烟实夜蛾细胞色素P 450 cDNA片段的克隆与序列分析

Clone and Sequence Analysis of cDNA Fragment of Cytochrome P 450 from Oriental Tobacco Budworm (Helicoverpa assulta Güenée)

昆虫细胞色素P 450与昆虫抗药性关系密切,本研究以烟实夜蛾5龄幼虫中肠的总RNA为模板,设计并合成简并引物,利用反转录-多聚酶链式反应(RT-PCR)扩增出了烟实夜蛾细胞色素P 450基因的cDNA片段,将其克隆到pMD 18-T载体进行序列测定,测序得到的491 bp的片断编码163个氨基酸残基,且该片断在阅读框内.与已公布的谷实夜蛾、棉铃虫和烟芽夜蛾细胞色素P 450氨基酸序列进行比较,其一致性分别为98.7%,98.7%和92.1%.

Cytochrome P 450 is closely related with the insecticide resistance of insect, cDNA fragment of cytochrome P 450 gene in the fifth instar larva of oriental tobacco budworm was amplified by RTPCR with total RNA as template and using a degenerate primer. The target fragment was further sequenced after being cloned into pMD 18-T vector plasmid. The sequenced result showed that 491 bp nucleotide sequence that encodes 163 amino acid residues was in the encoding frame. Compared with the known cytochrome P 450 nucleotide sequence of Helicoverpa zea,Helicoverpa armigera, and Heliothis virescens, the identity were 98.7% ,98.7% and 92.1% , respectively.