条锈菌几丁质脱乙酰基酶基因的沉默与提高小麦抗性的关系研究

Wheat stipe rust is a widespread disease in China and causes huge yield losses in wheat production. Currently, the situation of controlling rust epidemics is very urgent in China. Breeding of resistant wheat cultivars carrying genes conferring resistace to wheat stripe rust is an effective strategy to relieve rust damage and ensure the food security. However, it is not an efficient solution to combat rust epidemics due to several limitation. For example, those major resistance genes could be easily defeated by new races of rust due to the rapid evolution of its pathogenicity. Pyramiding several minor resistance genes or QTLs in one cultivar helps to broaden plant resistance spectrum and realize durable resistance, however, currently gene pyramiding could not be performed with enough efficiency because only few functional markers are avilable. Therefore, developing novel strategies is of great importance for wheat breeding and the rust control. Chitin deacetylase is an enzyme that catalyzes the hydrolysis of acetamido groups of N-acetylglucosamine in chitin, promoting the conversion to chitosan. This conversion affects not only the important structues, such as hyphae and haustorium of the fungi, but also the interactions between the fungi and the host plants. In this study, we have annotated the CDA genes in the rust genomes. Studies of gene transcription revealed different expression patterns of the CDA genes, and sequence alignments showed that the CDA gene memebers shared very low similarity. These results suggest different CDA genes probably have distinct functions or mechanisms in wheat-rust interaction. We have constructed RNAi vectors for several CDA genes and obtained transgenic plants of common wheat. Based on these works, we will further innoculate the transgenic wheat with stripe rust isolate, and evaluate the efficiency of gene silencing for CDA genes via the Host Induced Gene Silencing (HIGS), and study the correlation between silencing of CDA genes and the improvement of wheat stripe rust resistance. The results will contribute to understanding the function of CDA genes in the wheat-rust interaction and to developing new strategies for stripe rust resistance.

小麦条锈病在我国普遍发生，严重威胁小麦安全，防治形势十分严峻。利用抗病基因培育抗病品种，是防治条锈病的有效措施。然而，主效抗性基因很容易被快速变异的条锈菌生理小种所攻克；而微效的慢条锈菌基因，由于目前缺乏功能标记又难以实现分子聚合。探索新的防治策略及育种方法，对小麦生产意义重大。真菌几丁质脱乙酰基酶（CDA）在侵染过程中，催化几丁质转变为壳聚糖，不但影响菌丝、吸器等重要结构的发育，而且影响条锈菌与寄主的互作。本研究对小麦条锈菌的CDA基因进行了注释，序列差异及表达模式不同表明它们可能存在不同的功能或作用方式。已利用部分基因构建了RNAi载体并获得了转基因小麦。在此基础上，将对转基因后代进行接种处理，研究寄主诱导的转基因沉默（HIGS）对CDA基因的沉默效率，及其对提高小麦抗条锈病能力的影响。研究结果对于理解CDA基因在小麦-条锈菌互作中的作用及探索小麦条锈病防治新策略具有重要指导意义。

小麦条锈病是威胁小麦生产的重要病害，深入研究条锈菌侵染致病关键基因以及深入理解小麦-条锈菌互作机理，对于探索小麦条锈病防治策略、指导小麦抗病育种具有重要意义。本课题围绕小麦条锈菌生长发育及接触寄主的重要结构—细胞壁，针对细胞壁合成相关基因几丁质脱乙酰基酶（CDA），从基因注释、基因表达、转基因功能验证等方面开展了研究。共鉴定20个CDA基因家族成员，可归为3个分支，其中一个分支内（11个基因）存在多个基因重复。各CDA基因的表达模式类似，均在侵染后上调表达至1-2天后达到峰值，随后下调至最低点并在侵染后期再次上调表达。利用8个CDA基因构建了RNAi沉默表达载体，并获得了普通小麦的稳定转基因后代。接种小麦条锈菌后，转基因小麦叶片中条锈菌靶标基因的相对表达量显著降低，表型鉴定为高抗小麦条锈病，而且T2、T3世代表型稳定，均表现高抗条锈病。研究结果不但表明了寄主诱导的转基因沉默可有效应用于小麦-条锈菌互作研究，而且表明条锈菌细胞壁合成基因在条锈菌侵染致病过程中发挥重要作用。研究结果将有助于促进条锈菌关键基因的功能验证及小麦-条锈菌互作机制的深入研究。