摘要
目的:构建颗粒溶解素(granulysin)cDNA真核表达载体。方法:采用逆转录聚合酶链反应(RT-PCR)技术从健康志愿者外周血单个核细胞(PBMC)中扩增出颗粒溶解素蛋白cDNA,经限制性核酸内切酶消化后,插入真核表达载体pcDNA3.1(-)的相应酶切位点,将此重组质粒进行序列测定,并将重组质粒转染COS7细胞,进行基因表达鉴定。结果:重组质粒插入基因序列测定证实为颗粒溶解素cDNA,并能在COS7细胞稳定表达。结论:成功构建颗粒溶解素真核表达载体,为进一步研究其抗结核作用奠定了基础。
Objective: To construct eukaryotic expression vector of Granulysin. Methods: Granulysin cDNA was amplified from PBMC of healthy volunteer by RT-PCR, and correctly inserted into corresponding sites of eukaryotic expression vector pcDNA3. 1 (-) after restriction endonuclease digestion, and the recombinant plasmid was transfected into COS7 cells, gene expression was determined by RT-PCR. Results: The gene fragment inserted into the vector pcDNA3.1(-) was confirmed by nucleotide sequencing, and the mRNA of the recombinant plasmid was successfully expressed in mammalian cells. Conclusion: The successful construction of recombinant eukaryotic expression vector based on the cDNA of granulysin, provides the experiment basis for further researching.
出处
《武汉大学学报(医学版)》
CAS
2005年第6期804-807,共4页
Medical Journal of Wuhan University
基金
湖北省科技厅重大课题资助项目(301121028)