华西口腔医学杂志2016，Vol.34Issue(4)：398-403,6.DOI:10.7518/hxkq.2016.04.016
上皮膜蛋白1真核表达载体的构建及其对人舌鳞状细胞癌细胞迁移和侵袭的影响
Construction of epithelial membrane protein 1 eukaryotic expression vectorand its influence on migration and invasion of human oral tongue squamous carcinoma cells
摘要
Abstract
ObjectiveThis study aimed to construct a eukaryotic expression vector pEGFP-N1-EMP1 of epithelial mem-brane protein 1 (EMP1) and investigate its influence on migration and invasion of human oral tongue squamous carcinoma cells.Methods The human EMP1 gene was amplified by reverse transcription polymerase chain reaction and then ligated into the pEGFP-N1 vector by double restriction endonuclease digestion to construct pEGFP-N1-EMP1 recombinant plasmid. After sequencing identification, pEGFP-N1-EMP1 recombinant plasmid and pEGFP-N1 plasmid were transfected into human oral tongue squamous carcinoma Tb3.1 cell line. The expression of green fluorescent protein in cells was observed after transfection using an inverted fluorescence microscope. The overexpression of EMP1 mRNA was identified at 24, 48, and 72 h after transfection by real-time fluorescence quantitative polymerase chain reaction. The effect of EMP1 overexpression on migration and invasion of Tb3.1 cells was detected by Transwell assay.Results The full-length EMP1 gene sequence was successfully obtained. Sequence analysis showed that the EMP1 gene was inserted into the pEGFP-N1 vector correctly. Green fluorescence was observed in the transfected cells under fluorescence microscopy. The results of real-time fluorescence quantitative polymerase chain reaction indi-cated that the expression of EMP1 at 24 h after pEGFP-N1-EMP1 transfection was significantly higher than the other groups. Transwell assays indicated that overexpression of the EMP1 gene could significantly inhibit the migration and invasion ability of Tb3.1 cells.Conclusion The eukaryotic expres-sion vector of EMP1 was successfully constructed, and EMP1 overexpression was confirmed to inhibit the migration and inva-sion of oral tongue squamous carcinoma cellsin vitro. This study laid a foundation for further investigation on the influence of the EMP1 gene on the metastasis of oral tongue squamous carcinoma and its molecular mechanism.关键词
上皮膜蛋白1/舌鳞状细胞癌细胞/载体/转染/细胞迁移/细胞侵袭Key words
epithelial membrane protein 1/oral tongue squamous carcinoma cell/vector/transfection/cell migration/cell invasion分类
医药卫生引用本文复制引用
代晓华,张军,邹慧儒,连小丽,李燕妮,王冠华,颜艳..上皮膜蛋白1真核表达载体的构建及其对人舌鳞状细胞癌细胞迁移和侵袭的影响[J].华西口腔医学杂志,2016,34(4):398-403,6.基金项目
天津市卫计委基金 （）
