重组原核表达载体pQE-30-luxS的构建与表达
详细信息Construction and expression of recombinant prokaryotic vector pQE-30-luxS
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摘要: 构建原核表达载体pQE-30-luxS,并在大肠杆菌中诱导表达融合蛋白。以植物乳杆菌KLDS1.0391的基因组DNA为模板,采用PCR方法扩增luxS基因,同时克隆到pMD18-T simple中,经鉴定正确后与表达载体pQE-30连接,将重组质粒pQE-30-luxS转化到E.coli M15中,异丙基-β-D-硫代吡喃半乳糖苷(IPTG)诱导表达,表达产物进行SDS-PAGE鉴定。结果表明原核表达载体pQE-30-luxS构建成功,测序证实插入的目的片段与已知luxS基因序列一致,且LuxS蛋白在大肠杆菌中成功表达。该结果为体外合成信号分子AI-2奠定了基础。Abstract: To construct the prokaryotic expression vector pQE- 30-luxS and express the fusion protein. The luxS gene was amplified by polymerase chain reaction ( PCR) using the DNA of Lactobacillus plantarum KLDS1.0391 as template, then it was cloned to vector pMD18-T simple.After the luxS gene was identified to be correct, the target gene was connected with expression vector pQE-30 and the recombinant plasmid pQE-30-luxS was transformed into E.coli M15.The recombinant expressed proteins induced by Isopropyl β- D-1- Thiogalactopyranoside ( IPTG) were analyzed and identified with SDS-PAGE.The results showed that the prokaryotic expression vector pQE-30- luxS was successfully constructed.The sequence of cloned luxS was identical with the known sequences, and LuxS protein was successfully expressed in E.coli M15.The results referred to above underlaid the relationship between the synthesis of AI-2 in vitro and the synthesis of bacteriocins of Lactobacillus plantarum KLDS1.0391.
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Keywords:
- luxS gene;recombinant plasmid;gene expression; /
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