中国野葡萄抗黑痘病分子机理的研究

葡萄黑痘病是危害欧洲葡萄重要的真菌病害之一。中国野葡萄不仅抗病，而且和欧洲葡萄杂交亲和性好，是葡萄抗病育种重要的种质材料。因而，克隆中国野葡萄抗病基因，进行抗病分子机理研究，具有重要的育种价值。本项目在前期研究基础上，以抗病中国野葡萄商-24、感病的红地球及其种间杂交F1代为研究对象，进行商-24与红地球在黑痘菌侵染后不同时期的转录组研究，筛选出黑痘菌诱导下特异表达和增强表达基因片段，明确抗病过程中参与的代谢途径；采用qRT-PCR进行商-24、红地球及其杂种后代抗病、感病植株在黑痘菌及信号分子处理下的抗病基因表达分析，并结合Affymetrix芯片数据表达分析，获得抗黑痘病新基因；构建抗黑痘病新基因过表达载体和RNAi干扰载体，转化商-24和拟南芥，通过过表达、RNAi干扰及未转化葡萄或拟南芥的抗病性表现及基因表达分析，研究基因功能，分析抗黑痘病分子机理，为葡萄抗病育种提供理论依据。

Anthracnose, caused by the biotrophic fungus Elsinoe ampelina (de Bary) Shear, is one of the most devastating disease of grapevine (Vitis vinifera L.) prevalent in warm and humid regions of the world. European grapvine is one of the finest grapes grown in the world both for wine and table purposes, but the prevalence of anthracnose is a major risk to grpae production. As Chinese wild Vitis species are more anthracnose tolerant than European cultivars and the inheritance of the resistance to anthracnose to offspring is very strong, it turns out to be the important germplasm resources for developing resistant cultivars. Thus, cloning genes related to E. ampelina-resistance and studying the mechanisms of anthracnose resistance are of important breeding value. Based on our previous studies, Chinese wild V. quinquangularis clone 'Shang-24' which is highly resistant to E. ampelina, and V. vinifera cv 'Red Globe' which is susceptible to anthracnose, and the interspecific hybrid F1 generation, will be used as materials in this study. RNA-Seq, qRT-PCR, Affymetrix microarry data bioinformatic analysis and genetic transformation techniques, will be used to study the transcriptomics and molecular mechanism during Chinese wild Vitis species-Elsinoe ampelina interaction. We aim to isolate target genes related to anthracnose resistance, further identify the resistance relevant metabolic pathways.The full length cDNA sequence of new genes resistant to anthracnose will be isolated by rapid amplification of cDNA ends (RACE)or RT-PCR technique. The over-expression vecator and RNA interference (RNAi) vecator of new genes will be constructed and will be successfully transferred into Arabidopsis thaliana and Chinese wild V. quinquangularis Shang-24 via Agrobacterium tumefaciens mediated transformation method.Phynotype investigation and anthracnose disease resistance identification of different transgenic plants will carried out in field conditions.Information obtained from this study will enhance the understanding of the resistance mechanisms and resistant breeding in grapevine plants to E. ampelina infection.