Related Experiment Video
Updated: May 16, 2026
An Efficient Method for Adenovirus Production
Published on: June 10, 2021
[Construction and package of a recombinant adenoviral vector to inhibit miR-21 expression and its effects on HepG2
Yi-lin Zhang1, Jian-zhong Tang, Hong-peng Liu
1Institute of Laboratory Medicine, General Hospital, Ningxia Medical University, Yinchuan 750004, China. zyl-870331@163.com
Aim:
To construct the recombinant adenoviral vector which can efficiently inhibit mature miR-21 expression and explore its effects and the underlying mechanisms on hepatoma cell line HepG2.
Methods:
Based on miRNA-sponge technology, we synthesized 8 duplicated fragments, fully complementary with the miR-21 sequence and cloned them into a shuttle vector pAdTrack-CMV. The constructed plasmid was sequenced and linearized for homologous recombination with pAdEasy-1 vector in BJ5183 bacteria, then transfected into 293A cells. The recombinant adenovirus was used to challenge HepG2 cells. Mature miR-21 level was detected by real-time PCR. Apoptosis and proliferation of the HepG2 cells were detected by Hochest 33258 staining, Western blotting and MTT assay.
Results:
The restriction enzyme digestion, DNA sequencing and detection of GFP expression demonstrated that recombinant adenoviral vector was constructed successfully. The recombinant adenovirus inhibited the expression of miR-21 in HepG2 cells, and also depressed the proliferation of HepG2 cells and promoted the apoptosis.
Conclusion:
The recombinant adenoviral vector we successfully constructed could efficiently reduce the expression of miR-21 in HepG2 cells and depress the proliferation of HepG2 cells.
