产肠毒素大肠杆菌987P菌毛fasG亚单位的优化表达

本试验选用两种翻译起始区不同的表达载体pET28a(+)、pGEX-KG和两种对稀有密码子效应不同的表达宿主菌BL21(DE3)、Rossetta(DE3)，构建出四种重组表达菌（pET-fasG BL21(DE3), pET-fasG Rossetta(DE3), pKG-fasG BL21(DE3)和pKG-fasG Rossetta(DE3)）来考查翻译起始区的二级结构和稀有密码子对fasG基因表达的影响。对重组表达菌进行诱导表达的结果表明，翻译起始区的二级结构和稀有密码子的存在都能影响到目的基因的表达，通过优化翻译起始区的二级结构或优化稀有密码子都能提高目的蛋白的表达量。由于pKG-fasG优化了mRNA翻译起始区的二级结构，同时Rossetta(DE3)缓解了稀有密码子对目的蛋白表达的影响，pKG-fasG Rossetta(DE3)目的蛋白的表达量达到16%，较pET-fasG BL21(DE3)的表达量（8%）提高了100%。试验结果表明，对外源基因的表达，选择合适的表达载体和宿主菌，对获得较高的表达量是非常重要的。

Optimizing Expression of the Subunit fasG of 987P Fimbriae of Enterotoxigenic Escherichia coli(ETEC)

In our experiment, two E. coli expression vectors(pET28a(+)、pGEX-KG) with different translation intiation regions(TIR) and two E. coli expression strains(BL21(DE3)、Rossetta(DE3)) with different tRNA profiles had been used to obtain four bioengineered bacterium of fasG gene. The aim of the experiment was to examine the influence of secondary structures of TIR and the rare codens to the expression of fasG gene. The result of inducing expression of the bioengineered bacteriums showed that the secondary structures of TIR and the existence of rare codens can both influence the expression of target genes, optimizing either could both enhance the expression lever of target gene. The plasmid pKG-fasG had optimized the secondary structures of TIR and Rossetta(DE3) had overcome the rare condons’effects, so the expression lever of pKG-fasG Rossetta(DE3) achieved 16%, add to 100% in comparison with pET-fasG BL21(DE3)(8%). The result of this experiment indicated that the selection of appropriate expression vectors and strains was very important in the expressions of heterologous genes.