家蚕核多角体病毒的多角体蛋白基因

本文用AcMNPV多角体蛋白质基因的克隆DNA作探针,从BmS NPV DNASal Ⅰ酶酶解片段基因库中筛选出一个有同源顺序的克隆pBN 61。该克隆含大小为1.65 kb插入DNA片段。本文报道了该片段的Hind Ⅲ,Hpa Ⅱ和Alu Ⅰ等限制性内切核酸酶酶切图谱分析和部分DNA顺序分析结果。测得的DNA顺序中有138个碱基与已报道的BmSNPV多角体蛋白质一级结构中的46个氨基酸顺序相比较,除一个氨基酸发生变化外,其余完全相一致。克隆pBN 61可能包含完整的Bm SNPV多角体蛋白质结构基因。     

带肝细胞受体结合区的HBV表面抗原蛋白性质的研究——preS区有不同缺失的表面抗原蛋白性质的比较

本文用聚合酶链反应(PCR)构建了四对preS区有缺失突变的乙型肝炎病毒表面抗原基因。对这些突变体及我们以前构建的缺失突变体在哺乳动物细胞中的表达和产物性质进行了比较,结果表明,preS区的部分缺失并不影响羧端S区的基本结构和氨端preS区的表面分布性。当缺失preS1区滞留顺序(a.a.2—19)后,preS区对S区带主要抗原决定簇的区域的掩蔽明显减弱。我们发现过长的preS区严重影响表面抗原蛋白从哺乳动物细胞中的分泌,除已知的滞留顺序外,preS1的另一区域(a.a.48—65)可能也有阻滞表面抗原蛋白分泌的作用,而preS2区对LS蛋白分泌的阻滞不起主要作用。preS1的肝细胞受体结合区(a.a.21—47)和S区直接融合所得蛋白性质稳定,分泌良好,有很强的抗preS1抗原性,是值得发展为新型疫苗的带preS1肝细胞受体结合区的表面抗原蛋白。     

adr亚型乙型肝炎病毒表面抗原基因的核苷酸顺序

本文应用化学降解法测定了adr亚型乙型肝炎病毒pADR-1 DNA中包含表面抗原基因(s基因)的XhaI BamHI片段顺序,共1279碱基对。比较了adr亚型与已知的adw,adyw,ayw的s基因的核苷酸顺序及其编码的HBsAg氨基酸顺序的差异,发现了一些新的变异位点,差异集中分布在HBsAg的亲水区域。但在一些可能具有生物功能的位点,各亚型间并无变异,在遗传和进化中是相对保守的。比较s基因区和非s基因区的核苷酸变异情况表明,非s基因区的变异频率高一倍,s基因区是相对保守的。     

THE EXPRESSION OF HBsAg GENE IN YEAST UNDER GAL-10 PROMOTER CONTROL

The surface antigen gene of HBV (adr subtype) is inserted into the YEP 51 which is replicated and selected in E. coli and baker yeast Saccharomyces cerevisiae. The HBsAg gene can be expressed in yeast under GAL-10 promoter control. The protein synthesized in yeast is assembled into particles that have the same size and shape as particles isolated frown plasma. One μg surface antigen can be yielded in 100ml culture.     

EXPRESSION AND SECRETION OF preS CONTAINING HEPATITIS B SURFACE ANTIGEN IN VACCINIA VIRUS SYSTEM

The expression and secretion of preS containing hepatitis B surface antigen in vaccinia virus system was investigated. The human TK~- 143 cells were infected with the recombinant vaccinia viruses vTMS-1 or vTLS-1. Cells infected with vTMS-1, which contains the preS2+S gene, produced preS2 containing middle HBsAg proteins. Similarly, cells produced preS1 containing large HBsAg proteins upon infection with vTLS-1, which carries the preS1+preS2+S gene. The expression products could be secreted and form 22 nm particles. They reacted specifically with anti-preS1 and/or anti-preS2 monoclonal antibodies, and exhibited pHSA-receptor (for polymerized human serum albumin) activity. In addition, the major S components of hepatitis B surface antigen were also present in the products expressed by vTMS-1 and vTLS-1.     

CLONING AND RESTRICTION MAPPING OF HUMAN HBV GENOME SEROTYPE adr

A Bam HI cleaved 3.2 Kb fragment from the HBV adr genome was cloned in E. coli, using pBR322 as vector. sixteen restriction sites from the action of Bam HI, Hind Ⅲ, Bgl Ⅰ, Bgl Ⅱ, Ava Ⅰ, Hinc Ⅱ, Sph Ⅰ, Xba Ⅰ, Xho Ⅰ and Sst Ⅱ were determined and mapped. No cleavage sites were found for Eco RI, Pst Ⅰ, Sma Ⅰ, Hpa Ⅰ, Kpn Ⅰ, Pvu Ⅱ and Sst Ⅰ.The restriction map for HBV adr is significantly different from those reported for the subtypes adw, ayw and adyw.     

化学合成的亮氨酸脑啡肽基因在大肠杆菌中的克隆和表达

化学合成的亮氨酸脑啡肽(LEK)基因与质粒pBR322重组,转化大肠杆菌,经过原位杂交筛选,限制性图谱分析和Southern杂交鉴定,获得一批LEK基因重组体。插入乳糖操纵子(1ac)启动基因控制LEK基因的表达。一个lac转录方向和LEK基因转录方向相同的表达质粒,pLE103,能在大肠杆菌中产生LEK。用放射免疫分析方法检测,LEK的产量可达每毫克细菌蛋白426毫微克。     

A CLONE OF HEPATITIS B VIRUS (SUBTYPE adr) DNA WITH A NEW HINDIII SITE

We have cloned the HBV genome subtype adr through its HindⅢ site into plasmid pBR.322. Twelve recombinant plasmids each with an insert of the HBV genome were obtained. The restriction map of one of the recombinants, plasmid pADR-H1, was analyzed. The location of the sites for BamHⅠ, BglⅠ, BglⅡ, SstⅡ, XbaⅠ and XhoⅠ in pADR-HⅠ were found to be the same as that in pADR-1, but the HindⅢ site of pADR-H1 is distinctly different from that of pADR-1. The BamHⅠ-HindⅢ fragment is 316 bp long in the gcnome of pADR-1. However, it is only 82 bp in pADR-Hl. No deletion of sequence between BamHⅠ and HindⅢ sites has been found in the HBV genome of pADR-H1, The sequencing data around the HindⅢ site of pADR-H1 in both pADR-H1 and pADR-1 show that there is a stretch of AAGTTT in pADR-1 compared to an AAGCTT in pADR-H1. In addition, other differences were found. There are three sites for AvaⅠ, four for HincⅡ, one for HpaⅠ and none for SphⅠ in the genome of pADR-H1 compared to four sites for AvaⅠ, three f     

一个带有新的Hind Ⅲ切点的HBVadr基因组克隆株

本文报道利用HBVadr DNA的Hind Ⅲ切点,将HBV DNA切开,插进pBR322的Hind Ⅲ位点,获得12株带HBVadr基因组的克隆株。限制性酶切分析,发现pADR-H1虽然也只有一个Hind Ⅲ切点,但它在基因组中的位置与pADR-1不同。pADR-1的HBV DNA中,其Hind Ⅲ-Bam HI片段的大小为315 bp,而pADR-H1中的相应片段只有82bp。序列分析表明,前者82 bp处的一段AAGTTT结构,在pADR-H1中变为AAGCTT而成了Hind Ⅲ识别位点。此外,限制性内切酶AvaⅠ,HincⅡ,HpaⅠ和SphⅠ对二克隆株的作用情况也有差别。本文探讨了这些差别的意义。     

一个HBsAg基因在大肠杆菌中的强表达系统

利用筛选启动基因的载体系统,从大肠杆菌染色体DNA中获得一个启动基因片段。在它的控制下HBsAg基因在大肠杆菌中有较强的表达。通过抗-HBs亲和层析柱分离纯化了表达产物。用琼脂糖双扩散免疫分析法和聚丙烯酰胺——SDS凝胶电泳对分离纯化的表达产物进行了鉴定。