水稻线粒体延伸因子OsEF-Tu基因的克隆及表达

Cloning and prokaryotic expression of a mitochondria elongation factor gene OsEF-Tu in rice

[目的]克隆水稻线粒体基因Os EF-Tu并进行表达。[方法]RT-PCR分别扩增Os EF-Tu基因c DNA的5’端和3’端序列,重叠PCR克隆Os EF-Tu的c DNA序列。生物信息学分析Os EF-Tu基因,构建水稻线粒体基因Os EF-Tu原核表达载体并转化BL21(DE3),IPTG诱导表达融合蛋白。[结果]Os EF-Tu蛋白与多个高等植物线粒体延长因子EF-Tu蛋白具有同源性,p GEX-4T1-Os EF-Tu重组子构建成功并表达出带有GST标签的Os EF-Tu融合蛋白。[结论]水稻Os EF-Tu在高等植物中较为保守,原核表达Os EF-Tu融合蛋白在28℃诱导条件下包涵体中表达量高。

[ Objective ] Clone rice mitochondria elongation gene OsEF - Tu and express the EF - Tu fusion protein. [ Meth- ods ] The 5＇ and 3＇ fragment of eDNA of the OsEF - Tu gene, respectively, were amplified by RT - PCR. Then, the full - length eDNA was amplified by over - lapping PCR. OsEF - Tu gene was analyzed by bioinformatics, and prokaryotic expression vector pGEX - 4T1 - OsEF - Tu was constructed. Then the fusion protein was induced by IPTG after transformed pGEX - 4T1 - OsEF - Tu into B[21 （ DE3 ）. [ Results] OsEF - Tu gene has a high similarity with several higher plants＇ mitochondria EF - Tu genes. The prokaryotic expression vector pGEX - 4T1 - OsEF - Tu has been constructed and expressed EF - Tu fusion protein that contains GST tag, successfully. [ Conclusion ] OsEF - Tu is conserved in mitochondrial of higher plants. The OsEF - Tu fu- sion protein induced in 28 ~C has a high yield in inclusion body.