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[Construction of rat bowel transforming growth factor beta1 recombinant adenovirus vector]
Ji-peng Li1, Yi Huang, Wei-zhong Wang
1Department of Gastrointestinal Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an 710033, China.
Objective:
To construct a recombinant adenovirus vector which regulates the expression of rat transforming growth factor beta (TGF-beta1).
Methods:
Total RNA was extracted from F344/N rat small intestine pre-treated with Con A to clone TGF-beta1. pTRE-shuttle vector was used as mediator to ligate TGF-beta1 gene and backbone of replication-incompetent adenoviral vector. The constructed recombinant adenovirus contained tetracycline-responsive element which could regulate the expression of inserted genes. After identification, the desired recombinant adenovirus was packaged in HEK 293 cells. Supernatant of high titer adenovirus was collected to detect the TGF-beta1 gene expression by green fluorescent protein(GFP).
Results:
The constructed recombinant adenovirus was identified by restriction endonucleases cutting, sequencing, PCR and GFP examination.
Conclusion:
Rat TGF-beta1 recombinant adenovirus is established successfully, which provides material and evidence for further research of dendritic cell (DC) modified by TGF-beta1 to induce immune tolerance in rat heterotopic small bowel transplantation.
