采用逆转录PCR方法克隆获得1个虫草棒束孢类枯草杆菌蛋白酶基因的开放阅读框(ORF)序列,并对其进行生物信息学分析。该基因的开放阅读框全长1599bp,编码的蛋白质由532个氨基酸组成(包括18个氨基酸的信号肽)。预测蛋白质的等电点和分子量大小分别为6.256、56.936ku。进行序列分析和进化树构建,发现该蛋白酶与已知的几个种类枯草杆菌蛋白酶具有很高的同源性,且与虎甲寄生菌的粉拟青霉位于同一进化分支。实验首次鉴定了虫草棒束孢类枯草杆菌蛋白酶基因的编码区序列,为进一步研究虫草棒束孢致死蝙蝠蛾幼虫的作用机理提供理论依据。
In the present study, the coding cDNA sequence oflsariafarinosa subtilisin-like protease ( IsSUB ) was cloned by reverse transcription PCR and analyzed by bioinformatieal tools in detail. The subtilisin contains 1 599 nucleotides with a putative open reading frame ( ORF ) encoding 532 amino acid residues ( with a signal peptide of 18 amino acid residues ) . The isoelectric point ( pI ) and predicted molecular weight of IsSUB were 6.256 and 56.936 kDa, respectively. Amino acid sequence and phylogenetic tree analysis among L farinosa and other insects represented that these fungal subtilisin had a high homology, and IsSUB existed in one evolutionary branching with Cicindelidac sp. Paecilomyces farinosus. It was concluded that the ORF of subtilisin-like protease from L farinosa provided theoretical basis for damaging on Hepialus larvae body wall.