Identification Of Trihelix Transcription Factor And Study On The Function Of Grape VvTrihelix5 In Response To Salt

Grape is one of important economic fruits in China,and its cultivation area and annual yeild are among the top in the world.However,due to environmental and soil problems,viticulture is currently facing more serious problems.Among them,soil salinization has become one of important factors restricting viticulture.Genetic engineering is an effective breeding method,which can greatly shorten the breeding period.Therefore,screening salt-tolerant genes and studying their functions and mechanisms are great significance to improve salt-tolerant of grape.In this study,through bioinformatics identification and q RT-PCR analysis showed that the VvTrihelix5 transcription factor has obvious response to salt stress in grape.However,the research on the response mechanism under salt stress of this transcription factor is still unclear.Therefore,this experiment acquired a VvTrihelix5 transcription factor through gene cloned,and the sequence characteristics of the transcription factor and the mechanism of its response to salt stress were analyzed.The main research results were as follows:1 The bioinformatics analysis of the Trihelix transcription factor family of grape showed that 27 members are distributed on 11 chromosomes.The prediction of subcellular location shows that all transcription factors are mainly located in the nucleus;the results of the phylogenetic tree analysis indicate that this family can be divided into five sub-families.The results of cis-acting elements show that 2000 bp in front of the promoters contain numerous stress-related elements,including methyl jasmonate,abscisic acid,anaerobic inducing elements,and so on,respectively.q RTPCR finds that the expression levels of VvTrihelix5,VvTrihelix6,and VvTrihelix7 transcription factors were significantly up-regulated under 200 mmol/L NaCl stress,which were 7.3,15.4,and 7.2 times compared with the control,respectively.In addition,this family of transcription factors are also induced by ABA and PEG stress.2 The subcellular localization of the Trihelix5-GFP fusion protein in Arabidopsis protoplasts show that it is mainly localized in the nucleus,and the VvTrihelix5 transcription factor belongs to the SIP1 subfamily;multiple sequence alignments and conservation of nine species find that Motif 1,Motif 2,and Motif 4 contain the entire functional domain of transcription factors,namely the helix-loop-helix structure.3 After heterelogous expression of the grape VvTrihelix5 transcription factor in Micro-Tom tomatoes,wild-type and transgenic tomatoes are treated with 300 mmol/L NaCl,respectively.Phenotypic observation showed that the leaves of wild-type tomatoes are obviously curled after treatment for 144 h.The measurement results of the relative conductivity,root activity,and antioxidant enzyme activity index of tomatoes in different treatment periods show that the VvTrihelix5 transcription factor may increase the tolerance of tomatoes to resist salt stress by activating the activity of the antioxidant enzyme system.Yeast two-hybrid screening results show that VvTrihelix5 and VvHAL3 A protein have significant interaction relationship.The analysis of bimolecular fluorescence complementation(Bi FC)find that the interaction position of these two proteins may be located at the cell membrane.