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.

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 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.

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 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.

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.

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.

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.

In order to reveal the molecular mechanism of MiPDCD6 protein suppressing tomato PTI immunity,the MiPDCD6 overexpression seedlings of tomato variety Xinjinfeng 1 were used as experimental materials,and the tissue culture seedlings of tomato variety Xinjinfeng 1 were used as control.Transcriptome sequencing was performed on tomato MiPDCD6 overexpressing seedlings and control seedlings,respectively.With tomato cultivars Heinz 1706,comparing the genome as a reference genome,FPKM method was used to calculate quantity of gene expression,set parameters(|log₂ FC|>1 and P<0.05)in screening the differentially expressed genes(DEGs).Gene ontology(GO)database was used to analyze the GO functional enrichment of DEGs,count the number of DEGs in each GO term,calculate the significance of gene enrichment,and find out the functional term with significant enrichment.KEGG database was used for Pathway enrichment analysis of DEGs,and hypergeometric distribution test was used to calculate the significance of enrichment of DEGs in each Pathway.The enrichment degree of KEGG was measured by FDR and gene number.Combined with gene differential expression analysis and functional enrichment analysis,the effect of MiPDCD6 protein on tomato PTI immune-related pathway genes was studied.The results showed that there were 2 366 DEGs in MiPDCD6 overexpressed tomato plants compared with wild-type tomato,including 1 354 up-regulated genes and 1 012 down-regulated genes.In these DEGs,a large number of differentially expressed genes were concentrated in KEGG pathways such as plant hormone signal transduction(sly4075),plant-pathogen interaction(sly04626),plant MAPK signal pathway(sly04016)and procycloid biosynthesis(sly00940)through GO and KEGG annotation.SA biosynthesis pathways included ICS and PAL.In the MiPDCD6 overexpressed tomato plants,PAL1 and PAL-like genes in SA synthesis pathways and TGA9,TGA10-like and PR1a2 genes in SA signal transduction pathways were significantly down-regulated,suggesting that MiPDCD6 may inhibit SA synthesis thus inhibiting plant PTI immunity.

Genomic-wide identification of systematic work on the SKS gene family in cotton has not been reported.Identifying SKS (Skewed5 similar) gene family members and analyzing evolutionary relationship in Gossypium hirsutum,and analyzing the mechanism of GhSKS13 gene regulating cotton disease resistance are expected to provide a new direction for the cultivation of resistant plants.Here,the members of upland cotton SKS family were identified by bioinformatics at the genome-wide based on published genomic data of upland cotton genetic standard line TM-1,and G.hirsutum variety CRI-14 was used as the material.And chromosome distribution,evolutionary relationships,gene structure,and collinearity of SKS family members were predicted.Then,the expression pattern of GhSKS13 was analyzed by Real-time Quantitative PCR and virus-induced gene silencing was employed for preliminary investigation of the function of GhSKS13 in cotton resistance to Verticillium dahliae.A total of 48 upland cotton SKS genes were identified,unevenly distributed on 19 chromosomes,clustered into 5 subgroups with highly conserved gene sequences.Collinearity analysis revealed that the upland cotton SKS gene family was subject to purifying selection.Expression pattern analysis showed that GhSKS13 was predominantly expressed in upland cotton root tissues and significantly up-regulated by V.dahliae infection. GhSKS13-silenced reduced plants resistance to V.dahliae and suppressed the expression of the pathogen-related genes including GhPR1, GhPR2,GhPR3 and GhPR5 compared to control plants. GhSKS13-silenced plants invaded by V.dahliae showed significantly lower hydrogen peroxide (H₂O₂) deposition compared to the control,suggesting that GhSKS13 promotes the formation of reactive oxygen species (ROS).In conclusion,this study clarified the phylogenetic relationships,chromosomal distribution characteristics and gene structure characteristics of upland cotton SKS family members,and elucidated the involvement of GhSKS13 in upland cotton plant resistance to V.dahliae.

Partial substitution of organic fertilizer nitrogen for chemical fertilizer nitrogen is one of the ways to achieve sustainable crop development.This study explored the appropriate ratio of wheat organic nitrogen partial substitution for chemical nitrogen,as well as the characteristics of nitrogen accumulation,transportation,and utilization after substitution,in order to provide a basis for nitrogen fertilizer reduction and efficiency enhancement technology for winter wheat in Hebei Region.Field experiments of the following nine treatments were conducted in Ningjin,Hebei from 2021 to 2023:T1,non-nitrogen,and application of chemical P and K fertilizer alone;T2,high efficiency fertilization treatment,and application of chemical N,P and K fertilizer alone;T3—T7,substitution of 20%,40%,60%,80% and 100% the chemical N rate of T2 with organic fertilizer,respectively;T8,traditional fertilization,and application of chemical N,P and K fertilizer alone;T9,substitution of 100% chemical N rate of T2 with organic fertilizer,and spraying liquid nitrogen fertilizer at the erecting stage.The results of two years of experiments showed that the 100% substitution rate+liquid nitrogen treatment could achieve the highest wheat yield.Secondly,the yield of the 40% substitution rate treatment was equivalent to that of the high efficiency fertilization treatment,and it was much higher than that of the traditional fertilization treatment in the second year of the experiment.The 100% substitution rate+liquid nitrogen treatment increased the nitrogen content in stems and leaves by spraying available nitrogen during the erecting period,and the nitrogen accumulation in plants was equivalent to that of high efficiency fertilization and traditional fertilization treatments.Treatment with 40% and 80% substitution rates also achieved nitrogen accumulation equivalent to the highly efficient fertilization treatment.The 20%—100% substitution rate treatment (including liquid nitrogen treatment) could achieve a higher nitrogen transfer rate in stems and leaves,as well as the contribution rate of nitrogen transfer to grain.Among them,the 100% substitution rate+liquid nitrogen treatment had good fertilizer nitrogen absorption and utilization effect,achieving higher fertilizer nitrogen accumulation,nitrogen utilization rate,and nitrogen harvest index.Nitrogen fertilizer effect of the 100% substitution rate+liquid nitrogen treatment was equivalent to or slightly higher than that of highly efficient fertilization treatment.Secondly,the treatment with the 40% substitution rate had a similar or slightly lower nitrogen fertilizer effect than the highly efficient fertilization treatment.In summary,the 100% substitution rate+liquid nitrogen treatment showed better wheat yield,plant nitrogen accumulation,nitrogen transport rate,grain nitrogen accumulation,and nitrogen efficiency,followed by the 40% substitution rate treatment.

In order to clarify the relationship between the morphological and structural characteristics of stem basal node 2 and lodging resistance in wheat,we explored key stem morphological indicators and Quantitative trait loci(QTL)sites for lodging resistance.120 RILs families were selected as research materials,and stem strength,basal second internode length,stem diameter,wall thickness,cellulose content and lignin content were measured in 2020 and 2021,respectively.Multiple regression analysis and QTL locations were performed by combining 55K SNP data.The results showed that the stem strength was significantly or extremely significantly positively correlated with the stem diameter and wall thickness of the second basal internode,and was extremely significantly positively correlated with the cellulose content and lignin content of the second basal internode.Multiple regression analysis showed that cellulose content in basal second internode was the key index affecting stalk strength of wheat.A total of 19 QTLs related to stem traits were detected on chromosomes 1A,1D,2B,2D,4D,5A,5B,5D and 7B,explaining 7.67% to 65.33% of the phenotypic variation.On chromosome 1D,the QTL linked to AX-110771095 and AX-109431570 simultaneously controlled the basal second internode length,wall thickness and cellulose content,explaining the phenotypic contribution of 7.96%-10.76%.

To explore the genetic relationship of alfalfa germplasm resources and provide theoretical basis for the identification of alfalfa varieties and the breeding of new varieties.Genetic diversity of 30 alfalfa materials was analyzed by ISSR and SSR markers.A total of 125 bands and 117 polymorphic bands were amplified by 12 ISSR labeled primers.The polymorphic band ratio(PPB)was 93.01%.The average effective allele number(Ne),Nei's gene diversity index(H)and Shannon diversity index(I)were 1.465 9,0.281 3 and 0.431 4,respectively.A total of 152 bands and 144 polymorphic bands were amplified by 12 SSR markers,and the polymorphism band ratio(PPB)was 94.05%.The average effective allele number(Ne),Nei's gene diversity index(H)and Shannon diversity index(I)were 1.542 7,0.313 9 and 0.470 1,respectively.The genetic similarity coefficient and cluster analysis of the two markers showed that Zxy2010p-7900 and Zxy2010p-7740 could be divided into a separate category and were far related to other materials.Qingshui alfalfa,Longdong alfalfa and Gannong No.4 mixed alfalfa can be divided into one group,the genetic similarity coefficient between the varieties is large,and the relationship is close.The phylogenetic relationship of the tested alfalfa germplasm was objectively determined by the two labeling methods.

In order to study the response mechanism of pepper CaWRKY30 transcription factor and Tomato spotted wilt orthotospovirus,it was experimental materials with pepper Xiangyan 11.The CaWRKY30 coding sequence was obtained by RNA extraction,RT-PCR,split gel and cloning.Biological information analysis results showed that CaWRKY30 full length was 1 122 bp,encoding 373 amino acids,the gene encoded protein contains 1 WRKY conservative domain and 1 C₂H₂ domain,belonged to a typical Ⅱ(e)subfamily member.System evolution analysis showed that the relative relationship with the potato StWRKY22 amino acid sequence was recently.It was found that CaWRKY30 was positioned in the nucleus and cell membranes in its cigarette seedlings,and leads to cell membranes.The results of Real-time fluorescence quantitative PCR analysis showed that the viral accumulation of Tomato spotted wilt orthotospovirus mechanical friction-vaccination was found that the viral accumulation was gradually increased from 1 to 14 days after inoculation,and virus accumulation reached its maximum in 14 days,after inoculation 14 days,viral accumulation gradually declined.At the same time,CaWRKY30 was induced by Tomato spotted wilt orthotospovirus,when the inoculation 1—14 days,the CaWRKY30 expression was raised,and the peak was reached in 14 days,the expression in 14 days gradually decreased.In summary,it obtained the CaWRKY30 transcription factor gene sequence,which was located in the nucleus and cell membrane,and preliminarily explained the expression trend of CaWRKY30 transcription factors under the stress of Tomato spotted wilt orthotospovirus.

The bioinformatic analysis of SAUR genes aims to unveil their structure,function,and evolutionary relationships.Simultaneously,through subcellular localization,the goal was to understand their position within the cell,thereby inferring their biological functions.Such analyses contribute to a deeper understanding of the gene's mechanistic role and potential applications within an organism.This held particular theoretical and practical significance for regulating the germination of windproof seeds,reducing seed dormancy duration,and facilitating green cultivation and breeding of wind-resistant plants.In order to explore the biological characteristics and functions of SAUR genes,SdSAUR32 and SdSAUR23 were cloned from Saposhnikovia divaricata,and bioinformatics analysis was conducted to analyze the expression and subcellular localization of these two genes in different dormancy stages of Saposhnikovia divaricata seeds.The findings indicated that the SdSAUR32 protein had a molecular weight of approximately 14.48 ku(pI 6.45),while the SdSAUR23 protein had a molecular weight of around 10.29 ku(pI 8.00),confirming that both proteins were hydrophilic.The conserved domain analysis of SdSAURs gene encoded protein showed that SdSAUR32 and SdSAUR23 had the same conserved domain,which belonged to auxin induced superfamily members.The phylogenetic analysis showed that both SdSAUR32 and SdSAUR23 encoded protein was the closest relative to Daucus carota. The subcellular localization results indicated that both SdSAUR32 and SdSAUR23 were found within the nucleus.The SdSAUR32 and SdSAUR23 genes were found to be expressed during different stages of Saposhnikovia divaricata seed development.It was observed that the expression levels of these two genes were significantly higher during the germinal stage compared to the release dormancy stage and dormancy stage.This suggested that SdSAUR32 and SdSAUR23 may have a crucial role in the germination,growth,and development of Saposhnikovia divaricata seeds.This study offers not only insights for investigating the role of the SAUR gene family but also establishes atheoretical framework for examining the function of other genes in the parsnip family at the molecular level.

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.

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.

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.

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.

The identification of leaf shape mutant and genes controlling leaf shape can not only provide germplasm resources for genetic improvement of leaf shape,but also help to analyze the genetic regulation mechanism of leaf development.vrnl11 was identified from an EMS induced Wankelü 3(WK3)mutant library.Progeny populations derived from vrnl11/WK3 and vrnl11/Zhonglü 1 were used for genetic analysis,and the segregation pattern of different phenotypic plants in F₂ populations was determined by χ² test.Two F₂ populations constructed by crossing vrnl11 with Zhonglü 1 and Zhonglü 5 were used as mapping populations.Fine mapping for vrnl11 was completed by using BSA sequencing technology and map-based cloning strategy.Phenotype identification results showed that,compared with wild-type WK3,the leaf width and leaf area of vrnl11 decreased by 25.7% and 21.7%,respectively.Genetic analysis showed the narrow leaf phenotype of vrnl11 was controlled by a single recessive nuclear gene.BSA sequencing analysis located the mutation site within a 4.7 Mb interval from 15.0 Mb to the chromosomal end on chromosome 11.vrnl11 was finally located in the 186.5 kb interval between the markers nl-61 and nl-46 by using these newly developed polymorphic molecular markers.The mapping interval contained 9 predicted genes.These results provide a theoretical basis for cloning vrnl11 and understanding the molecular regulation mechanism of leaf development in mungbean.

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.

In order to explore the molecular mechanism of Zoysia grass in resistance to abiotic stresses,in the present study we reported the function of a cold responding gene from Zoysia matrella designated as ZmCOR410. The sequence information of the gene was obtained on the base of transcriptome and genomic data. The expression profile of ZmCOR410 in response to low temperature was detected by qRT-PCR,and the functions of the gene under abiotic stresses were estimated in Arabidopsis and yeast by genetic transformation.The results showed that the CDS of the gene was 927 bp in length,encoding an acid dehydrin that contains 308 amino acids. In the polygenetic tree of COR410 homologues from gramineous grasses,ZmCOR410 had a close relationship with the homologues of Cleistogenes songorica and Eragrostis curvula,two species that were high tolerant to drought stress. In the genomic DNA,there were four copies of core sequence of DRE-cis element in the 1 700 bp region upstream of the CDS of ZmCOR410,and its mRNA was accumulated in leaves exposed to cold. Compared to WT plants,Arabidopsis plants over expressing ZmCOR410 showed reduced injury in leaves after a freezing-temperature exposure,and showed higher survival rates under drought and high temperature stresses. Yeast cells harboring ZmCOR410 were also more tolerant to high temperature stress than the control cells. The results indicated that the product of ZmCOR410 could enhance cells resistance to freezing cold,high temperature and drought stresses,which would help Zoysia matrella going through adverse environments.

To further explore the molecular mechanism of pericarp thickness development in sweet corn, pericarp from sweet corn inbred lines M03 and M08 were used for transcriptome sequencing,these materials provided by the sweet corn breeding laboratory of South China Agricultural University.RNA-sequencing material were sampled at 15,19 and 23 days after pollination.Genes’ differentially expressed analysis combined with weighted gene co-expression network(WGCNA)analysis were used to identify the co-expressed gene modules related to pericarp thickness at the milk stage of sweet corn,and the hub genes were also identified.Quantitative real time-PCR(qRT-PCR)was used to validate the expression level of the hub genes.Here,a total of 4 748 differentially expressed genes(DEGs)were identified by comparing different development stages and different samples.Gene annotation analysis revealed that these DEGs were mainly enriched in carbohydrate metabolic processes,plant-pathogen interactions,and plant MAPK signaling pathways.Co-expression analysis identified 18 modules,in which four modules(Turquois,Yellow,Magenta and Pink)were significantly associated with pericarp thickness.GO and KEGG functional enrichment analysis were performed based on these four specific modules,which could be enriched in alanine aspartate and glutamate metabolism,MAPK signaling pathway-plant and cysteine and methionine metabolism.The top 20 hub genes were screed by calculating the gene connectivity in the corresponding net-works,13 hub genes including MYB transcription factor,cell cycle protein(CYC15),β-amylase,apoptosis regulatory gene(BCL-2),etc.were finally screened by functional annotation.Transcription factors such as auxin response factor(ARF,AUX/IAA),ethylene response element binding factor(ERF),and leucine zipper(bZIP)were also included in the co-expression network,and these genes may play a crucial role in pericarp development process.

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.

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.

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.

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.

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.

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 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.

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 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.

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 explore the function and possible molecular mechanisms of amino acid transporters (AATs),firstly,a new hidden markov model was established to identify the members of the rice AAT family,and a phylogenetic tree was constructed to analyze the evolutionary relationship of AAT members between rice and Arabidopsis;secondly,the promoter,protein structure,and functional domain of rice polyamine transporter gene 1 (OsPUT1) were analyzed using bioinformatics methods;thirdly,the subcellular localization of OsPUT1 was determined by constructing the OsPUT1-GFP vector and expressing it in rice protoplasts;fourthly,the expression patterns of OsPUT1 gene in different tissues and under different stress conditions were detected using qRT-PCR;finally,the function of OsPUT1 gene was preliminarily studied using the OE,Nip and RNAi lines.The results showed that there were 96 OsAATs divided into 13 subfamilies in rice;the PUT subfamily contained 6 OsPUTs,among which OsPUT1 and AtPUT3 had the closest genetic distance and were distributed in the same branch.The promoter of OsPUT1 gene contained cis-acting elements related to growth and development,light regulation,plant hormones,and stress response;OsPUT1 protein contained the polyamine transporter domain PotE,and subcellular localization experiment indicated that it was located on the cell membrane;the expression of OsPUT1 gene was relatively high in leaves,while was low in flowers;gene expression was inhibited by JA,mannitol,and ABA,while decreased and then returned to normal under low temperature stress,yet increased and then decreased under SA,Spm and paraquat treatment,however,under sodium chloride treatment,the gene expression level first increased,reached its peak at 1 h,then decreased,reached its lowest at 12 h,and returned to normal levels at 24 h;OE lines significantly reduced resistance to paraquat,while RNAi lines significantly increased resistance to paraquat.Overall,the OsPUT1 protein might have the function of transporting PA and participating in stress response.

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.

In order to alleviate the continuous cropping obstacle of millet,provide a reference for optimizing millet planting mode,millet continuous cropping(Si)was named as control(CK),millet-corn(Si-Zm),millet-potato-corn(Si-St-Zm),millet-corn-soybean(Si-Zm-Gm)and millet-soybean-potato(Si-Gm-St)were used to analyze the effects of different rotation patterns on the physiological indicators,photosynthetic characteristics,agronomic characters,yield and downy mildew incidence rate during the critical millet growth periods.The results showed that compared with CK,in the Si-St-Zm,Si-Zm-Gm and Si-Gm-St rotation patterns,the activeness of superoxide dismutase,peroxidase and polyphenol oxidase in millet flag leaves were significantly increased,with the largest increases of 45.55%,41.55% and 109.09%,respectively.In the Si-Zm-Gm and Si-Gm-St rotation patterns,millet plant height,stem thickness,root length and root branch number were significantly increased,with the largest increases of 30.48%,30.50%,31.76% and 13.79%, respectively.In addition,compared with CK,under the Si-Gm-St rotation system,the H₂O₂ and MDA content in the millet flag leaves were significantly decreased,with the maximum reductions of 18.78% and 47.29%, respectively;and the stomatal conductance,net photosynthetic rate,transpiration rate and relative chlorophyll content were significantly improved by 31.94%—101.43%,35.74%—234.00%,16.44%—46.97% and 24.15%—66.16%,respectively;with millet ear length,1000-grain weight and yield increased by 14.90%,17.09% and 10.58%,respectively;and millet downy mildew incidence rate significantly reduced by 12.33%.In short,compared with CK,the Si-Gm-St rotation system significantly increased the activeness of SOD,POD and PPO,and improved photosynthetic efficiency in millet flag leaves,meanwhile,the millet yield and disease resistance enhanced.Therefore,compared with Si-Zm,Si-St-Zm and Si-Zm-Gm rotation patterns,Si-Gm-St rotation system has the best effect on alleviating continuous cropping obstacles,which can provide a reference for optimizing millet planting system.

LMI1 is a key gene in the regulation of serrated leaf development.In order to study the mechanism of okra leaf development in cotton,we cloned GaLMI1-like gene and GaLMI1-like promoter by PCR amplification technology from Shixiya 1,which is an Asia cotton species of a genome.The sequence lengths were 681,1 439 bp,respectively.Domain analysis found that GaLMI1-like contained a homeobox domain.Then,we constructed a reorganization overexpression vector p6MYC-GaLMI1-like.The transgenic Arabidopsis showed incised leaf shape when GaLMI1-like was overexpressed in Arabidopsis.Moreover,we analyzed the cis-acting elements of the promoter sequence of GaLMI1-like.It was found that it had basic acting elements,such as CACA-box and TATA-box,and also had photoresponsive element,root,stem and mesophyll-specific expression elements.GUS fusion expression vector of GaLMI1-like promoter was constructed and transformed into Arabidopsis thaliana.GUS staining results showed that the promoter could drive GUS gene expression in root column,stem and leaf,and the expression was especially high in leaf.These results indicated that GaLMI1-like had the function of regulating the formation of incised leaves,and its function was realized by the strong expression of GaLMI1-like regulated by GaLMI1-like promoter in cotton leaves.

In order to study the effects of different fertilization pattern on the soil aggregates composition,the contents of total organic carbon and its components,and to clarify the optimal fertilization pattern for improving soil fertility in yellow clayey paddy soil.Based on the long-term fertilization experiment in Jinqu Basin,four treatments were conducted,including no fertilizer,chemical fertilizer alone,chemical fertilizer combined with cattle manure and chemical fertilizer combined with straw.The results showed that chemical fertilizers plus cattle manure resulted in the highest percentage of >0.250 mm aggregate,increased by 7.1,11.0 percentage point compared to the chemical fertilizers alone in 0-15 cm and 15-30 cm,respectively.By contrast,there were no significant differences in the composition of aggregate between chemical fertilizers plus straw and chemical fertilizers alone.On the other hand,chemical fertilizers plus cattle manure showed significantly higher in soil total organic carbon content,particulate organic carbon content in the surface soil,increased by 18.7%,98.7%,respectively.The order of POC/SOC value in the surface soil was as follows:chemical fertilizer plus cattle manure > chemical fertilizer plus rice straw> chemical fertilizers alone ≈ no fertilizer.Compared with the single application of chemical fertilizers,the combined application of chemical fertilizers and cattle manure or straw increased the relative absorption peak intensity of 1 030 cm^-1,indicating that the relative content of polysaccharides was higher under the combined application of organic and inorganic fertilizers treatments.Overall,the combined application of chemical fertilizers and cattle manure was the best practice for improving soil structure and soil carbon pool.

In order to explore a new way to reduce nitrogen and increase efficiency of spring maize,in the agricultural high tech demonstration park of Horqin district,Inner Mongolia,two different ratios of reduced urea gradients and corresponding combined application of urea ammonium nitrate solution(UAN)were set in comparison with conventional topdressing urea(CK1,570 kg/ha)(N1U treatment 225 kg/ha urea + 75 kg/ha UAN and N2U treatment 375 kg/ha urea + 75 kg/ha UAN),the effects of shallow drip irrigation with reduced urea application combined with compared with CK treatment,N2U treatment did not reduce the yield of spring maize when the total nitrogen input was reduced by 25.12%.The total nitrogen input of N1U treatment decreased by 51.42%,and the yield decreased significantly.The total dry matter accumulation of N2U treatment was also significantly higher than that of CK treatment,and increased by 12.84%—16.40% and 6.05%—9.76% at silking stage and mature stage,respectively.Compared with CK,the total nitrogen accumulation in the mature stage increased by 20.55%—42.29%.N2U treatment did not reduce the yield of spring maize under the condition of 25.12% reduction of total nitrogen input,and it was a more reasonable fertilization mode of shallow buried drip irrigation for maize under the same soil fertility in the Xiliaohe plain.