焦磷酸测序法和Sanger测序法检测丙型肝炎病毒基因分型的方法学比较

Comparsion of Pyrosequencing and Sanger sequencing in detection of hepatitis C virus genotypes

目的探讨焦磷酸测序法检测丙肝分型在临床应用中的准确性和敏感性,评价其方法的优缺点。方法收集100例丙肝患者的血浆标本,分装相同的2份,通过焦磷酸测序法和Sanger测序法分别进行丙型肝炎病毒基因分型的检测,对两种测序方法的结果进行比较。结果焦磷酸测序法和Sanger测序法检测HCVR N A基因分型时,100例标本中,92例结果一致,8例结果不一致,一致率为92%。其中,HCV-R N A病毒载量>1×104IU/ml有80例,其测序结果完全一致为100%,HCV-R N A病毒载量<1×104 IU/ml有20例,其中焦磷酸测序共检测20例,Sanger测序法共检测14例,12例结果相同。结论焦磷酸测序法检测丙型肝炎病毒基因分型,不仅结果准确可靠,而且与Sanger测序法比较,其敏感性更高。此外,焦磷酸测序还可以进行定量分析。

Objective To compare the accuracy and sensitivity of Pyrosequencing and Sanger sequencing in detection of hepatitis C virus genotypes. Methods Pyrosequencing and Sanger sequencing were used to detect the hepatitis C virus genotypes using plasma of the same 100 patients, results of the two methods were analyzed. Results The results were consistent in 92 of 100 cases when detecting the HCV-RNA genotyping,of which the concordance rate was 92%. There were 80 samples whose HCV-RNA concentration was above1 ×104IU/ml, with completely consistent sequencing results of the two methods. And there were 20 samples whose HCV-RNA concentration was below 1 ×104IU/ml, all of which were detected successfully by Pyrosequencing, while 14 of which were detected successfully by Sanger sequencing, 12 of which had identical results. Conclusions Compared with Sanger sequencing, Pyrosequencing can make results accurate and reliable, and the sensitivity of which is higher. In addition, Pyrosequencing can be used for quantitative analysis.