利用降落PCR扩增KS基因

通过优化SDS法提取链霉菌(Streptomyces spiramyceticus)和2株由生物技术与资源利用国家民委——教育部重点实验室分离获得的放线菌D8和D18基因组DNA,同时应用降落PCR和普通PCR扩增酮基合酶基因(ketosynthase,KS)。结果表明,扩增出的特异性条带与目的基因片段长度一致,降落PCR法较普通PCR法扩增KS基因特异性更高。使用普通Taq酶,降落PCR程序能明显提高PCR的特异性,它可用于扩增普通PCR难以扩增的基因片段,或在假阳性难以去除的情况下提高PCR的特异性。

Touch Down PCR for Amplification of Ketosvnthase Genes

Objective: To get target KS genes by a simple and efficient touch down PCR(TD-PCR).Methods: Genomic DNA from three strains of actinomyces was extracted by modified SDS method.The ketosynthase(KS) gene fragments from three strains of actinomyces were amplified with touch down PCR and common PCR.Results:Target genes were obtained by TD-PCR.The results indicated that touch down PCR was efficient in amplifying the gene from three strains of actinomyces and it was more specific than common PCR.Conclusion: TD-PCR with Tag DNA polymerase has higher specificity than common PCR.It can efficiently improve specificity of PCR and be used to clone gene fragments which are difficult to clone by the common PCR method,or to increase the specificity of PCR when false positivity is difficult to eliminate.