转化医学杂志2017，Vol.6Issue(1)：12-15,4.DOI:10.3969/j.issn.2095-3097.2017.01.003
可溶性白细胞分化抗原40配体原核表达载体的构建与鉴定
Construction and identification of soluble cluster of differentiation antigen 40 ligand prokaryotic expression vector
摘要
Abstract
Objective To construct containing human soluble cluster of differentiation anti-gen 40 ligand ( sCD40L) recombinant plasmid pGEX-4T-sCD40L. Methods The gene fragment of human leukocyte sCD40L was amplified by reverse transcription-polymerase chain reaction ( RT-PCR) . The fragment was ligated into T vector and ligated with 5-bromo-4-chloro-3-indolyl-β-D-ga-lactoside ( X-gal ) and isopropyl-β-D-thiogalactopyranoside ( IPTG ) . The positive clones were screened by IPTG. The positive clones were digested with BamHⅠand XhoⅠ, then ligated with pGEX-4T prokaryotic expression vector, and identified by restriction enzyme digestion and sequen-cing. Results The target DNA fragment of 474 bp was amplified by the complementary DNA ( cDNA) of L02 cells. The recombinant plasmid pGEX-4T-sCD40L was constructed by ligating the target frag-ment with T vector and screening for blue-white. The recombinants were obtained by transformation and screening, and the sequences were confirmed by restriction enzyme digestion and gene sequen-cing. Conclusion The prokaryotic expression vector pGEX-4T-sCD40L has been successfully con-structed, which lays a foundation for further study on the role of sCD40L in the diagnosis of hepato-cellular carcinoma ( HCC) .关键词
可溶性白细胞分化抗原40配体/原核表达载体/融合蛋白Key words
Soluble cluster of differentiation antigen 40 ligand ( sCD40L)/Prokaryotic ex-pression vector/Fusion protein分类
医药卫生引用本文复制引用
张志强,乔录新,吴涛,刘东杰,纪乐,陈德喜,吴刚..可溶性白细胞分化抗原40配体原核表达载体的构建与鉴定[J].转化医学杂志,2017,6(1):12-15,4.基金项目
国家自然科学基金资助项目(81361120401)；首都卫生发展科研专项项目(首发2014-1-1151) （81361120401）
