Construction of vascular endothelial growth factor modified muscle-derived stem cells and its expression / 实用医学杂志
The Journal of Practical Medicine
;
(24): 2954-2956, 2015.
Article
in Chinese
| WPRIM
| ID: wpr-481119
ABSTRACT
Objective To construct the lentiviral vector encoding vascular endothelial growth factor gene and detect the vascular endothelial growth factor (VEGF) expression in muscle-derived stem cells (MDSCs). Methods To culture MDSCs and detect the CD34,CD45,Bcl-2 and Desmin expression in MDSCs by immunofluorescence. A cDNA encoding VEGF gene was amplified by PCR. This fragment was cut with EcoRI and BamHI and ligated with an EcoRI- and BamHI-reated lentiviral vector pCDH-CMV-MCS-EF1-copGFP. Then DNA sequencing analysis was performed to confirm successful construction of pCDH-CMV-MCS-EF1-copGFP -VEGF. The expression of VEGF was confirmed using enzyme-linked immunosorbent assay (ELISA), Western blot, and Real-time PCR analyses. Results The pCDH-CMV-MCS-EF1-copGFP-VEGF lentiviral vector was constructed successfully. When MOI values in the transfection efficiency MDSCs by FCM. were 1,5,15, the transfection rate reached to 16.7%, 45.6%, 66.3% and 85.6% respectively. When MOI value was of 20, the rate was up to 90.1%. Real-time PCR, Western blot and ELISA showed stable expression of VEGF in MDSCs. Conclusion We successfully constructed lentiviral vector carrying the VEGF and stable expression in MDSCs.
Full text:
Available
Index:
WPRIM (Western Pacific)
Language:
Chinese
Journal:
The Journal of Practical Medicine
Year:
2015
Type:
Article
Similar
MEDLINE
...
LILACS
LIS