Construction of human BTG2 eukaryotic expression vector with FLAG tag and its expression in HeLa cells
10.13481/j.1671-587x.20140605
- VernacularTitle:携带FLAG标签的人BTG2真核表达载体的构建及其在HeLa细胞中的表达
- Author:
Jinxia ZHAO
;
Zhiping WANG
;
Yan TAO
;
Zhenhua HE
;
Qi GUO
;
Mei HONG
- Publication Type:Journal Article
- Keywords:
B-cell translocation gene 2;
tumor suppressor gene;
FLAG tag;
eukaryotic expression vector;
fusion protein
- From:
Journal of Jilin University(Medicine Edition)
2014;(6):1149-1154
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct an eukaryotic expression vector of human B-cell translocation gene 2 (BTG2), to express the FLAG-tagged BTG2 protein in HeLa cells,and to supply an experimental tool for investigating the function of BTG2 gene.Methods The full-length BTG2 fragment was obtained by PCR and inserted into the multiple cloning site of pcDNA3.1 (+)vector. Oligo DNA encoding FLAG tag was designed and inserted into pcDNA3.1(+)-BTG2 to construct another vector pcDNA3.1(+)-FLAG-BTG2.The HeLa cells were divided into pcDNA3.1(+)empty vector group,pcDNA3.1(+)-BTG2 group and pcDNA3.1(+)-FLAG-BTG2 group.The HeLa cells were transfected with recombinant plasmids.Western blotting using anti-FLAG antibody was performed to detect the expression of FLAG-BTG2 protein in HeLa cells.Results The sequence of the vector was verified by both BamH Ⅰ endonuclese digestion and DNA sequencing. The Western blotting analysis confirmed that FLAG-fused BTG2 was detected in pcDNA3.1(+)-FLAG-BTG2 group but not in empty vector or pcDNA3.1(+)-BTG2 groups. Conclusion The eukaryotic expression vector pcDNA3.1(+)-FLAG-BTG2 is successfully constructed and FLAG-tagged BTG2 protein is expressed in HeLa cells.
