Preliminary Mechanism Study Of Amelioration Of Water Defict Stress By 24-epibrassinolide Treatment And Identification And Expression Analysis Of The BZR Gene Family In Grapevine(Vitis Vinifera L.)

Brassinosteroid（BR）is a plant hormone widely found in plants similar to animal sterol hormones and plays an important role in plant response to abiotic stress.BZR1 is a key transcription factor in BR signal transduction process.It can regulate the expression of BR response gene and play a genetic role in the promotion of cell elongation,seed germination and photomorphogenesis of seedlings.In addition,in cotton,wheat,corn and other plants,researchers have conducted more research on the BZR gene family,and found that most TFs have the function of responding to abiotic stress.Our previous studies have demonstrated the role of BR in improving the resistance of grape seedlings to water stress,but there are few studies on its mechanism.In this experiment,we first carried out genome-wide identification of the grape BZR gene,and systematically analyzed their chromosomal distribution,gene structure and protein conserved domain.Real-time quantitative q RT-PCR was used to detect the expression pattern of BZR gene family under different tissues and water treatment stresses,in order to identify the BZR gene related to grape water stress.In addition,we also studied the expression pattern of drought-resistant genes in grape seedlings under water stress by EBR pretreatment,and laid a foundation for further study on the molecular mechanism of EBR treatment to improve drought resistance of grapes.The results are as follows:（1）Alleviation of grape seedlings under water stress by exogenous BR pretreatment.The water stress caused the grape to have a drought response,the content of H₂O₂ and O₂·^-in the leaves significantly increased,and the contents of endogenous hormones IAA and GA₃decreased.EBR pretreatment could slow down the accumulation of reactive oxygen species in stressed plants,increase the content of ascorbic acid（As A）and glutathione（GSH）in grape seedlings,induce the expression of VviAPX、VviDHAR、VviGR and VviGST genes at different stages and improve APX,DHAR,MDHAR and GR activity,alleviating the inhibitory effect of stress on IAA and GA₃ content,ultimately mitigate the drought response.（2）The effect of exogenous BR pretreatment on the expression of stress-related genes.EBR pretreatment could increase the content of ABA in early seedlings and up-regulate the expression of key genes involved in ABA synthesis and ABA signaling.In addition to ABA-related genes,EBR pretreatment also increased the expression of drought-related genes VviP5CS,VviRD22a,VviMYB60,VviNAC17,and VviWRKY45.It indicates that there is synergy between EBR and ABA,and these genes play an important role in BR regulation of grape resistance to water stress.（3）Identification and structural analysis of grape BZR gene family.We identified eight BZR homologs within the genome of the grape,and named it VviBZR1-8 based on its location on the chromosome.All N-terminals of this family have an atypical basic helix-loop-helix（b HLH）DNA binding motif,the BZR domain,indicating that VviBZR1-8can regulate the expression of related genes by binding to BR response elements.VviBZR2,VviBZR3,VviBZR6,VviBZR7,and VviBZR8 contain a phosphorylation site rich in serine（S）.VviBZR2,VviBZR3,VviBZR6,and VviBZR7 contain a C-terminal domain,which is consistent with the conserved domain of BZR TFs in Arabidopsis thaliana.（4）Tissue specific analysis of grape VviBZR gene family.Except VviBZR7,the other 7VviBZR genes can be specifically expressed in different tissues.VviBZR2 and VviBZR4 have higher transcription levels in roots,and VviBZR1,VviBZR3,VviBZR4 and VviBZR6 are highly expressed in tendril.The relationship between BZR family and grape fruit maturity was studied.It was found that VviBZR1 was highly expressed in the transfection and maturity pericarp,VviBZR5 and VviBZR8 were highly expressed in the fruit peel,and the VviBZR1,VviBZR4,VviBZR5 and VviBZR8 genes were in the green fruit stage.The high expression level indicates that the grape BZR gene family plays diverse functions in different tissues and organs and different developmental stages.（5）Analysis of the expression of grape VviBZR gene family after water stress treatment.Compared with normal growth control,the VviBZR1 and VviBZR2 genes in grape leaves were significantly up-regulated after PEG treatment for 12 h,while the expression levels of VviBZR4,VviBZR5 and VviBZR8 genes were increased to some extent in the late PEG treatment（after 48 h）.Except VviBZR1,grape BZR TFs showed differential expression after exogenous BR treatment,indicating that these genes may play an important role in grape resistance to water stress.