桔小实蝇电压门控钠离子通道基因cDNA克隆及其生物信息学分析

 【目的】克隆获得桔小实蝇（Bactrocera dorsalis）电压门控钠离子通道基因cDNA序列，明确其典型特征，为研究桔小实蝇抗性分子机理奠定基础。【方法】采用RT-PCR和PCR技术，克隆桔小实蝇钠离子通道基因cDNA序列，利用相关软件对其序列进行生物信息学分析。【结果】克隆得到1条长为6 446 bp的cDNA序列，包含1个6 405 bp的完整开放阅读框，共编码2 134个氨基酸。同源比对发现，桔小实蝇与果蝇（Drosophila melanogaster，NP_001188635）和家蝇（Musca domestica，AAB47604）钠离子通道基因相似度分别高达91.7%和86.9%，而与人的钠离子通道Nav1.2基因（Homo sapiens，NP_066287）相似度为42.3%。所克隆序列包含昆虫钠离子通道所有典型特征。【结论】成功地从桔小实蝇中克隆得到钠离子通道基因完整开放阅读框。该钠离子通道基因存在丰富的选择性剪接，发现了3个可能与抗性相关的碱基取代。钠离子通道基因有可能发展成为一种昆虫系统发育研究的分子标记。

Cloning and Bioinformatics Analysis of Voltage-Gated Sodium Channel Gene cDNA in Bactrocera dorsalis (Hendel)

【Objective】 The objective of this study is to clone voltage-gated sodium channel gene (VGSC) from Bactrocera dorsalis (Hendel), and to identify its typical hallmarks. It will provide basic molecular information for clarifying insecticide resistance mechanism of B. dorsalis.【Method】 The cDNA sequence was isolated using RT-PCR and PCR methods. Based on the sequencing results, the bioinformatics analysis of nucleic acid and putative amino acid was conducted.【Result】An almost full-length cDNA sequence (6 446 bp) of VGSC was obtained, including a complete open reading frame (ORF) of 6 405 bp, which encoded 2 134 amino acids and included all the typical hallmarks of VGSC. The amino acid shared 91.7%, 86.9% and 42.3% identity with sodium channel genes of Drosophila melanogaster (NP_001188635), Musca domestica (AAB47604), and Homo sapiens Nav1.2 (NP_066287), respectively. 【Conclusion】A complete ORF of VGSC was sequenced with clone strategy from oriental fruit fly for the first time. Abundant alternative splicing phenomena existed in the VGSC of B. dorsalis. Three potential mutation points, which may confer resistance to pyrethroids and DDT, were observed. It is suggested that VGSC would be used as a molecular marker in phylogenetic study of insect.