Construction of Hi FGF2 eukaryotic expression plasmids and its over-expression induced cell apoptosis
10.3969/j.issn.1001-1978.2014.11.011
- VernacularTitle:hi FGF2真核表达载体的构建及其高表达对细胞凋亡的影响
- Author:
Zhonglin CHEN
;
Hongyan JIANG
;
Xiaobing HONG
;
Zhonghua CHEN
;
Yanshan ZHENG
;
Han XU
;
Ganggang SHI
;
Zhanqin HUANG
- Publication Type:Journal Article
- Keywords:
Hi FGF2;
eukaryotic expression vector;
transfection;
apoptosis;
HEK293 cells;
flow cytometry
- From:
Chinese Pharmacological Bulletin
2014;(11):1535-1538
- CountryChina
- Language:Chinese
-
Abstract:
Aim To construct eukaryotic expressing plasmid of hi FGF2 ( high molecular weight isoform fi-broblast growth factor-2,hi FGF2) gene and to investi-gate its effect on apoptosis after its overexpression in HEK293 cells. Methods The DNA template primer was designed and synthesized. The pDsRed1-N1 plas-mids were digested by the restriction enzymes of Nhel and Hind III. The hi FGF2 was ligated with linearized pDsRed1-N1 by T4 DNA Ligase. The recombinant plasmid was identified by endonuclease digestion and sequenced. The recombinant hi FGF2 plasmid was transient transfected into HEK293 cells by Lipofectami-neTM 2000 Reagent. The transfection efficiency was de-tected by fluorescence inversion microscope. The cell apoptosis was detected by Annexin V-FITC/PI apopto-sis detection kit with flow cytometry analysis. Results The pDsRed1-N1 eukaryotic expression vector was consistent with the design. The recombinant hi FGF2 plasmid was transfected in HEK293 cells. The trans-fection rate was more than 70%. The FITC/PI dyeing rate in hi-FGF2 over-expression HEK297 cells was a-bout ( 29. 12 ± 2. 81 )%. Conclusions pDsRed1-N1 eukaryotic expression vector is successfully constructed and transfected into HEK293 cells. Over-expression of hi FGF2 induces cell apoptosis.
