应用错配PCR和限制性片段长度多态性分析技术检测HBV YMDD变异株

目的：建立简便易行的检测乙型肝炎病毒（HBV）多聚酶基因（P基因）上YMDD变异株的方法，评价运用拉米夫定治疗慢性乙型肝炎患对此变异的影响，方法：采用套式／错配聚合酶链反应（PCR）结合限制性片段长度多态性（RFLP）分析技术对108例治疗前HBV DNA（PCR）阳性正在拉米夫定治疗中的慢性乙型肝炎患者的HBV YMDD变异情况进行检测。结果：运用上述技术能有效区分HBV YMDD野生株和变异株，上述108例患者经拉米夫定治疗后，HBV DNA阴转者63例（58．3％），HBV YMDD野生株和变异株分别为23例（21．3％）和22例（20．4％），结论：建立的套式／错配PCR－RFLP分析技术可用于HBV YMDD变异株的临床监测，拉米夫定治疗慢性乙型肝炎患者可导致部分患者的HBV多聚酶基因上YMDD发生变异。

Detection of YMDD mutant HBV by mismatched-PCR and analysis of restriction fragment length polymorphism

Objective To establish a convenient method for detecting YMDD mutations in polymerase gene (P gene) of hepatitis B virus (HBV) and to evaluate the effect of lamivudine on these mutations. Methods Sera of 108 patients with chronic hepatitis B treated with lamivudine and showed positive HBV DNA (PCR) before treatment were detected for YMDD mutations in HBV by method of combining nested/mismatched PCR with analysis of restriction fragment length polymorphism (RFLP). Results The above method can be applied to distinguish well YMDD wild and mutant HBV; In patients treated with lamivudine, HBV DNA turned negative in 63 cases(58 3%). 23(21 3%) and 22(20 4%) showed YMDD wild and mutant HBV, respectively. Conclusion Nested/mismatched PCR-RFLP can be applied to identifying YMDD mutant HBV clinically; lamivudine may lead to YMDD mutations in P gene of HBV in some chronic hepatitis B patients treated with lamivudine.