R Gene Deployment Between Plant Families Facilitated By SRNA-Guided Gene Silencing

Plant endogenous sRNA(small RNA)are important for regulating of gene expression and have a variety of biological functions by activating and inhibiting the expression of specific genes,especially in the regulatory mechanism of host-pathogen interactions in plants.In the process of long-term interactions between plants and pathogens,plants evolve two basic immune systems: PAMP(Pathogen-associated molecular pattern)-triggered immunity and effector-triggered immunity.Pathogen effector has the features of fastevolving and great diversity,and plants evolve a large number of R genes to deal with the challenges from different pathogenic microorganisms.The R genes in the plant are strictly controlled,and the excessive expression of R genes in transgenic plants tends to cause abnormal growth.Small RNA(sRNA)is an important regulator in the whole life process of plants,and regulates gene expression at transcription,post-transcription or translation level.It also involves in the regulation of plant-pathogen ineraction.So we want to construct vectors of artificial miRNA targeting R gene to reduce the expression of R gene.When pathogens invade plants,they will reduce the expression of sRNA,to promote their infectivity,and destroy sRNA regulation mechanism of the plants.Thus mRNA transcribed by R gene can be translated,and express with a high abundance,which achieve the purpose of disease resistance in the end.In this study,Arabidopsis and Tobacco were used as research materials,and the Tobacco Mosaic Virus resistant N gene and Pseudomonas syringae resistant RPS4 gene were used as the research object.We try to use small RNA-mediated gene silencing to realize the use of disease resistant genes between two species.The main findings are as follows:1.The expression vector of amiR6019 was successfully constructed and transferred into Arabidopsis thaliana Col-0.On the basis of transgenic amiR6019 Arabidopsis plants,the N gene was successfully transferred.This result provides a new approach to study disease resistance: using R silencer to promote the interspecific R gene utilization.2.Four expression vectors of artificial microRNA targeting RPS4 were designed and constructed,and four transgenic tobacco plants were obtained.The expression vectors of RPS4,RRS1 and avrRPS4 were constructed and verified by transient expression.The optimum concentration of Pseudomonas syringae pv tomato DC3000 of infecting SR1(Nicotiana tabacum)was found.These results laid the foundation for the future study of the interaction between RPS4 and Pseudomonas syringae in tobacco.