HepG2.2.15细胞株中乙型肝炎病毒基因异质性的实验研究

The heterogeneity of HBV gene in HepG2.2.15 cells

目的以乙型肝炎病毒(HBV)前CC基因的异质性来探讨在HepG2.2.15细胞体内是否存在HBV准种。方法依据Genebank中HBVayw亚型全基因序列,选择保守区域设计出特异性多聚酶链反应引物,自HepG2.2.15细胞培养上清液中扩增HBV前CC基因序列,克隆入pGEM-TEasy载体,挑选15个克隆测序以比较病毒的变异程度。结果测序发现HepG2.2.15细胞HBV前CC基因存在异质性,碱基序列之间的同源性大于93%,突变类型全部为碱基替换。结论HepG2.2.15细胞内存在HBV准种。

Objective To investigate the HBV quasispecies in the long-term cultured HepG2.2.15 cells. Methods A set of specific primers was synthesized according to the HBV DNA sequence of the subtype ayw in the genebank, and the whole pre-core/core region was amplified by the PCR method from the supernatant of HepG2.2.15 cells. Then the PCR products were subcloned into pGEM-T Easy vectors. The selected clones were sequenced, and then the sequence comparison was performed. Results One sequence of selected clones was found to be different the homology of the nucleic acid sequence from the same patient was above 93.0%. The mutation type in this region was base pair substitution. Conclusion The results indicated that there were HBV quasispecies in the HepG2.2.15 cells. The HBV heterogeneity in the HepG2.2.15 cells may influence the application of hepatitis B gene therapy.