菊酯类杀虫剂抗性棉铃虫para型钠离子通道基因的部分序列测定与突变

NOVEL MUTATION IN THE para-HOMOLOGOUS SODIUM CHANNEL GENE ASSOCIATED WITH NERVE INSENSITIVITY RESISTANCE TO PYRETHROID IN HELICOVERPA ARMIGERA

昆虫神经系统 para型钠离子通道是菊酯类杀虫剂的主要靶标 ,对 10多种昆虫的研究表明 ,钠离子通道基因发生点突变与昆虫对菊酯类杀虫剂的抗性密切相关 .通过RT PCR扩增的方法 ,我们获得了编码溴氰菊酯抗性和敏感品系棉铃虫钠离子通道域II IV的cDNA片段 .核苷酸序列及推导的氨基酸序列分析结果表明 ,溴氰菊酯抗性品系棉铃虫钠离子通道基因不存在其它昆虫中报道的kdr和super kdr抗性突变 ,但是我们发现了一个甲硫氨酸 (M )到亮氨酸 (L)的新突变 ,该突变位于域II和III之间 .由于昆虫钠离子通道高度保守 ,因此 ,该突变很可能与棉铃虫对溴氰菊酯的抗性有关 .另外 ,通过PCR扩增的方法 ,克隆了棉铃虫钠离子通道域II IV的基因片段 ,该片段全长 7334bp,包含有 13个外显子和 12个内含子 ,与果蝇和烟蚜夜蛾相比 ,内含子在基因中的位置基本一致 ,进一步表明昆虫钠离子通道基因在进化过程中高度保守 .图 5表 1参 2

Para voltage dependent sodium channel in the nervous system of insects is the primary target site for pyrethroids. Point mutations in para sodium channel gene are tightly pertinent to insects' resistance to pyrethroids in more than a half dozen insect species. In this study, the cDNA fragments encoding domains II IV of sodium channel gene were cloned from resistant and susceptible strains of Helicoverpa armigera by RT PCR. Sequence comparison between susceptible and nerve insensitive strains showed there were no point mutations of kdr or super kdr associated with knockdown resistance to pyrethroid insecticides. However, a Met to Leu novel point mutation was found and located in the intracellular linker connecting domains II and III. The mutation from Met to Leu is most likely responsible for Helicoverpa armigera resistance to deltamethrin, since sodium channel was highly conservative in insect. In addition, genomic DNA fragment of domains II IV of sodium channel gene was also cloned by PCR amplification and the fragment was 7 334 bp in length, possessing 13 exons and 12 introns. Compared with that of Drosophila melanogaster and Heliothis virescens , these inserted sites of introns were nearly consistent, which further proved the conservatism of para sodium channel during evolution. Fig 5, Tab 1, Ref 22