| Verticillium wilt is a soil-borne vascular disease,the pathogenic fungus,Verticillium dahliae,is a semi-viviparous trophic funguswhich can cause the leaves to wither and fall off;The browning of vascular bundles has a very serious impact on cotton production.Therefore,it is of great significance to analyze the immune response mechanism of cotton to verticillium wilt and identify the key disease resistance genes to serve the cotton disease resistance molecular breeding for the prevention and control of cotton verticillium wilt.Zhongzhimian NO.2 is the first cotton variety with verticillium wilt resistance approved by the National Variety Approval Committee in China,with outstanding resistance performance.It is still a control variety in regional production trials for breeding new cotton varieties with verticillium wilt resistance.However,at present,the resistance related genes and resistance mechanisms of Zhongzhimian NO.2 have not been fully analyzed.In view of the extremely strong resistance of this variety to verticillium wilt,based on the genome of Zhongzhimian NO.2this study screened and identified the resistance genes of Zhongzhimian NO.2 through methods such as transcriptome analysis,genetic transformation in Arabidopsis thaliana,and resistance identification.The following results were obtained:1.Based on the previous work in the laboratory,the genome and transcriptome data were analyzed and compared with Junmian NO.1(susceptible cotton species).A total of 43 candidate disease resistance genes were screened,which were continuously up-regulated in Zhongzhimian NO.2 and down-regulated in Junmian NO.1 during the infection of Verticillium dahliae.2.Gene cloning of 43 candidate resistance genes,a total of 18 candidate genes were cloned and p CAMBIA1300-35 S overexpression vector was constructed3.A total of 10 transgenic materials including Gh-Vd-PR03,Gh-Vd-PR09,Gh-Vd-PR11,Gh-Vd-PR12,Gh-Vd-PR12,Gh-Vd-PR19,Gh-Vd-PR20,Gh-Vd-PR21,Gh-Vd-PR29,Gh-VdPR34,Gh-Vd-PR39 were obtained by agrobacterium transformation.4..A total of four of these transgenic materials,Gh-Vd-PR12,Gh-Vd-PR29,Gh-Vd-PR34,and Gh-Vd-PR39,were characterized for resistance.The overexpression of Gh-Vd-PR12,GhVd-PR29,Gh-Vd-PR34,and Gh-Vd-PR39 was found to significantly increase Arabidopsis thaliana resistance to V.dahliae;and the reproduction of the pathogen was significantly decreased in the transgenic strain compared with wild-type Col-0.5.RT-q PCR analysis revealed that the expression of jasmonic acid and ethylene signaling factors in Arabidopsis thaliana was upregulated after overexpression of Gh-Vd-PR34.It suggests that Gh-Vd-PR34 mediates plant resistance to verticillium wilt through jasmonic acid and ethylene signaling.In this study,we aimed to identify the resistance genes in this cotton species and transfer them into Arabidopsis thaliana for preliminary functional characterization,so as to provide possible candidate genes for the genetic engineering of cotton resistance to verticillium wilt,. |
