从杭州、兰州两地各一例乙型肝炎病毒 (HBV)表面抗原阳性血清中提取病毒DNA ,采取PCR技术扩增出前表面抗原 (preS)基因片段 ,重组到质粒载体上 ,对该基因进行了全序列测定 [GenBank索取号CpreS HZ :AF 32 5 6 74;preS LZ :AF 32 5 6 75 ]。克隆的HBVpreS基因杭州分离物 (preS HZ)和兰州分离物 (preS LZ)全长 5 2 2个核苷酸 ,编码174个氨基酸。preS HZ与已发表的HBVadr亚型上海分离物[8] 、北京分离物[9] 、日本分离物[5] 、HBVadw亚型[10 ] 和ayw亚型[11] preS基因的核苷酸序列同源性分别为 96 .7%、96 .2 %、97.3%、88.7%和 84.1% ,氨基酸序列同源性分别为 96 .0 %、94.9%、97.1%、85 .1%和 85 .4% ;preS LZ与相应序列的核苷酸序列同源性分别为 96 .4%、96 .2 %、96 .9%、88.7%、83 .7% ,氨基酸序列同源性分别为 94.9%、94.9%、96 .0 %、85 .1%、84.1%。分子进化分析 (DNASTAR ,1999)表明 ,克隆的两例HBVpreS基因属于adr亚型。preS LZ与preS HZ之间有两个核苷酸变异 (对应两个氨基酸变异 ) ,相对于以上报道的序列二者含有四个特异的氨基酸突变位点。在免疫保护区内二者具有较好的保守性 ,可用于表达乙肝重组亚单位疫苗。

PreS gene fragments were inserted into pBluescript SK after being amplified by PCR from HBV DNA,which was extracted from HBsAg-positive sera(Hangzhou and Lanzhou samples)and the cloned genes were sequenced.The complate nucleotide sequences of the cloned preS genes(preS-HZ and preS-LZ,GenBank accession numbers:Af325674 and AF325675 respectively) were 522bp long,encoding 174aa.The identity of preS-HZ with published preS gene Shagnhai isolate(adr subtype:Gan,et al.,1986),Beijing isolate (adr subtype:Qi,et al.,1989),Japan isolate(adr subtype:Ono,et al.,1983),adw subtype(Valenzuela,et al.,1980),ayw subtype (Galibert,et al.,1979)was respectively 96.7%,96.2%,97.3%,88.7%,84.1% for nucleotide sequence and 96.0%,94.9%,97.1%,85.1%,85.4% for amino acid sequence;correspondingly,preS-LZ was respectively 96.4%,96.2%,96.9%,88.7%,83.7 for nucleotide sequence and 94.9%,94.9%,96.0%,85.1%,84.1% for amino acid sequence.Only two nucleotide substitutions(resulting in two amino acid changes) were observed between preS-HZ and preS-LZ