湖北医药学院学报2023，Vol.42Issue(6)：608-611,619,封4,6.DOI:10.13819/j.issn.2096-708X.2023.06.006
过表达PD-L1慢病毒包装及稳转Ishikawa细胞株构建
Packaging of PD-L1 Overexpressing Lentivirus and Construction of Stable Lshikawa Cell Line
摘要
Abstract
Objective We aimed to construct a lentivirus expression vector carrying human programmed death protein 1 lig-and(PD-L1)and establish a stable Ishikawa cell line overexpressing PD-L1,providing a cell model for studying the bio-logical function mediated by PD-L1 in endometrial cancer.Methods Total RNA was extracted from Ishikawa cells,and the PDL1 CDS region sequence was cloned by RT-PCR.The PDL1 CDS region was then recombined onto a pCDH vector con-taining FLAG tags and purine resistance.After sequencing and identification,293T cells were co-transfected using a sec-ond-generation lentivirus packaging system to obtain the virus and infect Ishikawa cells.Stable transgenic strains were ob-tained through continuous screening with puromycin,and the expression of the target gene was detected by RT-qPCR and Western blot.Results PD-L1 CDS region sequence was successfully recombined into pCDH plasmid vector.After lentivirus packaging and infection,the mRNA and protein expression levels of PD-L1 were significantly up-regulated in Ishikawa.Conclusion This study successfully constructed a PD-L1 overexpressing lentiviral vector and established an Ishikawa stable transgenic strain overexpressing PD-L1 gene,providing a cell model for studying the role of PD-L1 in endometrial cancer.关键词
PD-L1/子宫内膜癌/慢病毒载体Key words
PD-L1/Endometrial carcinoma/Lentiviral vector引用本文复制引用
牛玉苗,黄浩,陈晓静..过表达PD-L1慢病毒包装及稳转Ishikawa细胞株构建[J].湖北医药学院学报,2023,42(6):608-611,619,封4,6.基金项目
国家自然科学基金面上项目(82273348) （82273348）
