多位点ARMS-PCR法进行HBVS区检测分型

DETECTING HBV S REGION WITH MULTILOCUS ARMS-PCR

目的建立一种准确有效的PCR法HBV分型的实验方案。方法对80例包含了HBV不同型别(B/C/D型)的血清样本进行了HBV S区测序,通过分析国人HBV B/C/D型S区碱基固有差异位点,设计得针对各种HBV型别的高度特异性ARMS引物与探针,并用于另外120例HBV样本的PCR检测实验,同时进行测序进行检测准确度验证。结果多位点ARMS-PCR法与测序法的型别检出吻合率为99.5%,其中一例HBV B/C型混合感染由ARMS-PCR法检出并经追溯确认。结论经验证可见设计于S区型别固有差异位点之上的ARMS引物与探针于PCR检测中能以极高的特异性对HBV血清样本进分型,并能检出HBV混合型中的劣势病毒株。PCR法以其低成本、结果容易判定以及高灵敏度等优点,于临床HBV分型上是更为推荐的分子诊断学方法。

Objective To establishan accurate and effective PCR system of HBV genotyping detection . Methods S-region genes of 80 HBV serum samples which contains HBV genotype B/C/D were sequenced . High specific ARMS-primers and probes for each genotype were designed with the differentiation analysis of the S gene among HBV genotype B/C/D ,the genotyping PCR of the other 120 HBV samples was per-formed ,and the DNA sequencing of each sample was performed simultaneously .Results The coincidence rate of Mutilocus ARMS-PCR and DNA sequencing was 99 .5% .One sample with the mixed infection of HBV genotype B/C was detected by ARMS-PCR and confirmed by tracing back .Conclusion Compared to the results of DNA sequencing ,the high specific ARMS-primers and probes for each genotype worked ex-cellently in the PCR of HBV genotyping detection ,and the inferior viral strain in the mixed infection sam-ples was detected in PCR .PCR is a recommended method in HBV genotyping for its low cost ,easy judg-ment and high sensitivity .