抑制差减杂交分离赤霉病菌诱导的小麦特异表达基因

Isolation of Genes Involved in the Response of Wheat to Fusarium graminearum by Means of Suppression Subtractive Hybridization

为了全面探索小麦赤霉病抗性机理，以抗赤霉病小麦品种‘苏麦3号’及其感病的近等基因系（Hw04）为实验材料，构建了一个‘苏麦3号’受赤霉病菌诱导表达的正向差减杂交文库。随机选取了141个阳性克隆测序，共获得133条通读EST。将获得的EST去除载体及接头序列后，利用CAP3软件进行聚类分析，133条EST被聚成90个序列重叠群（contigs），片段大小介于106-643bp间，平均长度为274bp。利用NCBI的Blastx软件对序列进行蛋白序列同源性比对，结果显示76条序列在蛋白数据库中可以找到同源序列，功能涉及能量代谢、物质代谢、疾病／防御、转录及细胞结构等。其中以能量及物质代谢相关的基因数量最多，分别占总EST的21．1％及17．1％；其次是与抗病及防御反应有关的EST，数量占总EST的15．8％。进一步的分析表明与抗病及防御相关的EST功能主要涉及抗氧化、细胞解毒及相关物质的代谢。

Wheat cultivar, ＇Sumai 3＇ with a good FHB resistance was widely used all over the world. Currently,the molecular mechanisms of resistance in wheat to FHB had been focused on inheritance law and quantitative trait locus （QTL） mapping. In order to investigate the genes related to FHB resistance,a forward subtractive cDNA library was constructed using F. graminearum infected spikes of ＇Sumai 3 ＇ and its susceptible near-isogenic line as tester and driver, respectively. A total of 141 positive clones were picked randomly and sequenced and 133 of them had good quality. Based on the clustering result,the 133 ESTs could be grouped into 90 non-redundant sequences. The size of the ESTs ranged from 106 to 643 bp with mean value 274 bp. Seventy-six of the ESTs showed similarity to known sequences in GenBank,the other fourteen ESTs seemed to be novel sequences. The functions of the ESTs involved in metabolism, energy, disease/defence,transcription and cell structure and so on. The ESTs associated to energy and metabolism accounted for 21.1% and 17.1%, respectively,which numbers were the most of the ESTs. Then the other major ESTs were ascribed to disease/defence system. Further analysis showed that the putative stress-related genes were mainly involved in anti-oxidation,detoxification and metabolism of disease resistance sub-stances.