B.t.c. cry1C全长基因的克隆及其在增产菌中的表达

CLONING OF FULL CRY1C GENE FROM B.T.C.AND EXPRESSION IN CROPS BENEFICIAL BACTERIA BACILLUS CEREUS

设计的一对引物 ,对苏云金芽孢杆菌科默尔亚种 (Bacillusthuringiensissubsp .colmeri)1 5A3菌株中的cry1C基因进行PCR扩增 ,得到包括结构基因 ,调节基因在内的全长为4 0kb的PCR产物。经两步克隆 ,将此基因连接至穿梭表达载体pHT31 5上 ,得到重组质粒pHT 1C。通过电转化将其导入一株增产菌蜡状芽孢杆菌 (Bacilluscereus) 95 0 9菌株 ,SDS PAGE检测到 1条 60kD左右的蛋白带 ,镜检观察到菱形晶体 ,生测结果表明 ,cry1C基因的导入使Bc95 0 9菌株获得了对甜菜夜蛾的杀虫活性。

Specific primers 1CaA/1CaB for full cry1C gene in Bacillus thuringiensis subsp.colmeri strain 15A3 were designed. The 4.0kb PCR product included the whole ORF and regulation region of cry1C gene. This PCR product was linked with shuttle vector pHT315 by two cloning steps. The recombined plasmid pHT-1C was electroporated into Bacillus cereus 9509, a kind of bacteria that beneficial to crops. The transformant could produce bipyramidal-shaped parasporal inclusions. The 60kD protein band was detected by SDS-PAGE. The bioassay result showed that the cry1C gene transformant of Bc 9509 had insecticidal activity to Spodoptera exigua.