SEF14菌毛基因操纵子在肠炎沙门氏菌和D-群沙门氏菌的变异分析

【目的】本文旨在探索SEF14菌毛特异性表达于D-群沙门氏菌,特别是肠炎沙门氏菌以及都柏林沙门氏菌的原因。【方法】应用PCR扩增以及序列测定检测了18株鸡白痢沙门氏菌,11株肠炎沙门氏菌以及1株都柏林沙门氏菌标准株中sefA,sefD和sefR基因序列,并分析比对其序列变异。【结果】以11株肠炎沙门氏菌以及1株都柏林沙门氏菌染色体DNA为模板能成功扩增sefA,sefD以及sefR基因;从18株鸡白痢沙门氏菌中均能成功扩增sefA基因,但只有分离于1980年之前的7株分离菌能成功扩增sefD和sefR基因,而另11株1980年后分离菌PCR扩增sefD和sefR基因却无任何产物。比对PCR扩增产物测序结果发现,11株肠炎沙门氏菌以及1株都柏林沙门氏菌株中sefA,sefD以及sefR基因序列和已发表的序列(GenBank登录号为L11008,U07129和AF233854)100%同源;7株鸡白痢沙门氏菌sefD基因测序结果表明,在196位点处发生碱基缺失,造成移码突变,提前于氨基酸残基71位点处产生终止密码子。优化菌毛表达条件,体外抽提和纯化菌毛并进一步试验证明:肠炎沙门氏菌以及都柏林沙门氏菌体外能很好表达SEF14菌毛,但鸡白痢沙门氏菌在相同培养条件下却无任何表达迹象。【结论】SEF14菌毛操纵子亚单位基因sefA,sefD以及调节基因sefR在不同沙门氏菌中的变异情况可能是SEF14菌毛局限性表达的原因之一。     

在兽疫链球菌中表达vgb基因和HA合成基因提高透明质酸产量

透明质酸(HA)是一种在医药及化妆品领域具有广泛应用的天然粘多糖。兽疫链球菌(Streptococcuszooepidemicus)是工业上生产透明质酸的菌种之一。透明颤菌血红蛋白(VHb)具有增强细胞摄氧的作用。对生产透明质酸的兽疫链球菌进行了基因改造:将兽疫链球菌HA的合成基因hasABC以及合成透明颤菌血红蛋白的vgb基因(Vitreoscillahemoglobingene,vgb)分别或同时插入阳性菌表达质粒pEU308中,通过电转化导入兽疫链球菌中。通过一氧化碳(CO)差光谱检测到了VHb的表达。在摇瓶实验中,同时带有hasABC和vgb基因的重组菌比野生菌的透明质酸产量提高了30％。而在发酵罐中,带有这2个基因的重组菌的透明质酸产量达到了6．9g／L,高于重组菌5．5g／L的产量。实验结果表明,vgb基因的存在促进了细胞的生长,hasABC操纵子的过表达增强了透明质酸的合成。首次将VHb导入兽疫链球菌中,获得了表达,并证明其对菌体生长及透明质酸合成有促进作用。通过研究,VHb将可以在阳性菌中获得更广泛的应用。     

Synergistic effect ofrib80 andhit Mutations on riboflavin biosynthesis and iron transport in the yeastPichia guilliermondii

Mutant strains of the yeastPichia guilliermondii, carrying bothrib80 andhit mutations in a haploid genome, were derived from previously obtained strains with defectiverib80 orhit genes, exerting negative control of the riboflavin biosynthesis and iron transport inPichia guilliermondii. The double mutant rib80hit strains exhibited an increased level of riboflavin biosynthesis and higher activities of GTP cyclohydrolase and riboflavin synthetase. Iron deficiency caused an additional increase in riboflavin overproduction. These results suggest the synergistic interaction of therib80 andhit mutations. A combination of both mutations in a single genome did not affect iron assimilation by the cells: ferrireductase activity, the rate of⁵⁵Fe uptake, and the iron content in cells of the double mutants remained at the level characteristic of the parent strains.     

霍乱毒素A基因内部翻译调控元件具有翻译起始功能

通过大肠杆菌体外转录－体外翻译系统,证明霍乱毒素Ａ基因内部的翻译调控元件具有翻译起始功能,且其翻译起始效率较ｃｔｘＡ基因高得多,当ｃｔｘＡ的起始密码突变时,从该元件起始的翻译效率下降,说明基因内翻译起以ｃｔｘＡ翻译起始的调控。结果进一步证实了霍乱毒素Ａ、Ｂ亚工比例表达调控的翻译弱化－翻译偶联机理。     

Characterization of the str operon genes from Spirulina platensis and their evolutionary relationship to those of other prokaryotes

Summary A 5.3 kb DNA segment containing the str operon (ca. 4.5 kb) of the cyanobacterium Spirulina platensis has been sequenced. The str operon includes the structural genes rpsL (ribosomal protein S12), rpsG (ribosomal protein S7), fus (translation elngation factor EF-G) and tuf (translation elongation factor EF-Tu). From the nucleotide sequence of this operon, the primary structures of the four gene products have been derived and compared with the available corresponding structures from eubacteria, archaebacteria and chloroplasts. Extensive homologies were found in almost all cases and in the order S12>EF-Tu>EF-G>S7; the largest homologies were generally found between the cyanobacterial proteins and the corresponding chloroplast gene products. Overall codon usage in S. platensis was found to be rather unbiased.     

The meta cleavage operon of TOL degradative plasmid pWWO comprises 13 genes

Summary The meta-cleavage operon of TOL plasmid pWWO of Pseudomonas putida encodes a set of enzymes which transform benzoate/toluates to Krebs cycle intermediates via extradiol (meta-) cleavage of (methyl)catechol. The genetic organization of the operon was characterized by cloning of the meta-cleavage genes into an expression vector and identification of their products in Escherichia coli maxicells. This analysis showed that the meta-cleavage operon contains 13 genes whose order and products (in kilodaltons) are The xyIXYZ genes encode three subunits of toluate 1,2-dioxygenase. The xylL, xyIE, xyIG, xylF, xylJ, xylK, xylI and xylH genes encode 1,2-dihydroxy-3,5-cyclohexadiene-1-carboxylate dehydrogenase, catechol 2,3-dioxygenase, 2-hydroxymuconic semialdehyde dehydrogenase, 2-hydroxymuconic semialdehyde hydrolase, 2-oxopent-4-enoate hydratase, 4-hydroxy-2-oxovalerate aldolase, 4-oxalocrotonate decarboxylase and 4-oxalocrotonate tautomerase, respectively. The functions of xyIT and xylQ are not known at present. The comparison of the coding capacity and the sizes of the products of the meta-cleavage operon genes indicated that most of the DNA between xyIX and xyIH consists of coding sequences.