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目的为从全基因水平研究乙型肝炎病毒变异与致病性的关系，建立血清标本经聚合酶链反应（ＰＣＲ）加酶切方法扩增及克隆ＨＢＶ全基因组的新方法。方法设计了含ＳｐｅⅠ，ＳａｌⅠ及ＳａｐⅠ酶切点的引物，分别扩增１．１５ｋｂ和２．０５ｋｂ的单链及双链ＤＮＡ区。经酶切及连接后获得ＨＢＶ全基因克隆。结果用新建立的方法直接从少量血清中克隆了ＨＢＶ全基因，转染ＨｅｐＧ２细胞可表达抗原并有胞内复制。结论该法可用于对临床上ＨＢＶ毒株的基因结构和功能的研究
Objective To study the relationship between hepatitis B virus (HBV) mutation and pathogenicity at the whole gene level and to establish a new method for the amplification and cloning of HBV whole genome by polymerase chain reaction (PCR) and enzyme digestion. Methods Primers containing Spe Ⅰ, Sal Ⅰ and Sap Ⅰ cleavage sites were designed to amplify single- and double-stranded DNA regions of 1.15 kb and 2.05 kb, respectively. After digestion and ligation obtained HBV whole gene cloning. Results The whole HBV gene was cloned directly from a small amount of serum using the newly established method. HepG2 cells transfected with HepG2 cells could express the antigen and replicate intracellularly. Conclusion This method can be used to study the gene structure and function of clinical HBV strains