摘要
丙型肝炎病毒(Hepatitis C virus,HCV)是散发性及输血后肝炎的主要病原体,易致慢性肝炎、肝硬变或肝细胞癌。为了研究中国HCV基因组的变异及结构特征,我们从中国丙型肝炎(丙肝,Hepatitis C,HC)患者血浆中提取HCV基因RNA,用随机引物合成HCV cDNA并构建其λgtll cDNA库。对此基因库进行了免疫筛选,并对2个阳性克隆(Q349和Q653)在pUC18质粒中进行了亚克隆。序列分析结果表明:Q349和Q653分别来自HCV基因组核心(C)区(第554～902位)和非结构3(NS3)区(第4175～4827位)。Q349和Q653与HCV原型相应序列在核苷酸和氨基酸水平上的同源性分别为86.8％,80.2％和97.3％,93.1％;与日本HCV相应序列间有更高的同源性,故属于HCVⅡ组。特异性灾验表明,Q349和Q653的编码多肽可与HC人群血清特异反应而不与正常人血清反应,且Q653与中国HC血清反应的阳性率(95.8％)比日本HC血清的阳性率(85.7％)高。表明根据中国HCV基因序列(特别是非结构区序列)设计引物或合成多肽将更适合中国人群HCV感染的检出。
Hepatitis C virus (HCV) is a major causative agent for sporadic and post-transfusion hepatitis, which frequently progresses into chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. In order to clarify the genomic variations and the structural characteristics of Chinese HCV isolate, we extracted HCV genomic RNA from plasma specimens of Chinese hepatitis C (HC) patients, reverse-transcribed to HCV cDNA with random primers and constructed successfully an HCV cDNA R gt11 library of Chinese type. Two positive clones (Q349 and Q653) were selected by immunoscreening from the library and subcloned into plasmid vector pUC18. Sequence analyses indicated that Q349 was derived from core region (positions 554-902) of HCV genome while Q653 was from NS3 region (positions 4175-4827) corresponding with the prototype HCV nucleotide sequence. The homologies of Q349 and Q653 with the equivalent sequences of HCV prototype were 86.8% and 80.2% at the nucleotide level, and 97.3% and 93.1% at the amino acid level, respectively. It was found that Chinese HCV clones had higher homologies with Japanese HCV isolates, and should belong to HCV group II. Specificity test proved that the encoded peptides of the 2 Chinese HCV cDNA clones reacted specifically with sera from HC patients and had no reaction with sera from healthy individuals. More importantly, clone Q653 showed higher positive reaction rates with Chinese HC sera (95.8%) than those with Japanese ones (85.7%), which strongly suggests that the sequences from Chinese HCV genome (especially from NS regions) would be more suitable for primer designing or peptide synthesis for the use in the detection of HCV infection among Chinese people.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
1992年第5期401-410,共10页
Academic Journal of Second Military Medical University
关键词
肝炎病毒
基因库
免疫技术
hepatitis virus
gene pool
immunization
