Construction of a recombinant adenovirus co-expressing bone morphogenic proteins 9 and 6 and its effect on osteogenesis in C3H10 cells
10.3969/j.issn.1673-4254.2013.09.04
- VernacularTitle:骨形态发生蛋白9、6双表达腺病毒载体的构建及其成骨诱导作用
- Author:
Xiaohong DIE
1
;
Qing LUO
;
Cong CHEN
;
Guangjin LUO
;
Quan KANG
Author Information
1. 重庆医科大学附属儿童医院儿童发育疾病研究教育部重点实验室//儿科学重庆市重点实验室// 重庆市(儿童发育重大疾病诊治与预防)国际科技合作基地
- Keywords:
co-expression vector;
bone morphogenic protein 9;
bone morphogenic protein 6;
recombinant adenovirus;
osteogenesis
- From:
Journal of Southern Medical University
2013;(9):1273-1279
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct a recombinant adenovirus co-expressing bone morphogenic protein (BMP) 9 and BMP6 and observe its effect on the osteogenesis in C3H10 cells. Method The full-length sequences of BMP9 and BMP6 were amplified from AdEasy vector by PCR and cloned into the shuttle plasmid pASG2 vector to construct the co-expression shuttle plasmid pASG2-BMP9, 6 followed by homologous recombination with plasmid pAdeasy-1 in BJ5183. After confirmation by restriction endonuclease digestion, the recombinant vector was transfected into HEK293 cells, and high-titer recombinant adenovirus (Ad-BMP9, 6) was collected after amplification. Ad-BMP9, 6 was then transduced into C3H10 cells in vitro, and the mRNA expression of BMP9 and BMP6 was detected by RT-PCR. The osteogenic capability of the transfected cells was observed by alkaline phosphatase staining and calcium-alizarin red staining. Results AdBMP9,6 was constructed successfully and effectively infected in C3H10 cells, in which high expressions of BMP6 and BMP9 were detected. C3H10 cells infected with Ad-BMP9,6 showed stronger alkaline phosphatase and calcium-alizarin red staining than the cells trasnfected by either BMP9 or BMP6 alone. Conclusion The recombinant adenovirus co-expressing BMP9 and BMP6 we constructed shows a more potent effect than the adenoviruses expressing either BMP9 or BMP6 alone in inducing the osteogenic differentiation of C3H10 cells into osteoblasts.
