Investigation And Studies On Resistance To White-rot Disease Of Chinese Wild Grape

The grape is one of the most widely cultivation fruit tree in our country. The total yield of grape is the first of the world in2010. Meanwhile, the grape disease occurrence and harm associated with the development of the grape industry. The white rot disease is caused by fungus, which is one of the serious disease in grape. The grape can be suffered fatal effect by white rot disease. How to improve the grape resistance to white rot disease is the key problem. There is abundant resistance and excellent agronomic characters in Chinese wild grape, Chinese wild grape have strong resistance to cold hardiness, drought resistance, disease resistance and better resistance to damp and hot. In this study, we use Chinese wild grape as research object. The purpose is mining Chinese wild grape resistance resources to white rot disease, cloning its resistance gene.First, using field investigation combine with artificial inoculation methods, system for evaluation Chinese wild grape white rot disease resistance. Investigations in the field showed that our country wild grape berries are not infected white rot disease in the condition of nature. In the artificial inoculation conditions, Chinese wild grape show differences resistance in the same variety or in different variety. It was found that within the same variety exist resistant and susceptible varieties. The investigation discovered that Tangwei grape and Amur grape exhibits the character of high resistance to white rot disease, but Yanshan grape and HuNan brier grape exhibits the character of high sensitive. It was found that within the same intraspecific exist resistant and susceptible varieties.Secondly, based on investigation, choose Vitis davidit as research materials, using the homologous cloning method to get resistance gene analogs (RGA), and through the fluorescence quantitative PCR analysis the relationship between these genes and white rot disease. According to the NBS-LRR kind of resistance gene conserved region design primer, extract DNA and RNA from resistance materials, through PCR amplification and screening, we get10RGAs, these10RGAs coded amino acid sequence are all contain four conservative structure of NBS domain. That is P-loop (GGVGKTT)、Kinase-2a (VLDDVW/D)、Kinase-3a (GSR/KILVTTR) and HD (GLPLAL). These10RGAs, six fragments isolated from DNA (NB-1、NB-3、NB-5、NB-7、NB-11and NB-12), four fragments isolated from cDNA (NBS-6、NBS-4、NBS-13and NBS-16). RGAs coding amino acids and four known R genes the amino acid sequence homology is20.8%-46.2%. The homology of NB7gene and Arabidopsis RPS5gene is46.2%. Furthermore, ten of grape resistance gene analogs show rich polymorphism in the level of the homology amino acid, the homology distribution between23.5%-98.8%. Further analysis found that among10RGAs, four belong to non-TIR-NBS-LRR kind of resistance genes, six belong to TIR-NBS-LRR kind of resistance genes. After NCBI matching, these RGAs are highly homology with assume disease resistant protein mRNA, which has positioned in grape chromosome number of5,9,13,18and19. Fluorescence quantitative analysis show that NB7gene is induced by white-rot fungus, indicate that NB7gene may as potential the white rot disease resistant gene in grape, which can lay the foundation for final cloning grape white rot disease resistant gene. The exact function needs to be further study.Through research, we get the main conclusion that Chinese wild grape berries are not infected white rot disease in the condition of nature. In the vaccination condition, Chinese wild grape show differences resistance in the same variety or in different variety. By using molecular techniques for research, from resistant materials Vitis davidii, we get10RGAs in which NB7gene is induced by white rot fungus, which provides a theoretical basis for getting white rot disease resistant genes.