中国人丙型肝炎病毒基因组3‘端非编码区的研究

目的 分析中国丙型肝炎病人ＨＣＶ基因组３‘端非编码区，以促进对ＨＣＶ基因组复制机制的研究。方法 采用两种方法，从上海地区感染ＨＣＶ的病人血清中，扩增获得ＨＣＶ基因组３’端非编码区：一是用套式ＰＣＲ直接的增，二是先分别ＨＣＶ３‘ＮＣＲ的前半部分和后半部分，再将两片段进行融合ＰＣＲ。ＰＣＲ产物进行测序后作同源性分析。

Study of the 3'noncoding region of Chinese hepatitis C virus genome]

OBJECTIVE: To analyze the 3' noncoding region (3' NCR) of HCV genome from Chinese hepatitis C patients so as to facilitate further study of mechanism of HCV gene replication. METHODS: Two different strategies were employed to amplify the full-length of the 3' noncoding region of HCV genome from sera of HCV infected patients in Shanghai area: one was to amplify the full-length fragment directly by nested PCR and the other amplify two overlapping fragments. The PCR products were further analyzed by sequencing and nucleotide alignments. A HCV genome 3'NCR based RT-PCR was developed and its specificity and sensitivity for HCV RNA detection in sera was compared with the established 5'NCR based RT-PCR. RESULTS: Sequence analysis showed that Chinese HCV genomic 3' NCR consists of three parts: the 5' region, poly (U-UC) tract and the 98-base region. Sequence alignments revealed that, while the 98-base regions were completely conserved in different isolates and were identical to the reported sequences, the poly (U-UC) region shared highly diversities. A high degree of concordance(95%) between the 3'NCR and 5'NCR RT-PCR for detection of HCV RNA in sera was found. CONCLUSION: The high conservation at the 3' NCR(98 bases) of HCV genome among different isolates indicated that this region may be critical for HCV gene replication The 3'NCR based RT-PCR may be a useful addition to available systems to diagnosis HCV infection.