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高世同, 刘会娟, 张仁利, 秦莉, 耿艺介, 黄达娜, 吴少庭. 金葡菌野生株肠毒素B基因克隆及分子特征[J]. 中国公共卫生, 2006, 22(1): 72-73. DOI: 10.11847/zgggws2006-22-01-38
引用本文: 高世同, 刘会娟, 张仁利, 秦莉, 耿艺介, 黄达娜, 吴少庭. 金葡菌野生株肠毒素B基因克隆及分子特征[J]. 中国公共卫生, 2006, 22(1): 72-73. DOI: 10.11847/zgggws2006-22-01-38
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GAO Shitong, LIU Huijuan, ZHANG Renli, . Cloning and molecular characteristics of enterotoxin B gene from wild staphylococcus aureus strain[J]. Chinese Journal of Public Health, 2006, 22(1): 72-73. DOI: 10.11847/zgggws2006-22-01-38
Citation: GAO Shitong, LIU Huijuan, ZHANG Renli, . Cloning and molecular characteristics of enterotoxin B gene from wild staphylococcus aureus strain[J]. Chinese Journal of Public Health, 2006, 22(1): 72-73. DOI: 10.11847/zgggws2006-22-01-38
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金葡菌野生株肠毒素B基因克隆及分子特征

Cloning and molecular characteristics of enterotoxin B gene from wild staphylococcus aureus strain

    摘要
  • 摘要:
      目的   克隆金黄色葡萄球菌野生株S407030肠毒素B(SEB)基因, 并分析其分子特征.
      方法   根据Genebank中金黄色葡萄球菌S6株肠毒素B基因设计1对引物, 采用PCR法从金黄色葡萄球菌野生株S407030基因组中扩增目的基因, 将目的片段与pMD-18T载体连接构建重组子pMD-18T-SEB, 并转化E.coli DH5a;阳性克隆以双酶切鉴定后, 双脱氧末端终止法作序列测定分析, 并以ExPasy Proteoic Tools蛋白分析软件预测其编码SEB蛋白的二级结构.
      结果   从金黄色葡萄球菌基因组DNA中扩增出约705bp的基因片段, 所构建的阳性克隆重组子PCR及双酶切鉴定与预期结果一致; 测序结果表明, 克隆的SEB基因含705个碱基, 与S6株序列相比无碱基的插入或缺失, 同源性为98.4%;存在11个碱基变异, 其中7个为无义突变, 4个为有义突变(突变位点位于第364, 699, 899, 988位, 编码氨基酸变化分别为:Ser→Ala、Gln→His、Asn→Ser、Met→Leu); 所预测的2菌株SEB蛋白的二级结构基本相同.
      结论   本实验成功克隆了金葡菌野生株S407030SEB基因, 其在不同菌株间相对保守; 由SEB基因点突变引起的个别氨基酸的改变对SEB生物活性与毒力的影响有待进一步研究.

     

    Abstract:
      Objective   To clone the enterotox in B gene from a wild stap hylococcus aureus strain S407030, and to analyze its molecular character istics.
      Methods   The Staphylococcal enterotoxin B gene was amplified using PCR mehtod, and ligated with the pMD-18T plasmid to construct the recombinant rpMD-SEB, Which was transformed into E.coli DH5A.The positive bacteria clones were identifiedty PCR and double enzymes dieg est ion methods.The inserted SEB gene fragment was finally sequenced and its deduced SEB protein secondary structure analyzed.
      Results   705 bp SEB gene fragement was correctly cloned.Compared with S.aureus strain S6, the overall nucleo tides identity was 98.4%.Among the 11 differ ent nucleotids, 4 nucleotids leading to the encoding aminor esidues different.Ho wever, the heterog eneity of nucleotides had no significant influence on the deduced SEB protein secondary structures between the S.aureus strain S407030 and S6.
      Conclusion   The Staphylococcal Enterotox in B gene of S.aureus strain S407030 was successfully cloned, and it was relatively conserved among different S.aureus strains.The problem that the heterog eneity of nucleotides of SEB gene lead to the variety of SEB virulence deserved further study.

     

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