乙型肝炎病毒C基因启动子变异检测方法的建立

Establishment of a Method for Detecting Mutation in Basic Core Promoter (BCP) in Hepatitis B Virus

目的建立乙型肝炎病毒(HBV)C基因启动子(BCP)变异的限制性片断长度多态性(RFLP)检测方法,更好地了解BCP变异的发生与乙型病毒性肝炎(乙肝)的病情、预后以及治疗的影响。方法结合引物设计软件Primer Premier5.0,选定保守区域,设计了2对最佳引物,建立了HBV BCP变异检测PCR-RFLP的实验方法。结果对34例不同乙肝患者进行BCP变异检测发现:肝硬化(LC)组和慢性乙型肝炎(CHB)组中发生变异的例数明显高于无症状感染者(Asl)(χ2值分别为6.39和3.98,P<0.05)。并且HBeAg(-)组发生变异的例数明显高于HBeAg(+)组(χ2=4.60,P<0.05),结论该方法操作简便、敏感度与特异性高、结果可靠,并可用于较大范围调查该变异株的流行情况,为临床进一步诊断和治疗提供了科学的依据,同时对人们进一步了解HBVBCP变异的发病机制具有重要意义。

[ Objective] To establish the detection method of mutation in basic core promoter （ BCP} in hepatitis B virus with restriction fragment length polymorphism analysis （RFLP} in order to further understand the effect of the BCP mutation on the situation, recover and therapy of hepatitis B. [ Methods] Two pairs of the most specific primers combined with Primer 5.0 in conservative region was applied to establish PCR-RFLP method for detecting BCP in HBV. [ Results] The rates of BCP mutation among liver cirrhosis （LC） and chronic hepatitis B （CHB} patients were significantly higher than that among asymptomatic HBV infectors （AsI） （ P 〈 0.05 } ; they were also higher among the patients with HBeAg negative than those with HBeAg positive （ P 〈 0.05 ）. [ Conclusion] The PCR-RFLP method is a simple, sensitive, efficient and accurate and is quite applicable for investigating the epidemic situation of BCP mutation and provide scientific basis for the diagnosis and therapy. It also has important significance to further understand the mechanism of BCP mutation in HBV.