PCR扩增循环数与低拷贝模板DNA的STR分型

A study on the STR genotyping of low copy number by increasing PCR amplification cycles

目的探讨PCR扩增循环数对低拷贝模板DNA的STR分型的影响。方法模板DNA(9947A)的不同扩增用量(含低拷贝模板量),采用ProfilerPlus试剂盒,扩增循环数分别为28、30、32、34、38次,3100型基因分析仪(ABI,美国)检测结果。结果循环数从28次增至38次,模板DNA量最低检出量可从0.25ng减少至0.0312ng;先循环28次后,每反应加0.3μlAmpliTaqGoldDNA聚合酶,再循环6次较1次34个循环的检测灵敏度高,相当于1次38个循环的效果。结论增加PCR扩增循环数,可能影响低拷贝模板DNA的STR分型。

Objective To investigate the influence of increasing PCR amplification cycles on low copy number DNA genotyping. Methods Various amount of template DNA (9947A) was analyzed by 3100 Genetic Analyzer with the multiplexing Profiler Plus Kit at 28, 30, 32, 34 and 38 amplification cycles. Results When PCR cycle numbers were increased from 28 to 38, the minimal amount of sample for extreme sensitivity of detection was reduced from 0.25ng to 0.0312ng. Conclusion The sensitivity of the profiling process may be enhanced by increasing the number of amplification cycles. DNA profiling is not available by increasing the cycle numbers when the sample of template DNA is lower than 0.01ng.