乙型肝炎病毒DNA聚合酶反式调节蛋白HBVDNAPTP1基因全长cDNA RACE克隆

RACE cloning of the full-length gene cDNA of hepatitis B virus DNA polymerase trans-activated protein HBVDNAPTP1

目的利用cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)技术克隆乙型肝炎病毒DNA聚合酶反式调节蛋白HBVDNAPTP1基因的全长cDNA序列。方法提取肝母细胞瘤细胞系HepG2总RNA,应用逆转录聚合酶链反应(reverse transcription-PCR,RT-PCR)技术建立RACE cDNA文库,进行RACE实验,扩增HBVDNAPTP1基因全长cDNA序列。结果经RACE实验,获得HBVDNAPTP1基因全长cDNA序列为2 537bp,经RT-PCR验证确定该基因真实存在,且与GenBank数据库中注释的其他基因无同源性。结论成功获取HBVDNAPTP1基因的全长cDNA序列,为进一步开展HBVDNAPTP1生物学功能及调控机制研究提供了线索和依据。

Objective To clone the full-length gene cDNA sequence of hepatitis B virus DNA polymerase trans-activated protein HBVDNAPTP1 by using rapid amplification of cDNA ends（RACE）experiment.Methods The total RNA of hepatoblastoma cell line HepG2 was extracted,and the RACE cDNA library was constructed by using reverse transcription polymerase chain reaction（RT-PCR）.The RACE experiment was performed to amplify the full length of HBVDNAPTP1 gene cDNA sequence.Results The fulllength cDNA sequence of HBVDNAPTP1 gene was obtained with rapid 3′ 5′ RACE experiment.The gene was found to truly exist in HepG2 by RT-PCR and had no homology with other genes in the GenBank database.Conclusion The full-length cDNA sequence of HBVDNAPTP1 gene was successfully obtained,which provided clues and basis for further study on the biological function and regulatory mechanism of HBVDNAPTP1.