Construction of rat interleukin-10 adenoviral vector and its expression in bone marrow mesenchymal stem cells / 中国当代儿科杂志
Chinese Journal of Contemporary Pediatrics
;
(12): 708-712, 2019.
Article
in Chinese
| WPRIM
| ID: wpr-775119
ABSTRACT
OBJECTIVE@#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.@*METHODS@#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.@*RESULTS@#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 virus solution containing the rIL-10 gene.@*CONCLUSIONS@#The constructed rIL-10 recombinant adenovirus can mediate the stable expression of rIL-10 gene in bone marrow mesenchymal stem cells, which provides a basis for gene transplantation therapy of inflammatory bowel disease.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Bone Marrow Cells
/
Transfection
/
Adenoviridae
/
Interleukin-10
/
HEK293 Cells
/
Mesenchymal Stem Cells
/
Genetic Vectors
Limits:
Animals
/
Humans
Language:
Chinese
Journal:
Chinese Journal of Contemporary Pediatrics
Year:
2019
Type:
Article
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