Construction of rat interleukin-10 adenoviral vector and its expression in bone marrow mesenchymal stem cells

To construct the recombinant adenoviral vector carrying the rat interleukin-10 (rIL-10) gene, and to investigate whether it is stably expressed in bone marrow mesenchymal stem cells. The rIL-10 gene was amplified by PCR from template rIL-10 cDNA, and the recovered 656 bp rIL-10 DNA fragment was clon...

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Published in:Zhongguo dang dai er ke za zhi 2019-07, Vol.21 (7), p.708-712
Main Authors: Zhang, Wen-Ting, Tang, Juan, Zhao, Hong-Mei, You, Jie-Yu
Format: Article
Language:Chinese
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Summary:To construct the recombinant adenoviral vector carrying the rat interleukin-10 (rIL-10) gene, and to investigate whether it is stably expressed in bone marrow mesenchymal stem cells. The rIL-10 gene was amplified by PCR from template rIL-10 cDNA, and the recovered 656 bp rIL-10 DNA fragment was cloned into pcDNA3.1 to construct pcDNA3.1-IL-10. Then HEK293 cells were transfected with pcDNA3.1-IL-10 and adenoviral vector for homologous recombination, and sequencing and PCR were used to evaluate whether recombination was successful. HEK293 cells were lysed by repeated freeze-thaw cycles, and bone marrow mesenchymal stem cells were infected with the virus solution containing the rIL-10 gene. Western blot was used to measure the expression of rIL-10 in bone marrow mesenchymal stem cells. Sequencing and PCR verified that the rIL-10 adenoviral vector was successfully constructed, with a virus titer of 4×10  PFU/mL. The expression of IL-10 was detected after bone marrow mesenchymal stem cells were infected by the vir
ISSN:1008-8830