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[Construction and identification of eukaryotic expression plasmid expressing siRNA targeting USP22 gene]
Jian-jun Xiong1, Li-jun Gan, Ling Lin
1Department of Pharmacology, College of Basic Medical Science, Jiujiang University, Jiujiang 332000, China.
Objective:
To construct eukaryotic expression plasmid expressing siRNA targeting ubiquitin specific peptidase 22 gene (USP22), and to investigate its effect on the growth of hepatoma carcinoma cells HepG2.
Methods:
siRNA templates were synthesized based on USP22 mRNA sequence and cloned into vector Pmscv/Hyg/U6. The resulting recombinant was identified by restriction enzyme digestion and DNA sequencing. Recombinants were than transfected into HepG2 cells mediated by liposome. The USP22 protein and mRNA in HepG2 cells were detected by western blot and RT-PCR, respectively. The cellular growth activity was evaluated with MTT assay.
Results:
Recombinant plasmid expressing siRNA targeting USP22 was successfully constructed. The down-regulated protein and mRNA level of USP22 and decreased cellular growth in HepG2 cells transfected with recombinant plasmid were observed.
Conclusion:
The eukaryotic expression vector for RNA interference USP22 gene is constructed successfully, which inhibits the expression of USP22 in HepG2 cells and suppresses cell proliferation.
