High-Throughput Screening Verticillium Wilt Resistance Related Gene By VIGS Technology In Cotton

Cotton is one of the world's most important economic crops and the main source of natural fiber,China is one of the world's major cotton production countries.Sustained and stable cotton production has important significance for the economic and social development in China.Verticillium wilt in cotton caused by the soil-borne fungal pathogen Verticillium dahliae is a vascular disease,which results in serious loss in yield and fiber quality yearly.However,traditional strategy doesn't work very well to control the disease but the local control,and it isn't an efficient way to develop a new cotton variety with resistances to Verticillium wilt via the conventional memod,and there are few successful reports with conventional breeding memod for disease resistance improvement due to shortage of high resistance germplasm,so the control of this disease has been an obstacle in cotton production.Currently,more and more research focused on disease resistance mechanism of cotton to Verticillium wilt.Thus,now a lot of studies still focus on unco,vering the molecular mechanism of cotton resistance to verticillium wilt,and try to improve cotton resistance to cotton Verticillium wilt.Because of the limitation of research materials and the lack of research techniques,it is difficult to explore the molecular mechanism of cotton resistance to verticillium wilt at the genetic level.Virus-induced gene silencing(VIGS),based on loss-of-function concept to research gene function in plant,which provides a new opportunity to reveal genes' functions that participate in the cotton-Verticillium dahliae interaction.The cotton(Gossypium hirsutum)strain TM-1 and H7124 was used in this study,which was breeded by the NanJing agricultural university,where is the Chinese key laboraty.On the basis of the results of RNA-Sequence and gene analysis,we chose 19 genes which possibly contact with the resistantion to verticillium wilt of cotton.Then,we silence these genes through VIGS technology to realize their function.The results of we found that two genes might be related to the resistance to Verticillium wilt,the AUXIN is a negative control gene,it can enhance the resistance to Verticillium wilt when we silence the gene in TM-1.While the LRR is a positive gene,it can weaken the resistance to Verticillium wilt when we silence the gene in H7124.To further demonstrate whether the selected gene is related to resistance,it was transformed in Arabidopsis by agrobacterium-mediated floral-dip technology and overexpressed driven by 35S promoter.Transgenic lines were screened by Kanamycin resistance and then verified by PCR and RT-PCR.Verticillium dahliae is inoculated to transgenic Arabidopsis,which showed that transgenic Arabidopsis are more sensitive to V.dahliae infection.Fouthermore,we found that the transgenic Arabidopsis can promote the plant flowering.