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

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

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者对此变异的影响。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 10 8例治疗前HBVDNA(PCR)阳性正在拉米夫定治疗中的慢性乙型肝炎患者的HBVYMDD变异情况进行检测。结果 运用上述技术能有效区分HBVYMDD野生株和变异株 ;上述 10 8例患者经拉米夫定治疗后 ,HBVDNA阴转者 63例 (5 8 3 % ) ,HBVYMDD野生株和变异株分别为 2 3例 (2 1 3 % )和 2 2例 (2 0 4% )。结论 建立的套式 /错配PCR -RFLP分析技术可用于HBVYMDD变异株的临床监测 ;

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.