Construction of recombinant adenovirus expressing BDNF and its expression in expanded rat mesenchymal stem cells in vitro
- VernacularTitle:BDNF重组腺病毒的构建及在大鼠骨髓间质干细胞的表达
- Author:
Hongle LI
;
Haowei LI
;
Feiyue XING
;
Xuegang SUN
;
Yubin DENG
;
Xiuming ZHANG
;
Yong JANG
;
Shunon LI
- Publication Type:Journal Article
- Keywords:
Bone marrow;
Stem cells;
Brain-derived neurotrophic factor;
Adenovirus;
Green fluorescent protein;
Gene fusion
- From:
Chinese Journal of Pathophysiology
1986;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To construct recombinant adenovirus vector containing brain derived neurotrophic factor, (BDNF) gene using bacterial homogenous recombination, and investigate the expression in expanded rat mesenchymal stem cells (rMSC) in vitro. METHODS: BDNF gene and proBDNF gene were subcloned into adenovirus shuttle plasmid pAdTrack-CMV containing enhanced green fluorescent protein gene (EGFP) expression cassette, forming shuttle vector of pAdTrack-BDNF, and pAdTrack-proBDNF, and co-transformed into BJ5183 bacterial cells with adenovirus backbone vector pAdEasy-1 using chemical transformation. After the recombinant adenovirus vector was obtained, the identified recombinant adenovirus plasmid DNA was digested with Pac I and transfected to 293 cells to package recombinant adenovirus particles. rMSC were infected by recombinant adenovirus and EGFP expression was detected using fluorescent microscope. Infection efficiency was assessed by flow cytometrics. Western blotting identified expression of Ad -proBDNF and Ad-BDNF in rMSC. rMSC infected with Ad -proBDNF and Ad-BDNF were induced to differentiate into neuron-like cells. rMSC infected with Ad -proBDNF and Ad-BDNF were injected into nude mice and assessd in vivo. RESULTS: We successfully constructed the recombinant adenovirus Ad -proBDNF and Ad-BDNF that expressed in expanded rMSC in vitro.CONCLUSION: Recombinant adenovirus high-effectively mediates Ad -proBDNF and Ad-BDNF expression in expanded rMSC in vitro and in vivo.
