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采用降落 PCR技术 ,根据昆虫 para型钠通道 区 S4至 S6及 与 连接区的保守区域设计简并引物 ,成功地克隆了棉蚜 para型钠离子通道 c DNA和基因组 DNA片段 (Gene Bank登录号分别为 :AF4 114 5 5、AF4 475 74 ) ,其中 c DNA片段为 6 5 9bp,编码 2 2 0个氨基酸。利用 c DNA片段设计特异性引物克隆获的基因组DNA片段为 132 8bp,共有 2个内含子。Blast序列分析表明 ,PCR扩增获得的棉蚜 para型钠离子通道 c DNA片段所编码的氨基酸与其他昆虫的 para型钠通道氨基酸具有很高的同源相似性 ,与马铃薯甲虫、德国小蠊、果蝇和家蝇的同源性分别为 6 5 %、6 3%、6 3%、5 9%。棉蚜 para型钠离子通道 c DNA和基因组 DNA片段的克隆对于农药分子设计与害虫抗性机制 ,特别是靶标抗性分子监测具有重要意义。
Based on the parasite sodium channel regions S4 to S6 and the degenerated primers with the conserved region of the junction, the lapin PCR technique was used to successfully clone the c-DNA and genomic DNA fragments of the para-type sodium ion channels of cotton aphid AF4 114 5 5, AF4 475 74), in which the c DNA fragment is 659 bp, encoding 220 amino acids. The genomic DNA fragment cloned using the specific primers designed by cDNA fragment was 132 8 bp with a total of 2 introns. Blast sequence analysis showed that the amino acids encoded by the para-sodium channel c DNA fragment of cotton aphid obtained by PCR amplification had high homology with para-type sodium channel amino acids of other insects and were highly homologous to potato beetle, German cockroach, Drosophila and housefly homology were 65%, 63%, 63%, 59%. Cloning of para-type sodium ion channel c DNA and genomic DNA fragments of cotton aphid is of great significance for pesticide molecular design and pest resistance mechanisms, especially target-resistant molecular monitoring.