构建乙型肝炎病毒基因参照品判断基因分型的探讨

Establishment of a method of hepatitis B virus genome reference standard to estimate genotyping

目的利用乙型肝炎病毒（HBV）基因参照品判断HBV基因分型。方法自行制备HBV—S区基因参照品A、B、C、D型并测序及比较同源性，利用限制性内切酶MboI、EarI酶切参照品和样品扩增产物，从而确定样品基因型。利用该方法对克拉玛依地区272例HBV—DNA阳性血清基因分型。结果测序证实，HBV基因参照品与39例GenBank序列同源性均在96％以上。272例样品成功分型241例（88．6％）。检出B、C、D型，未检测到A型。抽检12例测序同源性大于96％。结论依据基因参照品核酸碱基片段大小判断分型，简明直观，适用于大样本的筛检及临床应用。

Objective To estimate genotyping with hepatitis B virus genome reference standards. Methods Hepatitis B virus S gene reference standard A, B, C and D were established by our laboratory and sequencing and homology comparison were performed. By restriction endonuclease MboI and EarI digestion of reference standards and sample amplified product, genotyping was idenlified. Genotypes for 272 cases of HBV DNA posilive sera were eatablished. Results Gene sequencing con firmed that homology of HBV gene reference standards with 39 samples in GenBank was above 96%. 241 cases （88.6%） were genotyped successfully in 272 samples. Genotype B, C and D were detected, while genotype A was not detected. Sequencing analysis for 12 cases displayed 96% homology. Conclusion Estimating HBV genotypes by means of genome reference standards is confirmed to be highly sensitive and can be used in large scale surveys and clinical research.