[Experimental study of human umbilical cord blood derived stromal cells transfected with recombinant adenoviral vector co-expressing VCAM-1 and GFP].
This study was aimed to investigate the effect of vcam-1 gene-modified human umbilical cord blood derived stromal cells (CBDSCs) on hematopoietic regulation so as to establish the experimental foundation for further study. The target gene vcam-1 was cloned into the shuttle plasmid with the report gene GFP. The recombinant shuttle plasmid was transformed into BJ5183 bacteria to recombine with backbone vector pAdeasy-l, and the recombinant adenoviral vector ad-vcam-1-gfp was confirmed after transfection with CBDSCs. The results indicated that two fragments of about 9 kb and 2 kb were obtained after digestion of recombinant plasmid pAdTrack-vcam-1 with NotIand XhoI, and single fragment of 600 bp was obtained after amplification with PCR; two fragments of about 31 kb and 4 kb were obtained after digestion of recombinant plasmid pad-vcam-1-gfp with PacI, which suggested a successful homologous recombination. The expression of vcam-1 gene in ad-vcam-1-gfp transfected CBDSCs could be detected by immunocytochemistry, RT-PCR and fluorescent microscopy. It is concluded that the recombinant adenoviral vector ad-vcam-1-gfp has been constructed successfully, and the expression of vcam-1 is up-regulated in CBDSCs transfected by gene ad-vcam-1-gfp.
Zhang
Xi
X
Department of Hematology, Xinqiao Hospital, The Third Military Medical University, Chongqing 400037, China.
Si
Ying-Jian
YJ
Chen
Xing-Hua
XH
Liu
Yao
Y
Gao
Li
L
Gao
Lei
L
Peng
Xian-Gui
XG
Wang
Qing-Yu
QY
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism*; D005312:Fetal Blood / Q000166:cytology*; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017154:Stromal Cells / Q000166:cytology*; D014162:Transfection; D019010:Vascular Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism*
D012333:RNA, Messenger; D011994:Recombinant Proteins; D019010:Vascular Cell Adhesion Molecule-1; D049452:Green Fluorescent Proteins