程序化设计简并引物与克隆小菜蛾酯酶基因

Automated programming of degenerate primers and the cloning of the diamondback esterase gene.

利用遗传密码简并性 ,针对特定的氨基酸序列设计简并引物 ,是克隆蛋白质家族cDNA的常规方法。文章介绍了利用blastp ,blockmaker,CodeHop ,SwissProt,SpTrEMBL等网络工具及数据库设计昆虫抗性酯酶的简并引物。用这对引物从抗有机磷杀虫剂的小菜蛾Plutellaxylostella中克隆了 1段cDNA。经blastx检索genebank,发现此cDNA产物与其它昆虫抗性酯酶基因有高度的相似性。研究表明 ,程序化设计的简并引物可信性强 ,阳性率高 。

It is a normal strategy to clone protein family cDNA using the degeneracy of the code to design PCR primers from protein multiple\|sequence alignments. With the tools of blastp, blockmaker, CodeHop, SwissProt,and SpTrEMBL, degenerate primers were designed for cloning of the insect esterase cDNA. Using one pair of primers, a partial cDNA was obtained from the diamondback moth by RT\|PCR. Similarity alignment showed that the products of the cloned cDNA were very similar to those of insect esterases. The results indicated that these degenerate primers designed by this software could be used to obtaining specific RT\|PCR product.