Functional Verification Of Cotton GhIQM1,GhNAC90 And GhBsr-d1 Genes In Resistance To Verticillium Wilt

Verticillium wilt is one of the most destructive diseases that endanger cotton production,posing a major threat to cotton yield and quality.Due to the long cycle,high cost and low selection efficiency of traditional breeding methods,the process of cotton breeding for resistance to Verticillium wilt is slow.Therefore,it is particularly important to discover cotton disease resistance related genes and reveal the molecular mechanism of disease resistance genes regulating cotton verticillium wilt.In this study,the GhIQM1 and GhNAC90 genes related to Verticillium wilt were cloned and three homologous genes of Gh Bsr-d1 Ls were cloned in cotton by homologous comparison with rice Bsr-d1.Fluorescence quantitative PCR was used to analyze the expression levels of these genes in different cotton tissues and after inoculation with Verticillium dahliae.VIGS technology was used to silence the 5 genes to identify their disease resistance functions.Two disease resistance related genes,GhIQM1 and GhNAC90,were screened out.The two genes,GhIQM1 and GhNAC90,were transferred into Arabidopsis thaliana through heterologous overexpression to further verify their disease resistance.The results of the study are as follows:1.The analysis of expression patterns showed that GhIQM1 and GhNAC90 had the highest expression levels in roots and leaves,respectively,and Gh Bsr-d1 Ls genes were differentially expressed in different tissues.Through the induction treatment of Verticillium dahliae,the results showed that GhIQM1 and GhNAC90 were significantly up-regulated in response to the invasion of Verticillium dahliae after the stress of Verticillium dahliae.Among the remaining three candidate genes,Gh Bsr-d1L2 gene and Gh Bsrd1L5 gene quickly responded to the induced expression of Verticillium dahliae at the initial stage of inoculation.The expression level of Gh Bsr-d1L7 gene after vaccination showed a downward trend first and then an upward trend.It shows that these genes are induced by Verticillium dahliae to varying degrees.2.The VIGS vector of candidate genes was constructed to verify the disease resistance of Verticillium wilt.The silencing gene plants were obtained from the cotton’TM-1’ material.After inoculation with Verticillium dahliae,through phenotypic observation and disease index analysis,stubbing detection and stem recovery experiments were finally screened out of 3 plants that were compatible with the control group.The significantly different candidate genes are the negative regulator of GhIQM1 and the two positive regulators of GhNAC90 and Gh Bsr-d1L7.3.The GhIQM1 and GhNAC90 genes were screened out from the above 5 candidate genes to verify their disease resistance in the ‘ XLZ33 ’ material.The results showed that in both susceptible and resistant varieties,inhibiting the expression of GhIQM1 showed enhanced resistance to Verticillium wilt in cotton.In addition,GhIQM1 regulates the expression of disease resistance genes downstream of the SA/JA signaling pathway,Resistance of cotton to Verticillium wilt.Overexpression of disease resistance function analysis showed that overexpression of GhIQM1 weakened the resistance of Arabidopsis to Verticillium wilt.Therefore,GhIQM1 acts as a negative regulator in the process of resistance to Verticillium wilt.While silencing the GhNAC90 gene plays a positive regulatory role in the resistance of cotton to Verticillium wilt,overexpression of GhNAC90 exhibits a negative regulatory effect in Arabidopsis,which is inconsistent with the results of VIGS.The reason may be that the overexpression exceeds this.The threshold of gene regulation has led to a more susceptible phenotype,but its specific regulation mechanism remains to be explored in depth.