肝癌病人血清中庚型肝炎病毒非结构基因(NS)5区cDNA的序列分析

Sequencing of partial gene of hepatitis G virus in the sera of patients-with hepatocellular carcinoma

通过逆转录-套式-聚合酶链反应,从2例肝癌患者血清中扩增出长994bp的庚型肝炎病毒(HGV)cDNA序列,聚合酶链反应(PCR)产物纯化后以双脱氧末端终止法从双向直接测定其序列。结果,所分离的2株HGV NS5区994bp核苷酸与国外已发表的3株HGV序列比较,同源性为87.21%～93.67%;2株间的同源性为93.92%。根据所测得的cDNA序列推导出其编码的313氨基酸序列,在氨基酸水平上本文报告的2株与国外发表的3株序列相比,同源性在94.25%～97.44%之间,2株之间为96.49%。在此区内发现有16个保守的脯氨酸残基及8个保守的半胱氨酸残基。结论:首次从肝癌患者血清中分离出2株HGV,其NS5区部分序列在核苷酸水平及氨基酸水平均较保守,与非肝癌患者血清中分离的HGV相比,变异不大。

To sequence genome of hepatitis G virus (HGV) in the sera of patients with hepatocellular carcinoma (HCC), total nucleic acid were extracted from the sera of 6 HCC cases, then subjected to reverse transcription-nested-polymerase chain reaction (RT-nested-PCR) using the primers derived from the putative nonstructural 5 (NS5) region. HGV RNA was detected in 2 of the 6 HCC patients. The positive PCR products, 994bp in length, were directly sequenced using the Sanger's method after purification. Prtial HGV NS5 gene of 2 isolates (HG-CA1, HG-CA2) detected in this study were 87.21% to 93.67% identical compared with 3 reported HGV isolates (GBV-C, R10291, PNF2161) over the 994 nucleotides examined. Homology between 2 isolates was 93.92%. 94.25% to 97.44% homology with 3 published iso-lates were found in the amino acid sequences deduced from HG-CA1 and HG-CA2., and 96.49% identical between HG-CA1 and HG-CA2. In the region sequenced, there existed 16 conserved proline residues and 8 conserved cysteine residues. The partial nucleotide sequences, as well as amino acid sequences, of the putative NS5 region of HGV detected in this study were relatively conserved. There was no significant variability in NS5 region sequenced between the isolates from HCC cases and the isolates from non-HCC patients.