Construction of gastric cancer related gene GCRG213 eukaryotic expression vector and its effect on growth of gastric cancer cells
- VernacularTitle:胃癌相关基因GCRG213真核表达载体的构建及其对胃癌细胞生长特性的影响
- Author:
Lili GAO
;
Benyan WU
;
Mengwei WANG
- Publication Type:Journal Article
- Keywords:
stomach neoplasms;
genes, GCRG213;
transfection;
apoptosis
- From:
Medical Journal of Chinese People's Liberation Army
2001;0(09):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effect of gene GCRG213 transfection (sense, anti-sense) on growth of gastric cancer cell MKN45. Methods The sense and anti-sense fragment of GCRG213 were obtained by PCR. They were cloned into eukaryotic expression vector pcDNA3.1(+). The recombinant plasmid pcDNA3.1-a, pcDNA3.1-b and the vector pcDNA3.1 were transfected separately into MKN45 cells conducted by lipofectamine~ TM 2000. Expression of GCRG213 was assessed with semi-quantitative RT-PCR and Western Blot. The growth graph was plotted by the methods of cell counting of three steady transfected cells. FACS was used to determine the cell cycle, and Annexin V FITC/PI bi-labeling method was used to determine the cell apoptosis. Results By sequencing, sense GCRG213 and anti-sense GCRG213 were proved to be successfully cloned into pcDNA3.1. Transfecting the sense vector (pcDNA3.1-a) into the MKN45 significantly increased the expression of GCRG213, both in mRNA level and protein level. Transfecting the anti-sense vector (pcDNA3.1-b) into the MKN45 significantly decreased the expression of GCRG213, both in mRNA level and protein level. The growth of pcDNA3.1-a transfected cells was faster than that of vector transfected cells, and the cell apoptosis decreased. But the growth of pcDNA3.1-b transfected cells was slower in multiplication than that of vector transfected cells, and the cell apoptosis was increased. Conclusion Stable transfection showed that GCRG213 promoted cell to grow, inhibited the cell apoptosis. GCRG213 might be a new promoter to tumor.
