organic fertilizer could increase the content of soil organic matter,available phosphorus,available potassium,urease,catalase,etc.,and it was positively correlated with the application amount of organic fertilizer.Applied organic fertilizers could increase the diversity of soil fungal communities and reduce their richness.The dominant fungal communities in the soil under different fertilization rates were Ascomycota,Mortierellomycota,Basidiomycota,and Olpidiomycota,the relative abundance of Ascomycota and Basidiomycota was significantly higher in the treatment with a medium amount of cattle manure than in the control without organic fertilizer;from the perspective of fungal genera,the dominant genera include Fusarium,Humicola,Olpidium,and Microidium.The application of organic fertilizer increased the relative abundance of symbiotic and saprophytic nutrients,and with the increase of organic fertilizer application,the abundance of pathological nutrients showed a decreasing trend;the number of woody saprophytic fungi in the treatment with increased organic fertilizer application was significantly higher than that in the treatment without organic fertilizer application,while the number of plant and animal pathogens was lower than the control.Therefore,it is believed that increasing a certain amount of organic fertilizer application could optimize the soil microbial environment and promote the increase of corn plant yield.

Grapes are often subjected to various abiotic stress during their growth and development,such as drought,salt,heat or chilling,among them,drought stress is one of the most important abiotic stresses inhibiting vegetative and reproductive growth of grape.Although grapes are more resistant to drought stress,about half of our country's main grape cultivation area belongs to arid and semi-arid climates,drought is one of the main factors restricting the development of our country's grape industry.To ensure the healthy development of our country's grape industry,the research on the effects of drought stress on grape,the formulation of rational irrigation system and the breeding of drought-resistant varieties have become the research hotspots in recent years.Due to the extensive effects of drought stress on grape,in older to cope with drought stress,grapes have evolved many regulatory mechanisms to balance the effects of drought stress in grape growth.Firstly,the water demand degree of grape in different growth period was analyzed according to the law of grape water consumption,and the effects of drought stress on main physical and chemical indexes were exhibited,including photosynthesis,osmotic regulation and reactive oxygen regulation.And then,the quality and yield of grape fruit were analyzed under drought stress,the effects of drought stress on grape fruit quality were reviewed.We give the prospect for how to use drought stress strategy to improve grape quality and how to further study the molecular mechanism of grape response to drought stress at last.

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.

In order to clarify the effects of high temperature stress during grain filling stage on leaf senescence and yield,a field experiment from 2019 to 2020 was conducted with employing heat-sensitive variety(XY335)and heat-resistant variety(ZD958). Simple plastic greenhouse was constructed for heat treatment(HT)during 12-25 days after silking,the natural temperature was set as control(CK). The two years results showed that from 12 to 25 days after silking,high temperature stress significantly reduced the grain weight and yield of summer maize. For ZD958,the grain yield of HT was reduced by 18.8% and 19.3% than CK and thousand kernels weight(TKW)was decreased by 23.4% and 9.1%,respectively.For XY335,the grain yield and TKW of HT were reduced by 29.4%,26.3% and 20.1%,14.2%,respectively.High temperature during grain-filling stage accelerated leaf senescence. The two years results showed that leaf senescence rate(LSR)of ZD958 and XY335 under HT were increased by 76.4%,140.6% and 135.1%,139.6%,respectively.Leaf area duration(LAD)after silking with HT was reduced by 17.7%-36.5% compared with CK. Ear leaf photosynthesis under HT was lowered by 20.0%-42.9% than CK.Correlation analysis results indicated that accelerated LSR under HT had negative correlation with grain yield but without statistical significance(P=0.064 7).LAD after silking,photosynthesis,and SPAD significantly reduced under HT,which showed significant positive correlation with grain yield(P< 0.05). Taken together,heats tress during grain filling significantly accelerated leaf senescence,thus lowering kernel weight and resulting in yield loss. Under high temperature stress,heat-resistant maize variety had higher regulation ability than high temperature-sensitive maize variety,which could significantly mitigate yield loss caused by high temperature stress.

JAZ protein plays a key role in plant growth and development and stress signaling pathways.In order to explore the regulatory mechanism of JAZ protein in wheat late spring cold,TaJAZ6 gene was cloned from young spike of wheat,and its molecular characteristics,expression characteristics and subcellular localization were analyzed.The results showed that the full-length CDS sequence of the gene was 549 bp,encoding 178 amino acids.The predicted molecular weight of the encoded protein was 18.376 ku,the theoretical isoelectric point was 9.37,and the instability coefficient was 62.44,so it was an unstable protein.The protein encoded by this gene had a TIFY domain and a CCT_2 domain.Phylogenetic tree analysis showed that the protein had the closest relationship with TIFY 11b proteins of Triticum dicoccoides and Triticum urartu.In addition to the basic response elements such as CAAT-box and TATA-box,the promoter region of TaJAZ6 gene also contained hormone response elements,light response elements,low temperature response elements,defense and stress response elements.Real-time Quantitative PCR analysis showed that TaJAZ6 gene was expressed in roots,stems,leaves and young panicles,with the highest expression in roots.The expression of TaJAZ6 gene was also induced by low temperature and methyl jasmonate (MeJA).Under low temperature stress,the expression trends of TaJAZ6 in roots,stems and leaves of Aikang 58 (tolerant to late spring cold ) and Zhengmai 366 ( sensitive to late spring cold ) were the same trend,which significantly increased.After spraying 300,350 μmol/L MeJA,the expression of TaJAZ6 in plants treated with low temperature decreased significantly in both wheat varieties.The expression level of TaJAZ6 in the young panicles after low temperature stress showed an opposite trend.The expression level of TaJAZ6 in the young panicles of Aikang 58 decreased significantly,and increased significantly in the young panicles of Zhengmai 366.It was speculated that the gene might negatively regulate the defense response of wheat to late spring cold stress.By spraying MeJA,the relative expression of TaJAZ6 gene in young spikes of two wheat varieties under low temperature stress was significantly reduced,and the grain number of wheat was increased.Subcellular localization assay showed that TaJAZ6 protein was localized in the nucleus.The results above indicate that TaJAZ6 may play an important role in the response of wheat to late spring cold stress.

In order to study the biological function of NbEHD1 in Nicotiana benthamiana,its gene structure,conserved domains,phosphorylation sites,subcellular localization and evolutionary relationship were predicted by bioinformatics methods.The encoding sequence of NbEHD1 was 1 638 bp,and its genomic sequence contained 16 exons and 15 introns.NbEHD1 protein was predicted located in the cytoplasm,which had 42 phosphorylation sites and had no signal peptide and transmembrane region.NbEHD1 belonged to the P-loop_NTPase superfamily and had a conserved domain specific to the EHD family.Phylogenetic relationship showed that NbEHD1 was closely related to EHD sequences of Solanum tuberosum and Solanum lycopersicum.Blast analysis against N.benthamiana sub-data base in SNG was performed to obtain the predicted full length of NbEHD1,then its full nucleic acid sequence was amplified by gene-specific primers.After obtaining NbEHD1 CRISPR/Cas9 gene editing construct,the vector was successfully transformed into the leaves of N.benthamiana by Agrobacterium-mediated genetic transformation.Eventually 16 T₀-generation plantlets were successfully identified,which provided materials for further research to determine the biological function of NbEHD1.

To analyze the same protein of whey and milk fat globule membrane (MFGM)of jersey yak crossbreeds milk and yak milk by proteomics method. The milk yield of jersey yak crossbreeds was higher than that of yak. If the function of jersey yak crossbreeds milk is like that of yak milk,it will improve the economic level of local herdsmen. Whey and milk fat were separated by centrifugation,and proteins were extracted respectively. The same protein was obtained by qualitative and quantitative analysis of protein by tandem mass tag (TMT)technique. GO functional annotation,KEGG metabolic pathway,protein interaction and other bioinformatics analysis were performed on the same protein. Result analysis indicated that 651 whey proteins and 990 MFGM proteins were identified in the peak lactation period of jersey yak crossbreeds milk and yak milk by label quantitative proteomics,and 298 whey proteins and 283 MFGM proteins were screened. GO annotation analysis showed that the same proteins of jersey yak crossbreeds milk and yak milk whey were mainly involved in the negative regulation of endopeptidase activity,immune response and immunoglobulin production. The same proteins of MFGM were mainly involved in the negative regulation of endopeptidase activity and complement activation. The cell components of the same proteins of whey and MFGM in the classical pathway were mainly extracellular space and extracellular region. The molecular functions involved were mainly guanosine 5'-triphosphate (GTP)enzyme activity and GTP binding. KEGG enrichment analysis showed that the main enrichment pathways of the same proteins in jersey yak crossbreeds milk and yak milk whey and MFGM included complement and coagulation cascades,phage and protein processing in endoplasmic reticulum. During the peak lactation period,the proteins with higher abundance in the milk of jersey yak crossbreeds and yak showed excellent performance in immunity and promoting nutrient absorption. However,the milk yield of jersey yak crossbreeds was higher than that of yak,which would increase the economic income of local herdsmen and provide more valuable plateau dairy products for people.

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.

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.

Nitrogen is an essential element for plant growth and development.It is the main limiting factor of the primary productivity of terrestrial ecosystem and has important significance for plant growth and development.Mycorrhizal fungi form a symbiotic relationship with plants,promote plants to absorb nitrogen from the soil,reduce the restriction of nitrogen on plant growth,and enhance the nitrogen exchange between soil,mycorrhizal fungi,and plants,playing an important role in the nitrogen cycle.It reviewed the utilization of different forms of nitrogen by mycorrhizal fungi,their effects on plant nitrogen metabolism,and their ecological significance on soil nitrogen cycle.Based on the current research status of mycorrhizal fungi' impact on plant nitrogen utilization,it is suggested to combine genomics,transcriptome,proteomics and environmental genomics technologies,focus on studying the nitrogen transport mechanism of mycorrhizal fungi plant symbiosis,analyze the main path of nitrogen exchange and gene interaction network between mycorrhizal fungi, plant, and soil,so as to promote plant nitrogen utilization rate,reduce nitrogen fertilizer application,and promote sustainable agricultural development.

As one of the main products of photosynthesis,sucrose has a variety of physiological functions in plants.As a group of glycosyltransferase enzymes,sucrose synthase plays an important role in the process of sucrose metabolism.In plants,sucrose synthase reversibly catalyzes the decomposition of sucrose and nucleoside diphosphate(NDP)to produce nucleoside diphosphate glucose(NDPG)and fructose.This reaction is the most important rate-limiting step in sugar metabolism.Sucrose synthase exists in the form of a gene family in plants.Phylogeny shows that there are three subgroups of sucrose synthase,and different subgroups have different expression patterns.Recent studies have shown that sucrose synthase has several major physiological functions in plants,such as participating in the synthesis of macromolecular sugars,response to adversity,participating in reproductive growth and regulating fruit quality.This article reviews the research progress of sucrose synthase in recent years,and looks forward to future research directions in order to provide a reference for future research.

In order to explore the interaction between ScPRT1 and Sporisorium scitamineum genes in sugarcane,this study used Saccharum spontaneum and Saccharum as research objects.The ScPRT1(GenBank Accession Number:MT747433)gene was cloned by RT-PCR and was analyzed by bioinformatics,quantitative Real-time PCR and subcellular localization.Bioinformatics analysis showed that the full-length cDNA of the ScPRT1 gene was 1 621 bp,including a complete open reading frame with 1 260 bp in length and encodes 419 amino acids.The ScPRT1 protein with two RING domains and one ZZ domain,was 46.56 ku in molecular weight and was an acidic and unstable hydrophilic protein with nuclear localization signal and no signal peptide.The higher structural elements of protein were mainly random coil.The results of qRT-PCR analysis showed that ScPRT1 was higher expressed in stem pith than in bud,epidermis and leaf.And ScPRT1 was significantly up-regulated under abscisic acid(ABA)stress and in the smut-resistant variety but significantly down-regulated in the smut-susceptible variety under Sporisorium scitamineum stress.The result of subcellular localization showed that ScPRT1 was localized in the nucleus.In the co-expression network between ScPRT1 and Sporisorium scitamineum genes,three genes that encoded effectors with aromatic amino acid residues at the N-terminal were found co-expressing with ScPRT1,and their proteins may interact directly.It is speculated that ScPRT1 plays an important role in sugarcane hormone signal transduction and response to smut infection.

In order to explore the gene expression regulation mechanism in the formation of reactive oxygen species (ROS) induced by heat stress,heat stress treatments were set at seedling,heading and filling stage to investigate the dynamic change of ROS accumulation in rice,respectively.And the Quantitative Real-time PCR (qRT-PCR) was used to analyze the expression pattern of nine respiratory burst oxidase homologue (Rboh) encoding genes (OsRboh1-OsRboh9) in rice under different growth stages.Results showed that the ROS accumulation in rice leaves and grains significantly increased with the extension of heat stress.The ROS content increased slowly after 7 days of heat stress at seedling stage, while increased continuously after heat stress during the heading stage and early filling stage (1-10 d) in rice grains. Expression levels of the nine OsRboh family genes continuously increased with the extension of heat stress at seedling and heading stages,and OsRboh7 and OsRboh5 showed higher expression levels under heat stress.The expression levels of OsRboh1,OsRboh5 and OsRboh9 were continuously increased,while other genes showed a change tendency of increasing initially and then decreasing under heat stress at filling stage.The expression levels of OsRboh7 and OsRboh5 were all reached to a high level at seedling,heading and filling stage under heat stress.Furthermore,higher expression levels of OsRboh7 and OsRboh5 were showed in various tissues and organs of rice such as seedling leaves,flag leaf,floret,lemma,palea,stamen,pistil and grain.The higher induced range of gene expression levels in OsRboh7 and OsRboh5 by heat stress was shown in seedling leaves,floret,stamen,pistil and grain.Taken together,OsRboh7 and OsRboh5 were mostly remarkably responsive to heat stress at different growth stages among the OsRboh family genes in rice,which indicated that OsRboh7 and OsRboh5 played a vital role in the regulation of ROS formation pathway in rice under heat stress condition.

In order to determine the suitable sowing date of green manure for multiple cropping rape in Hetao irrigation area,field experiments were carried out in Hangjinhou Banner,Bayannur City,Inner Mongolia Autonomous Region in 2019 and Dalad Banner,Ordos City in 2022.There were five sowing dates in Hangjinhou Banner experimental site from July 26th to August 15th,2019,and one sowing date was set every five days,which were sown on July 26th,July 31st,August 5th,August 10th and August 15th,respectively.Similarly,from July 22nd to August 11th,2022,the experimental site in Dalad Banner was set up with five sowing dates,and one sowing date was set every five days to study the effects of different sowing dates on the green manure biomass,nutrient content and nutrient accumulation of rape.The results showed that although the biomass level and climate conditions of the two experimental sites were different,the overall trend showed a decline with the delay of sowing date.Compared with the first sowing stage,the rape biomass in the fifth sowing stage decreased by 90.3% and 75.4% respectively,and the average active accumulated temperature,effective accumulated temperature and sunshine hours in the two-point growth period decreased by 469.9 ℃,409.9 ℃ and 179.1 h,respectively.At the same time,when sowing in the first,second and third stages,compared with the nutrient input (N 27.3 kg/ha,P₂O₅ 34.5 kg/ha),the nutrient input of rape was higher than the input,and the amount of carbon and potassium returned to the field was at least C1 800 kg/ha and K₂O 200 kg/ha.Therefore,considering the biomass of rape green manure and the amount of nutrients returned to the field,rape could be sown early after wheat harvest in Hetao irrigation area,making full use of light and temperature resources to promote rape growth and nutrient accumulation.In order to achieve more than 5 t/ha of rape returning to the field and obtain a higher amount of nutrients returning to the field,at least 1 300 ℃ of active accumulated temperature,1 100 ℃ of effective accumulated temperature and 640 h of effective sunshine hours should be accepted during the rape growth period.

Obtaining the LaeA gene sequence of Ganoderma lucidum through electronic cloning,analyzing its gene sequence information and preliminarily exploring its regulatory function.This study adopts the method of electronic cloning,using the known LaeA protein sequence of Penicillium citrinum as a template,perform sequence similarity search and alignment(Blast)in the EST database of Ganoderma lucidum and obtain the cDNA sequence of the LaeA gene of Ganoderma lucidum through electronic cloning methods such as sequence splicing,sequence validation and sequence extension.Some characters of amino acids encoded by LaeA gene,including the physical and chemical properties,hydrophobicity/hydrophilicity,subcellular localization,secondary and tertiary structure of protein,and phylogenetic relationship were analyzed by bioinformatics tools.The length of LaeA gene from G.lucidum was 1 134 bp,encoded 378 amino acids.The protein encoded molecular weight of 42.895 3 ku.The protein was an instability protein that was present in the cytoplasm and not secreted to the extracellular.Furthermore,the structure of LaeA protein was mainly composed by 47.88% random coil,33.33%α-helix and 18.78% extension strand,contained SAM binding site,belonged to AdoMet_MTases superfamily proteins.Phylogenetic analysis showed that LaeA was closely related to white rot basidiomycetes such as Trametes versicolor and Dichomitus squalens;the Real-time Fluorescence Quantitative PCR results showed that the expression level of LaeA in Ganoderma lucidum cells during liquid static culture was significantly higher than in oscillatory culture.It is speculated that the LaeA protein as a methyltransferase protein that participates in histone methylation modification,thereby affecting the expression level of gene clusters.

To investigate the influence of green manure cultivation on the carbon and nitrogen content of saline-alkali soil under freshwater leaching,a field experiment was conducted from October 2021 to May 2022.Three treatments were set up:winter fallow(T1),Dongmu 70 rye(T2),and rape(T3).The spatial distribution of soil organic carbon(SOC),nitrate nitrogen($\mathrm{NO}_{3}{ }^{-}-\mathrm{N}$),and ammonium nitrogen($\mathrm{NH}_{4}^{+}-\mathrm{N}$)in the soil and leachate were measured.The results revealed that in the 0—30 cm soil layer,the soil organic carbon content of T1,T2,and T3 increased from 6.20,6.58,7.24 g/kg before leaching to 6.48,7.39,8.06 g/kg after leaching,representing an increase of 4.41%,12.20%,and 11.23%,respectively.After freshwater leaching,the nitrate nitrogen content in the 0—60 cm soil layer of T1 was significantly higher than that of T2 and T3.In the 0—30 cm soil layer,the respective reductions for each treatment were 42.42%,3.85%,and 10.84%.In the 60—90 cm soil layer,the reductions were 1.38%,7.96% and 18.11%.There were no significant differences in ammonium nitrogen content among the different treatments before leaching,but after leaching,the highest ammonium nitrogen content was observed in T2.In conclusion,after leaching irrigation,the soil organic carbon content in different soil layers increased compared to before leaching,while soil nitrogen showed a significant decrease.Analysis of nitrogen content in the soil and leachate indicated that the main nitrogen loss caused by leaching irrigation was in the form of nitrate nitrogen.Compared to winter fallow farmland,the cultivation of rapeseed green manure had a significant effect on increasing soil nitrogen content,while Dongmu 70 green manure was the the most effective in reducing soil nitrogen loss.

To improve the production quality of matcha in tea gardens,the effects of interaction of shading height and shading rate on matcha chlorophyll content,quality components,tea garden environment,etc.were studied through a field experiment,10 treatments in total,including non-shading treatment (CK),shading heights (1.0,1.5,2.0 m)of main treatments and shading rates (40%,70%,100%)of subtreatments were set. Compared with the CK,the results showed that interaction of shading height and shading rate could improve the daily average temperature,daily maximum temperature,and daily minimum temperature of the surrounding environment of tea garden.Interaction of shading height and shading rate were beneficial to reducing illuminance on the surface of tea canopy,increasing the relative chlorophyll content (SPAD value)and significantly increasing the chlorophyll content.The interaction of shading height and shading rate significantly increased the content of water extract and ammonia,and significantly decreased the content of tea polyphenols,caffeine content,and phenol ammonia ratio in tea.Redundancy analysis of tea quality components and chlorophyll content with corresponding environmental factors showed that illuminance and minimum temperature were the main indicators that affected the quality components and chlorophyll content of tea.The interaction of a shading height of 1.5 m and a shading rate of 70% is conducive to the production of high-quality matcha.

By analyzing different materials of Medicago sativa L. and Medicago falcata L.,this study aims to observe the leaf morphology and tissue anatomical structure of alfalfa under drought stress,and explore the adaptive changes in leaf morphology and anatomical structure of alfalfa seedlings under natural drought stress.This study used three alfalfa and two Medicago falcata L. as research materials,and set four different soil relative moisture contents(70%,55%,40%,and 25% of the maximum soil moisture capacity)for experiments. After 30 days of treatment,the morphological and anatomical structures of the leaves were determined. To investigate the response of alfalfa seedling leaves to natural drought stress by observing the leaf morphology and anatomical structure of purple and sickle alfalfa seedlings through electron microscopy. Drought stress would inhibit the growth of alfalfa leaves. With the increase of drought stress,the soil moisture decreased,the leaf length,leaf width,and leaf area of alfalfa and sickle alfalfa decreased,the thickness of upper epidermis increased,the thickness of leaf,mesophyll,and lower epidermis decreased,the thickness of palisade cell and sponge tissue decreased,the thickness of midrib center decreased,the palisade sea ratio gradually decreased,and the compactness of leaf tissue structure increased,the degree of looseness decreased and the degree of leaf vein protrusion increased. The intensity of drought stress was negatively correlated with leaf length,leaf width,leaf area,leaf thickness,mesophyll thickness,lower epidermis thickness,palisade cell thickness,sponge tissue thickness,midvein center thickness,palisade sea ratio,and leaf tissue structure porosity.It was positively correlated with the thickness of the upper epidermis,the tightness of leaf tissue structure,and the degree of leaf vein protrusion.

In order to explore the function of ACS gene in herbaceous peony,a full-length cDNA sequence of PlACS cDNA in Paeonia lactiflora was obtained,RACE technique and bioinformatic methods were used to analyze the protein sequence which it encoded.The CDS of PlACS was subcloned,the prokaryotic expression vector of PlACS was constructed based on pET32a vector,and then the highly efficient prokaryotic expression system was established.The results showed that the total length of PlACS cDNA(GenBank accession JX512359)was 1 752 bp,which encoded 492 amino acids.Seven conserved regions and active sites K₂₇₈ were detected in PlACS protein.Phylogenetic tree analysis showed that PlACS was highest homological with ACS of P.suffruticosa.PlACS protein was determined structurally to be 40.04% α-helix,16.26% β-extended strand,6.91% β-turn and 36.79% random coil.Protein 3D structure homology modeling predicted that PlACS existed as homodimers.The optimal expression condition of PlACS protein was that when the cell density of genetic engineering strain A₆₀₀ reached 0.2,IPTG with a final concentration of 0.1 mmol/L was added,and the recombinant protein was expressed for two hours at 37 ℃.It was of great significance to acquire PlACS recombinant protein with biological activity by denaturation & renaturation and identify its enzymatic activity in vitro.

To investigate the genetic polymorphism of the inbred lines and hybrids derived from Cucurbita moschata (MO)×Cucurbita maxima (MA),the genetic diversity of 53 pumpkins,including commercial varieties,core parent materials and inter-specific hybrids offspring,were investigated by thirty morphological traits and twenty SSR markers.The results showed that the core parent materials and offspring of inter-specific hybrids had more polymorphism in plant type,growth vigor,fruit skin color and shape than commercial varieties.Twenty SSR primers were screened and showed 90% of polymorphism ratio.Based on the principal components,the cluster results of morphological and SSR markers were consistent with each other very well.The two methods and three times cluster results showed that the commercial varieties and core parent materials of Cucurbita maxima were clustered into one group,firstly.This mean the commercial varieties and core parent materials of Cucurbita maxima had relatively close genetic distance and shared the severely homogenized genetic background.It's very hard to produce the new and different varieties in this situation.Secondly,the offspring of inter-specific hybrids derived from MO×MA were clustered into one group,which was distinguished from commercial varieties and core parent materials of Cucurbita maxima and the genotypes of Cucurbita moschata.The offspring of inter-specific hybrids had relatively long genetic distance and more polymorphism than commercial varieties.It's possible to derive the new and different pumpkin varieties by crossing with the genotypes having the heterogeneous genes produced from interspecies hybrids.Meanwhile,the 6 principal component factors derived from principal component analysis would be used to simplify the edible pumpkin survey indicators.

To further investigated the effect of different allelic variants of TaGAMyb-B genes in wheat on stem elongation, used the rice Agrobacterium transformation system,RT-qPCR,tissue section and cell tissue specific analysis to systematically study the function of the 84 bp InDel of TaGAMyb-B.The results showed that,in the over-expressed transgenic rice lines,transcript expression of TaGAMyb-B was detected in seeds,roots,stems and leaves;TaGAMyb-Bb-GFP transgenic seeds were more sensitive under the treatment of NaCl,GA and mannitol compared with TaGAMyb-Ba-GFP;the diameters of first,second and third stem inter-nodes,spike length and tiller number of TaGAMyb-Ba-GFP were significantly larger than that of TaGAMyb-Bb-GFP;and the analysis of cell tissue sections showed that the average thickness of the thick-walled tissue cells in the transverse sections of the transgenic TaGAMyb-Ba-GFP rice was significantly greater than that of the transgenic TaGAMyb-Bb-GFP event,the average length of thick-walled cells was extremely significantly shorter than that of transgenic TaGAMyb-Bb-GFP event.The above mentioned results indicated that the 84 bp deletion in TaGAMyb-B increased not only abiotic stress resistance and plant lodging resistance,but also spike length and tiller number in transgenic rice.

Conservation tillage and organic fertilizer application are one of the effective solutions to alleviate ecological vulnerability in dry farming areas of the Loess Plateau.Reasonable tillage and fertilization measures are of great significance for the realization of two crops a year in the arid area of central Shanxi.The dry farmland in the Loess Plateau was taken as the research object,and the split-plot experimental design was adopted,the main area was three tillage methods (deep tillage (DT),deep loosening (SS),no tillage (NT)),and the sub-area was four fertilization levels (No fertilizer control (CK),full chemical fertilizer (CF),50% chemical fertilizer+50% organic fertilizer (OF),full organic fertilizer (OM)).The changes of soil bulk density,available nutrients and annual yield of wheat and maize under different tillage and fertilization methods were explored.The results showed that in 0-20 cm the soil bulk density of SS and NT treatments were lower than that of DT treatment after organic fertilizer application.The soil bulk density of SS+OF treatment was 1.13 g/cm³,which was significantly lower than that of DT+OF treatment in wheat season.In the whole annual rotation system,the soil mass water content of NT+OM treatment was the highest,which was 7.88 percent point higher than that of other fertilization treatments,and the three-phase ratio of soil under SS+OM treatment was more ideal,and the deviation value of three phase was low.Under the three tillage methods,the content of available nitrogen and available phosphorus generally showed the trend of OM>OF>CF>CK,and the content of available potassium was significantly related to the fertilization method,which was the highest under the CF treatment.The application of organic manure in maize season significantly increased the fresh ear yield of maize compared with a single application of chemical fertilizer,but the difference between them was not significant.The annual yield of wheat and maize under SS+OM treatment was the highest,reaching 19 145 kg/ha.In summary,under the conditions of this experiment,the combination of SS,NT and organic fertilizer application could significantly improve the physical properties of the soil,and the content of alkali-hydrolyzable nitrogen and available phosphorus also had a certain degree of improvement,and SS+OM treatment was beneficial to the increase of the yield of wheat-maize continuous cropping field in the Loess Plateau.

Calmodulin is an important Ca²⁺ receptor protein in plants,which plays an important role in calcium signaling pathway and stress resistance.It is of great significance to study the mechanism of CaM protein in tomato under low temperature stress for cold tolerance breeding of tomato.Tomato calmodulin genes SlCaM3, SlCaM4 and SlCaM5 were cloned from tomato varieties Heinz1706,LA3969,Jifen 2,Jifen 3 and Nongbofenba 15,and their sequences and protein sequences were analyzed by bioinformatics;the cis acting elements in the promoter regions of SlCaM3,SlCaM4 and SlCaM5 were analyzed by plantcare;Quantitative Real-time PCR was used to analyze the expression patterns of SlCaM3,SlCaM4 and SlCaM5 under 15,5 ℃ temperature stress in different tomato varieties,and the specific expression of SlCaM3,SlCaM4 and SlCaM5 in different tissues was analyzed in combination with RNA-seq data.The results showed that the coding sequence of SlCaM3,SlCaM4 and SlCaM5 were 450 bp,and their similarity was 93.63%;the encoded amino acid sequences were identical,belonging to acidic stable hydrophilic protein with typical conservative domain of cam protein.The analysis of cis acting elements showed that the promoter regions of the three genes contained not only the necessary core elements,but also a variety of biotic and abiotic stress response elements,and showed a complementary pattern.The analysis of the expression patterns of different degrees of low temperature stress showed that the expression patterns of SlCaM3,SlCaM4 and SlCaM5 in five tomato materials showed a trend of first decreasing and then increasing at 15 ℃,and the expression of SlCaM5 increased more significantly.At 5 ℃,SlCaM3 and SlCaM4 did not change significantly,but the expression level of SlCaM5 gene increased significantly in the later stage of treatment.The high expression of SlCaM5 gene in the process of tomato resistance to low temperature indicated that it maintained the translation level of cam protein under low temperature and ensured that the function of CaM protein was not affected.The analysis of SlCaM3,SlCaM4 and SlCaM5 specific expression in different tissues of Heinz1706 showed that SlCaM3 and SlCaM4 were highly expressed in meristem,while the expression of SlCaM5 different tissues was not significantly different.

To investigate the effects of Piriformospora indica combined with Arbuscular mycorrhizal fungi (AMF) on drought resistance of tobacco,Yunyan 87 was inoculated with sterile water (CC),P.indica (CP),AMF (PC),P.indica and AMF (PP),and the contents of proline (Pro),superoxide dismutase (SOD),peroxidase (POD),malondialdehyde (MDA) and the expression of drought-related genes NTNAC1 and NAC4 in tobacco leaves were determined under stress in the form of natural drought.The results showed that P.indica and AMF could promote the growth of above-ground and underground parts of tobacco,and chlorosis of leaves and symptoms of their drought were slight.After 7 days of drought stress,the contents of Pro in tobacco leaves of CP,PC and PP groups were 1.39,1.59 and 1.78 times higher than those of CC in group,respectively.The activities of SOD and POD in tobacco leaves increased first and then decreased.The activities of SOD in CP,PC and PP groups were 1.15,1.22 and 1.33 times higher than those in CC group,POD activity was 1.33,1.46 and 1.85 times higher than those of CC group,respectively.The content of MDA in tobacco leaves was decreased by 21.98%,23.98% and 24.84% in CP,PC and PP groups respectively.The expression levels of NTNAC1 and NAC4 in tobacco leaves were up-regulated.The expression levels of NTNAC1 in CP,PC and PP groups were 3.37,3.88 and 5.07 times higher than those in CC group,the expression levels of NAC4 gene were 3.04,3.59 and 5.56 times higher than those of CC group,respectively.This study indicates that P.indica and AMF showed significant synergistic effects,which could significantly improve the drought resistance of tobacco.

In order to clarify the similarities, differences and biological functions of Nodule inception(NIN)genes, which expressed core transcription factors NIN involved in all the biological processes from rhizobia infection to root nodule formation and nitrogen fixation in soybean, and integrates nodulation and autoregulation of nodulation signals to dynamically control the number of nodules, methods of bioinformatics were used to analyse the phylogeny, protein sequence, gene sequence and promoter elements of the four GmNINs, and Real-time PCR were carried out to verify the tissue expression pattern.The results showed that the four GmNINs proteins with similar protein sequences belonged to the specific NIN protein family in Legumes, located in the nucleus, had multiple phosphorylation sites and similar core tertiary structure with significant difference in corner. Acorrdingly, the above results indicate that the four GmNINs proteins may be redundant in core functions, but have differences in specific expression patterns.Analysis of the upstream-3 kb promoter sequence of GmNINs showed that the promoter sequence of GmNINs contained many elements such as ABRE, DRE1 and other elements related to stress and hormone response, except NBS and CYC elements.Transcriptome analysis and Real-time PCR results showed that GmNINs were highly expressed in nodules and inhibited in high nitrogen condition; they responded to salt and drought stress of varying degrees, among which GmNIN2a and GmNIN2b were more sensitive and significant in responding to abiotic stress, and GmNIN2a and GmNIN2b may play an important role in the process of plant response to abiotic stress. In short, the results revealed that GmNINs not only regulate the nodule number of soybean and nodulation, but also participate in the process of root response to abiotic stress. The discovery of this regulatory mechanism provides essential clues for breeding new varieties with high quality and high yield.

In order to investigate the structural characteristics of Brassica juncea AHK family proteins and their regulatory functions related to seed weight traits,it employed bioinformatics methods to perform whole-genome identification,physicochemical properties analysis,phylogenetic tree construction,protein structure analysis,promoter cis-elements analysis,and differential tissue expression profiling of the BjAHK family members.The main findings were as follows:a total of 19 BjAHK protein sequences were identified from the B.juncea genome,and phylogenetic analysis classified the BjAHK family members into four branches:G1 (BjAHK2—BjAHK4),G2 (BjAHK5),G3 (BjAHK1),and G4 (BjCKI1).Analysis of functional domains revealed the presence of six conserved motifs (Motif1—Motif6) in BjAHK family members,corresponding to the HisKA,HATPase_c,and REC conserved domains.The N-terminal regions of BjAHK1—BjAHK4 and BjCKI1 proteins exhibited relatively conserved transmembrane domains,which might be associated with their transmembrane function.Differential tissue expression profiling revealed that the genes encoding cytokinin receptor proteins (BjAHK2—BjAHK4) were constitutively expressed genes,with the highest expression levels observed in root tissues.Through integration with existing transcriptomic data on seed weight traits in B.juncea,four candidate genes potentially involved in the regulation of seed weight traits were identified:BjA07.AHK3,BjB03.AHK3,BjA10.AHK5,and BjB05.AHK5.BjA07.AHK3 and BjB03.AHK3 might positively regulate seed development,while BjA10.AHK5 and BjB05.AHK5 might have the opposite effect.Based on the integration of gene family analysis and transcriptome result,it was inferred that BjAHK3 and BjAHK5 genes play an important role in the regulation of seed development in B.juncea.

To explore the function of the TCP gene family in the growth and development of Jasminum sambac,bioinformatics methods were used to identify and analyze the whole genome of J.sambac TCP(JsTCP)gene family.Moreover,the expression levels of TCP gene family in different stages of flower development and pollen-stigma interaction were analyzed.A total of 27 TCP gene family members were identified from the whole genome of J.sambac,named JsTCP1—JsTCP25.The protein contained 208—539 amino acid residues,with molecular weight of 22.95—56.96 ku and isoelectric point of 5.70—9.97,and all of them were unstable hydrophilic proteins.The subcellular localization prediction showed that all JsTCPs were located in the nucleus.JsTCPs were unevenly distributed on 10 chromosomes.Gene structure analysis showed that JsTCPs had 1—5 exons and 0—4 introns.Protein conserved motifs and phylogenetic analysis showed that JsTCPs contained conserved TCP domain,and they were divided into two subclasses:Class Ⅰ and Class Ⅱ.Promoter of JsTCPs contained cis-acting elements related to plant hormone response,stress response,and growth and development.The expression pattern analysis showed that 24 JsTCPs were expressed at different stages of flower development,and 22 genes were expressed in pollen-stigma interactions.In conclusion,27 TCP gene family members were identified from J.sambac,and it found that they expressed specifically at different stages of flower development and different stages after pollination.

Breeding resistant varieties is the most economical and feasible method to control wheat leaf rust.In order to further explore the resistance genes,50 wheat varieties were selected in this study,including wheat varieties from Shandong,Henan,Hebei,Shandong and other 8 provinces.Firstly,16 physiological races of leaf rust fungus (THFS,TGTS,THJS,FHKT,FGJN,KHKS,FCJQ,RFKS,THFM,MHGT,KHGS,KBGT,FHGT,PHHT,FHJT,FCJT) were inoculated to 36 vector cultivars containing known leaf rust resistance genes,and 50 wheat varieties tested during the seedling stage.Due to the different virulence of each strain,specific molecular markers closely linked to known disease resistance genes can be combined and analyzed based on the differences in phenotype,and it can be inferred that 50 wheat materials may contain leaf rust resistance genes.The leaf rust resistance genes were identified by genetic deduction,molecular markers and pedigree analysis.The results showed that a total of 9 known resistance genes (Lr1,Lr2c,Lr10,Lr16,Lr26,Lr34,Lr37,Lr45 and Lr46) and a few unknown genes were detected in 50 cultivars.There were 22 varieties including Zimai 12 that contained the Lr1 gene;there were 10 varieties including Lumai 14 that contained the Lr2c gene;Laizhou 9361 only contained the Lr10 gene;there were 25 varieties including Kenong 199 that contained the Lr16 gene;there were 15 varieties including Xuzhou 24 that contained the Lr26 gene;Baomai 3 and Jingdong 8 contained the Lr34 gene;The Lr37 gene was contained in Zimai 12,He 0927,and He 9946;there were 11 varieties including Lianmai 2 that contained the Lr45 gene;there were 38 varieties including Shannong 19 that contained the Lr46 gene.

Fruit development is the key stage which determines the yield and quality of grape.WRKY family transcription factors play important roles in regulating plant development and environment adaptation.Ethylene is the important plant hormone which participates in regulation of fruit development,and ACC synthase is the key enzyme that limits ethylene synthesis.Using grape variety Zuoyouhong, VvWRKY13 overexpressing grape callus,as well as VvWRKY13 heterologous over-expressed tomato lines as materials,the role of VvWRKY13 from grape in fruit development and its relationship with ethylene were studied though plant physiological and biochemical methods as well as molecular biological techniques.The results displayed that the expressions of VvWRKY13 as well as ACC synthase genes VvACS2 and VvACS7 significantly upregulated at early stage of fruit development,and in VvWRKY13 over-expression grape callus,the expression of VvACS7C was significantly higher than control,but the expression of VvACS2 displayed no significant difference compared with control.Yeast one hybrid experiment showed that VvWRKY13 could directly bind to the VvACS7 promoter,VvWRKY13 had no direct interaction with VvACS2.We also found that the ethylene content and the expression of ACS family members,such as SlACS1b,SlACS4 and SlACS6,the ethylene synthesis genes,were significantly induced in VvWRKY13 heterologous over-expressed tomato,and the time from flowering to breaking of tomato was 3—6 days shorter than wild type;the above results indicated that,VvWRKY13 could regulate ethylene synthesis by promoting the expression of ACC synthase gene to participate in regulation of fruit development.