摘要
目的比较普通聚合酶链反应(PCR)与降落聚合酶链反应(Touchdown PCR,TD-PCR)在临床医学科研中的价值。方法以人类外周血基因组DNA为模板,设计VHL基因3个外显子的3对引物,根据普通PCR及TD-PCR原理设计包括3个外显子片段在内的PCR程序,通过试验选择PCR最佳反应条件,在同一程序中分别对3个片段进行扩增,琼脂糖凝胶电泳检测扩增产物,纯化后PCR产物测序分析两种试验方法的差别。结果电泳检测及纯化后DNA产物测序分析均显示TD-PCR扩增产物条带特异性、效率较普通PCR扩增产物高。结论成功建立了TD-PCR方法,TD-PCR方法较普通PCR更为高效实用,为临床基因突变筛查研究提供了快速可靠的手段。
Objective To compare the ordinary polymerase chain reaction （PCR） and touchdown （TD）-PCR in clinical medical researeh. Methods We used genomic DNA from human peripheral blood as templates and designed three pairs of primers of VHL gene three exons. On the basis of the principle of PCR and TD-PCR, we designed the programs, including the three exons. We chose the better reaction conditions through the PCR tests. The same programs were carried out on three fragments. The PCR products were detected by gel agarose electrophoresis. We analyzed the difference between the two methods by sequencing purified PCR products. Results It was indicated that TD-PCR was more specific and effective than ordinary PCR, according to electrophoresis detection and sequencing analysis. Conclusion TD-PCR, which acts as a rapid and reliable method, is more efficient than ordinary PCR for clinical gene mutation screening.
出处
《临床军医杂志》
CAS
2008年第2期280-282,共3页
Clinical Journal of Medical Officers
关键词
基因
聚合酶链反应
降落聚合酶链反应
gene
polymerase chain reaction
touchdown polymerase chain reaction
