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目的建立稳定表达乙型肝炎病毒X基因肝细胞株。方法对含酶切位点Eco RⅠ、HindⅢ的X基因序列进行PCR扩增,构建HBVx基因质粒(pc DNA3.1(+)-HBVx),利用转基因技术将X基因转入正常肝细胞构建稳定表达X基因的细胞株。结果 X基因亚克隆入pc DNA3.1(+),有完整的X基因片段,转入正常肝细胞获得稳定表达X基因的细胞株,转染C57BL/6正常肝细胞有HBVx m RNA表达,且C57BL/6/HBVx有HBV蛋白表达。结论成功构建稳定表达乙型肝炎病毒X基因肝细胞株。
Objective To establish a stable expression of Hepatitis B virus X gene hepatocyte strain. Methods The X gene sequence of EcoRⅠand HindⅢ with restriction endonuclease sites was amplified by PCR, and the HBVx gene plasmid (pcDNA3.1 (+) - HBVx) was constructed. Transient gene transfer of X gene into normal hepatocytes was used to construct stable expression X gene cell line. Results The X gene was subcloned into pc DNA3.1 (+), and a complete X gene fragment was obtained. The X gene was transformed into normal hepatocytes to obtain a stable cell line expressing X gene. HBV xm RNA was expressed in C57BL / 6 normal hepatocytes C57BL / 6 / HBVx has HBV protein expression. Conclusion The stable expression of Hepatitis B virus X gene hepatocyte strain was successfully constructed.