超表达葡萄糖脱氢酶促进大肠杆菌生长
Overexpression glucose dehydrogenase can promote the growth of recombinant Escherichia coli
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摘要: 通过PCR扩增了肺炎克雷伯氏菌（Klebsiella pneumoniae）和大肠杆菌（Escherichia coli）各自的葡萄糖脱氢酶基因KPgdh和ECgdh,构建了表达载体p ET-28a-KPgdh和p ET-28a-ECgdh,转化大肠杆菌E.coli BL21,获得了重组菌BL21（p ET-28a-KPgdh）和BL21（p ET-28a-ECgdh）。经IPTG诱导,聚丙烯酰胺凝胶电泳显示重组菌实现了葡萄糖脱氢酶的高效表达,且葡萄糖脱氢酶活性分别是空质粒对照菌E.coli BL21（p ET-28a）的8.5倍和12倍。重组菌的生长速度快于空质粒对照菌E.coli BL21（p ET-28a）,并与原代菌E.coli BL21持平。如果扣除质粒复制造成的代谢负荷,过表达葡萄糖脱氢酶促进了大肠杆菌的生长。Abstract: The genes encoding glucose dehydrogenase(gdh) from Klebsiella pneumoniae and Escherichia coli were amplified by polymerase chain reaction(PCR). Subsequently,the corresponding expression vectors p ET-28a-KPgdh and p ET-28a-ECgdh were constructed and cloned into E. coli BL21,resulting in two recombinant strains E. coli(p ET-28a-KPgdh) and E. coli(p ET-28a-ECgdh). SDS-PAGE analysis showed the high level expression of both KPgdh and ECgdh upon IPTG induction. Compared with the control strain E. coli(p ET-28a)which harbored an empty vector,the recombinant strains E. coli(p ET-28a-KPgdh) and E. coli(p ET-28a-ECgdh) consumed more glucose,and the gdh activity toward glucose increased by 8.5-and 12-fold,respectively. If plasmid burden was not counted,overall these results indicated that gdh overexpression facilitated cell growth.
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