Construction of adenoviral vectors expressing miR-30a and miR-30e / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To construct adenoviral vectors expressing mature miRNA-30a and miRNA-30e.</p><p><b>METHODS</b>The target mmu-miR-30a and mmu-miR-30e genes amplified from mouse genome were digested and linked to the shuttle plasmid pSES-HUS, which was then transformed into competent AdEaseier cells for recombination. The confirmed recombinant plasmids were transfected into Hek-293 cells for production of the adenoviruses pAd-mmu-miR-30a and pAd-mmu-miR-30e. The obtained adenoviruses were used to infect Mefs cells, and the cellular expressions of mmu-miR-30a and mmu-miR-30e were detected using fluorescence quantitative PCR.</p><p><b>RESULTS</b>mmu-miR-30a (357 bp) and mmu-miR-30e (324 bp) containing the restriction sites were amplified and linked to the shuttle plasmid pSES-HUS, which was successfully recombined with AdEasy1. After packaging in Hek-293 cells, the adenoviral vectors were obtained, which caused an increase of mmu-miR-30a expression by 26.46∓7.46 folds and mmu-miR-30e expression by 2.76∓0.25 folds in transfected Mefs cells.</p><p><b>CONCLUSION</b>We have successfully constructed the adenoviral vectors expressing the mature miRNAs.</p>