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目的:克隆脂肪储存小滴蛋白5(LSDP5)基因,并在大肠杆菌中表达。方法:利用PCR技术从小鼠肝脏cDNA中扩增LSDP5基因,并将其克隆到pMD18-T载体中,进行测序。同时,通过疏水性和二级结构分析,将LSDP5的片段克隆到原核表达载体pEGX-4T-1,构建GST融合表达质粒pEGX-LSDP5,在大肠杆菌BL21中进行表达。结果:成功地克隆了LSDP5基因,DNA测序证实,与GenBank公布的序列一致。含LSDP5片段的pGEX-LSDP5基因表达质粒在大肠杆菌经IPTG诱导后,能够表达相对分子质量(Mr)40000的融合蛋白。结论:获得了LSDP5全长基因,成功构建了原核表达质粒pEGX-LSDP5,并在大肠杆菌中得到表达,为其相关研究奠定了一定的基础。
Objective: To clone the fat storage protein 5 (LSDP5) gene and express it in E. coli. Methods: LSDP5 gene was amplified from mouse liver cDNA by PCR and cloned into pMD18-T vector for sequencing. At the same time, the fragment of LSDP5 was cloned into the prokaryotic expression vector pEGX-4T-1 through hydrophobic and secondary structure analysis, and the GST fusion expression plasmid pEGX-LSDP5 was constructed and expressed in E. coli BL21. Results: The LSDP5 gene was successfully cloned and verified by DNA sequencing. The results showed that the sequence was identical with that published in GenBank. The pGEX-LSDP5 gene expression plasmid containing the LSDP5 fragment was able to express a fusion protein with a relative molecular mass (Mr) of 40,000 after induced by IPTG in Escherichia coli. CONCLUSION: The LSDP5 full-length gene was obtained and the prokaryotic expression plasmid pEGX-LSDP5 was successfully constructed and expressed in E. coli, which laid a foundation for its related research.