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Construction of human GDF-5 on adenovirus vector and its expression in MSCs

Cheng, X.; Lin, H.; Xue, J.; Zhang, T.

Sheng Wu Yi Xue Gong Cheng Xue Za Zhi 27(1): 120-125

2010

ISSN/ISBN: 1001-5515
PMID: 20337038
Document Number: 646103
This experimental study was aimed to construct the recombinant adenovirus vector containing human GDF-5 gene, and to use it for infecting human MSCs and detecting the expression of the gene GDF-5. The core sequence of human GDF-5 was amplified by PCR from pCMV-SPORT6, and then was cloned to pAdtrack-CMV. The linearized shuttle plasmid pAdtrack-CMV-GDF-5 was homogenously recombined with pAdeasy-1 in BJ5183. The potential clone was analyzed by restriction endonuclease digestion. The correct clone was linearized and transfected into QBI-293 cells for packing and amplifying so as to obtain adenovirus pAd-GDF-5 and identify it, while the titer was also determined by TCID50. MSCs were infected by the harvested virus, and the expression of GDF-5 was detected by RT-PCR. The recombinant adenovirus vector containing human GDF-5 gene was constructed successfully, its titer was 1 x 10(9) PFU/ml, and it could infect MSCs efficiently. The human MSCs infected by constructed adenovirus vector could continue expressing GDF-5 in a certain time, and the transgenic MSCs would be much potential on tissue regeneration.

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