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。     

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。     

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     

葡萄糖供应速率对大肠杆菌表达及分泌人表皮生长因子的影响

采用一株大肠杆菌BL 21(DE 3)[pBLMVL 2EGF]生产人表皮生长因子(hEGF),其中hEGF表达由PL启动子控制并通过phoA信号肽分泌到胞外。实验结果表明:诱导阶段以恒定比供应速率流加葡萄糖时,流加速率对hEGF的表达和分泌有很大影响。当葡萄糖比供应速率为0.122g/(g.h)时,hEGF表达水平最低,分泌效率最差,只有37.5%;随着葡萄糖比供应速率的增加,胞外和胞内hEGF比生产速率明显增加,当葡萄糖比供应速率为0.196 g/(g.h)时,单位菌体产生的hEGF水平最高(121.6 m g/g),其中分泌至胞外的hEGF占42%。此外,诱导前补充有机氮源有利于提高诱导初始hEGF的生产速率。     

THE COMPLETE NUCLEOTIDE SEQUENCE OF THE CLONED DNA OF HEPATITIS B VIRUS SUBTYPE adr IN pADR-1

The complete nucleotide sequence of the cloned hepatitis B virus DNA subtype adr in pADR-1 wasdetermined by Maxam and Gilbert's method. It is 3215 base pairs in size, which is 27 bp longer thanthe sequence of the adr pHBr330, as reported by Ono et al. The nucleotide difference between pADR-1and adr pHBr330 is about 2% while those between pADR-1 and adw as well as ayw are 9.3% and 9.7%respectively. In this paper, the heterogeneity and homogeneity of the S gene, the C gene and the othercoding regions in pADR-1 and in the other subtypes are compared and discussed.     

重组巴斯德毕赤酵母高密度培养中铵离子浓度的影响

采用Mut^s表型的重组毕赤酵母生产血管生长抑制素,表达阶段流加甘油—甲醇混合碳源以提高菌体密度和血管生长抑制素的表达水平,菌体密度可达174g／L,约是表达阶段采用甲醇为单一碳源的发酵过程的3倍。菌体密度的提高导致表达阶段发酵液中铵离子浓度下降很快,当发酵液中的铵离子浓度低至40mmol／L时,影响了血管生长抑制素的表达。改变pH调节方式并在发酵后期添加25mmol／L(NH4)2SO4使发酵液中铵离子浓度维持在150mmol／L以上,血管生长抑制素的表达产量达到108mg／L。     

克隆的adr亚型乙型肝炎病毒(pADR-1)DNA的全顺序

本文应用化学法测定了克隆的adr亚型HBV(_pADR-1)DNA全顺序,共长3215个碱基对(bp),比Ono等报道的adr_pHBr330长27个碱基对。_pADR-1和同一adr亚型的_pHBr330相比,核苷酸顺序中碱基的变异约为2％;而_pADR-1与adw和ayw的碱基差异则较大,分别为9.3％和9.7％。比较了_pADR-1与其它亚型的基因组中S基因、C基因,其它编码区的异同,并作了讨论。     

THE NUCLEOTIDE SEQUENCE OF SURFACE ANTIGEN GENE OF HEPATITIS B VIRUS SUBTYPE adr

The nucleotide sequence of the XhoI-BamHI fragment (1279bp), which contains the sur-face antigen gene (S gene) of HBVadr, was determined by Maxam and Gilbert's method.By comparing the differences both of the nuclectide sequence in the S gene and its codedamino acid sequence between adr and those reported for adw, ayw and adyw, some new var-iation sites were discovered. The differences were mainly distributed in the two hydrophilieregions. However, at those sites which might show biological function, there were no varia-tions among different subtypes, they are relatively conservative in heredity and evolution.Comparing the variation of the nucleotide sequence in the S gene region with that in thenon-S gene region, it is shown that the frequency of variation in the non-S gene regiondoubled that in the S gene region. Tim S gene region is more conservative.     

重组大肠杆菌BL21(pBAI)生产人干扰素α2b

通过培养重组大肠杆菌BL21(pBAI)表达人干扰素α2b(Human Interferon α2b,hIFNα2b).hIFNα2b表达由PL,启动子控制,通过升温至42℃诱导表达.本研究比较了分批培养和多种补料分批培养方式下hIFNα2b的生产,其中通过恒速流加葡萄糖,hIFNα2b的表达量达到6 540 mg/L,平均生产速率和比速率分别为546 mg/(L·h)和27 mg/(g·h).升温前1.5 h补充25 g酵母提取物,并以0.27 g/(g·h)的比速率供应葡萄糖,hIFNα2b的平均生产速率达到1 006 mg/(L·h),比生产速率为54 mg/(g·h),对有机氮源的得率提高到138 mg/g.