苏云金芽胞杆菌WZ-9菌株对马铃薯瓢虫的毒力及其杀虫基因的研究  

Identification of cry Gene and Its Toxicity from Bacillus thuringiensis Strain WZ-9 against Henosepilachna vigintioctomaculata

作　　者：宋萍[1] 王勤英[1] 吴会贤[1] 陆秀君[1] 王永[2] 

机构地区：[1]河北农业大学植物保护学院河北省农业病虫害生物防治工程技术研究中心,保定071001 [2]天津农业科学院中心实验室,天津300192

出　　处：《农业生物技术学报》2008年第3期515-520,共6页Journal of Agricultural Biotechnology

基　　金：河北省自然科学基金项目(No.303213);保定市科技局项目(No.05S001);河北农业大学发展基金资助

摘　　要：苏云金芽胞杆菌(Bacillus thuringiensis)WZ-9是从土壤中分离的对马铃薯瓢虫(Henosepilachna vigintioctomaculata)的幼虫有特异杀虫活性的新菌株。实验对WZ-9菌株的形态特征、生长特性、生物活性、基因型和蛋白型等进行了研究。结果表明,WZ-9菌株可产生菱形伴胞晶体,SDS-PAGE检测表达的主要蛋白条带分子量约为130kD;液体培养的生长周期是24h,随菌株的生长培养基pH发生较大波动,在潜伏期和稳定期,pH变化较小,在对数生长期pH呈先迅速下降、再缓慢回升的变化趋势;生物活性测定表明,WZ-9菌对马铃薯瓢虫2龄幼虫72h校正死亡率达100%,LC50为2.95×107细胞/mL;基因型鉴定表明,WZ-9菌株含有cry7基因,利用PCR扩增方法获得了1条总长为3781bp基因序列,其中包含了1个3414bp开放读码框,其编码蛋白由1138个氨基酸残基组成,与Cry7Ab2具有99.65%序列同源性,存在4个氨基酸差异,亲缘关系最近,被Bt杀虫晶体蛋白命名委员会命名为Cry7Ab3(登录号为BI1015188)。A high toxicity Bacillus thuringiensis （Bt） strain WZ-9 against larvae of Henosepilachna viginfioctomaculata was isolated from soil in Hebei, and bipyramidal inclusions could be observed in it by scanning electron microscopy. SDS-PAGE analysis indicated that the strain produced a 130 kD protein, growth cycle of this becteria was 24 h, and pH of the culture showed big fluctuation with the cell growth. In lag phase and stationary phase, pH varied less than that in exponential phase. The bioassay results showed that the WZ-9 strain was highly toxic to the 2 instar larvae ofH. viginfioctomaculata. Corrected mortality was 100% at 72 h, and LC50 was 2.95 × 107cell/mL. Genotype identify showed that this strain had cry7 gene. Sequence analysis showed that the encoding gene contained a ORF of 3 414 bp and encoded 1 138 amino acids residues. The deduced amino acid sequence was 99.65% identical to that of the reported Cry7Ab2 sequences. This gene was designated by Bt δ-endotoxin nomenclature committee as Cry7Ab3 with accession number BI 1015188 in GenBank database.

关 键 词：苏云金芽胞杆菌 Cry7Ab 马铃薯瓢虫 

分 类 号：S188[农业科学—农业基础科学]