Cloning And Functional Analysis Of A VaERF20 Transcription Factor In Chinese Wild Vitis Amurensis

Grape can be attacked by various pathogens during growth and development period,such as Botrytis cinerea,a kind of necrotrophic fungi with strong reproductive ability,which can seriously affect the normal growth of grape leaves and fruits,and also harm the development of grape plants in the second year.Ethylene response factor(ERF)transcription factors play important roles in plant defense fungi and bacterial infection.However,the function of grape ERF transcription factor is still not clear.Research group conducted RNA-Seq transcriptomic profiles of the highly resistant V.amurensis acc.'Shuangyou' and the susceptible V.vinifera cv.'Red Globe' after induced by B.cinerea.VaERF20 was selected according to differential expression induced by B.cinerea.In this study,VaERF20 was cloned from the Chinese wild V.amurensis acc.'Shuangyou',and transformed to the model plant Arabidopsis thaliana for explore the function and the mechanism.The main results are as follows.1.The expression of VaERF20 in the leaves of Chinese wild V.amurensis acc.'Shuangyou' was analyzed by real-time PCR.The results showed that VaERF20 was induced by B.cinerea and the expression level peaked 4h after treatment;Characterized VaERF20 from the Chinese wild Vitis genotype,V.amurensis acc 'Shuangyou'.The complete open reading frame length of VaERF20 is 459 bp which encodes a deduced protein of 153 amino acids.Chinese wild V.amurensis acc.“Shuangyou” ERF20 shows 84.53% sequence homology to V.vinifera cv.PN40024 and the protein sequence shared 85.06% identity.2.Overexpression vector 'pCAMBIA2300-35S-VaERF20' was constructed and then transformed to the model plant A.thaliana.Inoculate both A.thaliana plants and wild type(Col-0)with B.cinerea and the bacterium Pseudomonas syringae pv.tomato(PstDC3000),the result showed A.thaliana plants over-expressing VaERF20 displayed enhanced resistance to B.cinerea and PstDC3000.Patterns of pathogen-induced reactive oxygen species(ROS)accumulation were entirely distinct in B.cinerea and PstDC3000 inoculated plants.Examples of both salicylic acid(SA)and jasmonic acid/ethylene(JA/ET)responsive defense genes were up-regulated after B.cinerea and PstDC3000 inoculation of the VaERF20-overexpressing transgenic A.thaliana plants.After threatment Pst DC3000,flg22 and LPS,the transgenic A.thaliana leaves were accumulated more callose than the Col-0 plant as well as the stomatal closure was greater than that of Col-0.