苏云金芽孢杆菌杀虫晶体蛋白基因cry1A的克隆及生物信息学分析

Molecular Cloning and Sequence Analysis of cry1A Genes from Bacillus thuringiensis

以分离的3株苏云金芽孢杆菌质粒为模板,通过设计一对特异引物,PCR扩增得到3条全长3.6 kb的基因序列,其各自都包含一个完整的开放阅读框,编码序列全长分别是3468、3450、2697 bp(Genebank登录号分别为GQ385073,GQ385074,GQ385075);它们与cry1A类基因同源性都高达95%以上;分别编码1159、1150和902个氨基酸。并在此基础上采用生物信息学方法对这3个基因编码蛋白从氨基酸组成、理化性质、跨膜结构域、疏水性/亲水性、亚细胞定位、跨膜结构域以及功能域等方面进行了预测和分析。为转基因抗虫植物和微生物杀虫工程菌的构建提供了基因来源。

Specific primers,which were synthesized according to the conservative regions of crylA gene, were used to amplify genomic DNA from three Bacillus thuringiensis （Bt） strains sreened by polymerase-chain reaction （PCR）. Three sequences of crylA gene with whole length of 3.6kb was obtained ,which contained a complete open reading frame. These three full-length coding sequences were 3468,3450 and 2697 bp （ GenBank Accession Number GQ385073, GQ385074, GQ385075 ）. Nucleotide sequencing showed that the sequences for these three genes had more than 95 % identical to the known crylA gene,which encoded 1159,962 and 898 amino acids respectively. Some characters of these three genes that could encode amino acids were analyzed and predicted by the tools of bioinfonnatics in the following aspects,including the composition of amino acid sequences, hydrophobicity or hydrephilicity, membrane spaning domain and tertiary structure of protein and function. It provided new selection for development of novel Bt products based on its toxins.