Expression and functional analysis of CRT-E2-EGFP fusion protein in B16 cells
10.3321/j.issn:1007-8738.2009.10.010
- VernacularTitle:CRT-E2-EGFP表达、胞内定位及其对B16细胞凋亡的影响
- Author:
Jianlin YANG
;
Changbai LIU
;
Danli LI
;
Li HAN
- Publication Type:Journal Article
- Keywords:
human papillomavirus E2 protein;
calreticulin;
apoptosis
- From:
Journal of Cellular and Molecular Immunology
2009;25(10):897-899
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To detect the expression of the Calreticulin and HPV E2 Fusion Protein in B16, and study the effects on proliferation and apoptosis of B16 cell lines in vivo. METHODS: To construct eukaryotic fluoresce expression vector pEGFP-CRT-E2, pEGFP-CRT and pEGFP-E2. Then the recombinant plasmids were transfected into B16 cells by Lipofectamine 2000. The expression of proteins was detected by Western blot. The location of different GFP fusion proteins in B16 was tested by inverted fluoresce microscope. Flow cytometry was applied to detect the effects of fusion proteins on the growing of B16 and then the apoptosis effects of B16 induced by different proteins were observed. RESULTS: The correctly constructed recombinant plasmid pEGFP-CRT-E2 and the expression of CRT-E2 gene could be detected in B16 cells. Apoptosis of B16 cells could be detected after the transient transfection. Meanwhile, the apoptosis rate of B16 cells transfected by pEGFP-CRT-E2 was much higher than that of control cells. And cell cycle G1/G0 arrest was also found (P < 0.01). CONCLUSION: The eukaryotic expression plasmid pEGFP-CRT-E2 is successfully constructed and it could correctly express the fusion protein in B16 cells. And the B16 cells transfected by plasmid pEGFP-CRT-E2 could induce apoptosis.
