摘要
将甲型肝炎病毒龙甲株（ＬＪ）结构蛋白基因（包括部分５’端非编码基因，ｎｔ６３０－３０４９）ｃＤＮＡ序列与国外ＨＭ１７５、ＭＢＢ和ＬＡ三个毒株相应区域相比较，结果显示，ＬＪ株结构蛋白基因与ＭＢＢ株的同源性最高，为９６．７％，与ＨＭ１７５和ＬＡ的同源率分别为９５．４％和９１．４％。推导的氨基酸变异率分别为０．９１％，０．９１％和２．９８％。ＬＪ与ＬＡ株氨基酸差异在整个结构蛋白区均有分布，而与ＨＭ１７５和ＭＢＢ的差异仅出现在ＶＰ２和ＶＰ３区。５６种常用限制性内切酶位点分别增加１０、１３和３０个，分别减少１５、１０和２７个，其中ＢｓｔＥⅡ和ＰｖｕⅠ各有一个为ＬＪ株所特有的位点，ＨｉｎｄⅢ、ＰｓｔⅠ和ＳａｃⅠ位点四毒株均一致。将ＬＪ株结构蛋白基因（ｎｔ７４５－２９９３）置换ＨＭ１７５株相应基因区域，插入ｐＪＳＡ１１７５表达质粒７．５ｋ启动子下游，经转染和克隆纯化，获得ｖＪＳＡ１１７５ＨＡＶ５ＬＪ重组痘苗病毒。用甲型肝炎病毒单克隆抗体包被，ＥＬＩＳＡ夹心法检测表达产物，效价为１∶１６。Ｗｅｓｔｅｒｎｂｌｏｔ分析，表达产物与抗－ＶＰ０、抗－ＶＰ１和抗－ＶＰ１、２、３血清特异性结合的带型与已报道ＨＭ１７５重组病毒株的表达产?
The nucleotide and amino acid sequences of a Chinese hepatitis A virus Long Jia (LJ) strain were compared with that of HM175, MBB and LA strains in structural genes (nt 630-3049). The most extensive nucleotide homology was identified between LJ and MBB strains. The identity rates of nucleotide were 95.4%, 96.7% and 91.4%, respectively. Variation rates of amino acid were 0.91%, 0.91% and 2.98%, respectively. A total of 23 amino acid differences located in whole capsid region between LJ and LA strain, especially in VP1. Only 7 amino acid differences located in VP2 and VP3 between LJ and HM175/MBB strain. Restriction enzyme sites increased 10, 13 and 30 sites in 56 restriction enzymes tested, and decreases 15, 1 and 27 sites, respectively. BstE Ⅱ (nt 2810) and Pvu Ⅰ (nt 2013) were the peculiar sites of LJ strain. Hind Ⅲ, Pst Ⅰ and Sac Ⅰ sites were identical among the four strains. After structural gene (nt 745-2993) of HM175 strain was replaced by LJ strain, the complete hepatitis A virus cDNA open reading frame was inserted into pJSA1175 (vaccinia virus expression vector) downstream of promoter 7.5k. Hepatitis A virus antigen expressed was 1∶16 in titer by sandwich ELISA. Band patterns of anti-VP0, anti-VP1 and anti-VP1, 2, 3, were as same as that of HM175 strain and natural hepatitis A virus antigen by Western blot analysis.
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
1997年第3期208-211,共4页
Chinese Journal of Experimental and Clinical Virology
基金
国家863高科技生物技术领域基金
