[Construction and expression of eukaryotic expression vector containing hVEGF165 gene in rat bone marrow stroma cell].
OBJECTIVE
To construct the eukaryotic expression vector containing human vascular endothelial growth factor 165(VEGF165) gene and express it in rat bone marrow stroma cells(rMSCs).


METHODS
The recombinant plasmid pSP73-VEGF165 was digested with BamH I and Xho I. Then the hVEGF165 gene segment obtained was again cloned into pcDNA3.1 to construct recombinant eukaryotic expression vector pcDNA3.1-VEGF165. Then the recombinant vector was identified by enzyme digestion analysis and sequencing. The rMSCs were transformed by recombinant vector and positive clones were screened with G418. The expression of hVEGF165 gene in the transformed cells was detected by immunocytochemical staining.


RESULTS
Enzyme digestion analysis and sequencing showed that target gene had been cloned into recombinant vector. The expression of hVEGF165 gene in the transformed cells had been demonstrated by immunocytochemical staining.


CONCLUSIONS
The recombinant eukaryotic expression vector has been constructed and expressed successfully in the transformed cells. Therefore, it is possible to use the rMSCs expressing hVEGF165 gene as seed cells in the bone tissue-engineering.
Gao
Quan-wen
QW
Department of Oral Histology and Pathology, College of Stomatology, Fourth Military Medical University, Xi'an 710032, China. gaoqw2000@yahoo.com.cn
Dong
Shao-zhong
SZ
Chen
Xi-zhe
XZ
Chen
Yan
Y
Yang
Lian-jia
LJ
D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017154:Stromal Cells / Q000378:metabolism*; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A