Construction of pIRES2-EGFP-PELP1 eukaryotic expression vector and its expression in human periodontal ligament stem cells

VernacularTitle:真核表达载体pIRES2-EGFP-PELP1的构建及其在人牙周膜干细胞中的表达
Author: Yan YAN ¹ ; Zhong-Ying NIU ; Chu-Hua TANG ; Liang SHI ; Shao-Yan SI ; Shu-Jun SONG
Author Information

1. 解放军306医院全军口腔疾病诊治中心
Keywords: Periodontal ligament; Stem cells; Gene cloning; Vector
From: Chinese Journal of Stomatology 2013;48(z1):83-86
CountryChina
Language:Chinese
Abstract: Objective To construct an over-expressing plasmid containing full length of PELP1 gene and investigate the characterization of PELP1 expressions in different tissues.Methods Full length PELP1 gene was cloned using RT-PCR product of MCF-7 total RNA as the template.The PELP1 gene fragament was amplified by PCR and two enzyme sites EcoR I and Xho I were added.Then PELP1 gene was cloned into the pIRES2-EGFP palsimd (containing EcoR I and Xho I enzyme sites) to make reconstructed plasmid pIRES2-EGFP-PELP1.Enzyme digestion and sequencing were employed to assess the integrity and correctness of PELP1 gene cloned.The plasmid pIRES2-EGFP-PELP1 was transfected into human periodontal ligament stem cells,and the expression of PELP1 was examined by quantitative real-time-PCR or Western-blotting.Results The integrity and correctness of PELP1 gene were confirmed by digestion and sequencing.Conclusions The full length of PELP1 gene from MCF-7 cell was successfully cloned and over-expression plasmid pIRES2-EGFP-PELP1 constructed.