海藻糖合酶产生菌筛选、鉴定及目的基因克隆

Screening of Trehalose Synthase Producing Strain and Cloning of Trehalose Synthase Gene

目的:为筛选出一株产海藻糖合酶的菌株,并以此菌的全DNA为模板,克隆出产海藻糖合酶的目的基因片段。方法:实验过程中采用了常规筛选菌种、快速提取细菌全基因、显微镜观察菌种、热启动PCR技术、电泳纯化回收基因片段、EcoRⅠ和HindⅢ双酶切鉴定目的基因片段等方法。结果:在电镜下可观察到有芽孢、杆菌;菌株16S rRNA基因扩增产物共计1490个碱基;PCR方法扩增出阳性克隆大约1700bp的基因片段。结论:通过生理、形态、结构特征分析及16S rRNA基因全序列比较得出结论:筛选到一株短小芽孢杆菌;PCR扩增出阳性克隆片段,全长1722bp,为实验所要的编码海藻糖合酶的基因片段。

Objective：The strain which could produce trehalose synthase was screened from preserved microorganism in laboratory,The DNA was prepared from the bacillus and the gene of trehalose synthase was cloned.Methods：Some methods had been used that were screening of trehalose synthase producing strain,preparing the DNA from the bacillus,polymerase chain rection,purifying the gene fragment, cleaving the recombinant plasmid by the restriction endonucleases EcoRⅠand HindⅢ.Results：The spore stain and bacterium had been found under the transmission electron microscopic, A 1 490bp gene fragment was amplified from the genomic DNA with the primers of 16S rRNA, The gene of trehalose synthase from the screened bacterium was mnplified by the polymerase chain rection was about 1700bp. Conclusion： The bacterium had been screened whose enducellular enzymes could pnxtuce trehalose and the morphological, cultural and physiological characteristics of trehalose producing bacterium were described, A 1722bp gene fragment which was amplified from the genomic DNA with specific primers, and cloned into the PUCm - T vector was the wanted gene fragment.