斜纹夜蛾取食诱导棉花抑制性消减文库的初步构建及生物信息学分析

摘要

To explore insect resistant genes induced by Spodoptera litura (Fabricius) larvae from cotton, forward and reverse suppression subtractive hybridization(SSH) libraries were constructed. 1000 clones were selected from each library, the inserted fragment sizes ranged between 200 bp and 1000 bp, which meet the requirements of a plant cDNA library. 250 positive clones from the forward library were screened by using the Reverse Northern Dot-blotting technique, then sequenced and analyzed, and 35 expressed sequence Tags (ESTs) were obtained, in which 24 ESTs were obtained from the positive clone in reverse library. Based on BLASTx in GenBank analysis, it was found that those ESTs make important roles in signaling pathway regulation, gene regulation, anti-stress ability, defense responses, energy synthesis and metabolism, proliferation of cells, and so on. Those genes play an important role in regulating and defending against S.litura larvae directly and indirectly.%为了筛选棉花抗逆基因,实验室构建了棉花被斜纹夜蛾幼虫取食诱导的正、反向抑制性消减文库.文库随机各挑取1000个克隆进行PCR鉴定,发现插入片段长度主要集中在200～1000 bp之间.经反向Northern杂交筛选,正向文库随机挑取250个克隆测序,得到了35条非重复序列ESTs,反向挑取150个克隆,得到24条ESTs.GenBank数据库BLAST序列相似性比对功能分析表明,斜纹夜蛾幼虫取食胁迫棉花的应答过程涉及信号传导、基因调控、抗胁迫、防御反应、能量合成代谢、细胞生长延伸等多个生物途径.诱导表达的基因对斜纹夜蛾取食胁迫功能表现为一定的直接或间接保护作用.