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枯草芽孢杆菌XF1菌株中XFsacA基因的克隆及功能验证

DOI: 10.7671/j.issn.1001-411X.2012.02.007, PP. 154-158

Keywords: 枯草芽孢杆菌XF1,蔗糖,代谢,基因克隆

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Abstract:

枯草芽孢杆菌BacillussubtilisXF1是从土壤中分离的一株利用蔗糖快、能高效防治根肿病的专利菌株.为探明其高效的蔗糖代谢机制,用PCR从该菌中扩增到蔗糖代谢关键基因蔗糖-6-磷酸水解酶基因(XFsacA基因),其编码的蛋白质氨基酸序列与枯草芽孢杆菌B168菌株中的XFsacA基因有97%的相似性,且发生了12个氨基酸突变.将该基因中约1.4kb的编码框序列连接到表达载体pQE30上,构建了重组表达质粒pQE30-XFsacA,该质粒转入到不能利用蔗糖的EscherichiacoliBL21中,后者能在以蔗糖为唯一碳源的M9无机离子培养基中正常生长,并表达出了一条约54000蛋白条带.结果预示XFsacA基因氨基酸序列的替代可能是XF1高效利用蔗糖的基础,同时XFsacA基因的克隆与其在E.coli中的表达研究,为利用蔗糖发酵的工程E.coli构建奠定了基础.

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