Bt杀虫基因在桑粒肩天牛幼虫肠道内生优势菌中的转化研究

桑粒肩天牛(Apriona germariHope,Ag)幼虫是一种营钻蛀性生活的重要林业害虫,通过传统纯培养、生理生化鉴定和16SrDNA分子生物学分析等方法分离、鉴定出其肠道优势内生菌溶血葡萄球菌(Staphylococcus haem olyticus,S.haem olytic-us)Ag06菌株和人葡萄球菌(Staphylococcus hom is)Ag08菌株。从中筛选菌株S.haem olyticusAg06进行质粒消除后作为出发菌株,利用电转化技术将含有对鞘翅目昆虫具专一性毒力B t杀虫基因cry3A的Escherichia coli-Bacillus thuringiensis穿梭表达质粒pHT305 a和pHT7911分别转入其中。经质粒稳定性试验、转化子生长特性测试等分析,结果显示cry3A基因已经成功转入Ag幼虫的优势内生菌溶血葡萄球菌中。

Transformation of Bt Pesticidal Gene into Predominant Endogenetic Bacterium Isolated from Apriona germari (Hope) Larvae Intestines

2 strains of culturable predominant endogenetic bacteria(Staphylococcus haemolyticus Ag06 and Staphylococcus homis Ag08)were isolated from serious pest Apriona germari(Hope) larvae intestines and identified by traditional methods of culture,identification and molecular method based on 16SrDNA analysis.Furthermore,the plasmid of the strain S.haemolyticus Ag06 was cured.And then the plasmid-free strain was selected as starting strain to transform Escherichia coli-Bacillus thuringiensis shuttle plasmid pHT305a and pHT7911 which contained specific insecticidal gene Bt cry3A respectively by electrotransformation.The test of the plasmid stability in recombinants,transformants vegetal properties indicated that Bt cry3A gene had been successfully transformed into dominant endogenetic bacteria of A.germari larvae intestines.