Functional Analysis Of RxLR31197 Effect Factor Of Plasmopara Viticola

Grape downy mildew is one of the diseases that seriously damage grape growth.Downy grape can secrete Rx LR effect factors into plant cells and have physiological and biochemical reactions,which can effectively promote growth and reproduction and lead to host disease.These effects factors inhibit plant immune response and play a toxic role in host cells.Based on screening the molecular function of grape downy mildew effector factors previously,we selected the RxLR31197 effect factor to analyze.First,we cloned the RxLR31197 gene and constructed the related expression vector and carried out bioinformatics analysis and expression pattern analysis.We also used mouse Pro apoptotic protein Bax as elicitor to analyze its inhibitory effect on Bax induced allergic cell death and the function of inducing tobacco cell death.At the same time,the subcellular localization of it was analyzed.Then RxLR31197 was used as bait to screen the interaction proteins in the cDNA library of the wild yew grape "Liuba-8" leaf yeast.Finally,a stable transformation model of tobacco was carried out,and the disease resistance of transgenic plants was analyzed.The following results are as follows:1.We cloned RxLR31197 gene sequence and predicted secondary structure with 8 alpha helix and 1 beta folding.Multiple sequence alignment and phylogenetic tree analysis of Rx LR31197 protein sequences with other known avirulent proteins were performed and found that the genetic relationship between the RxLR31197 and Avr4 of Phytophthora infestans recently,studies showed that Phytophthora infestans Avr4 play an important role in R4 mediated resistance,speculation RxLR31197 may Participate in resistance response.2.Analysis of expression patterns,induction of lethal and inhibitory lethality,and subcellular localization analysis revealed that: RxLR31197 was induced up-regulated at the early stage of infection,which could inhibit the production of PTI(PAMP-triggered immunity)to the pathogen identification;RxLR31197 had a partial inhibition of Bax induced PCD,and the subcellular localization showed that RxLR31197 located in the nucleus and cytoplasm..3.The yeast two hybrid system was used to screen the interaction protein of grape downy mildew factor Rx LR31197 in Chinese wild grapes.Interacted proteins are: Vitis vinifera vesicle-associated protein,chaperone protein ClpB3,NACdomain-containing protein,tRNA deaminase.4.By tobacco stable genetic transformation technique for obtaining T2 transgenic plants,the transgenic resistance function analysis indicated that it has the ability to inhibit the bacteria infection.