The construction of psiRNA-hHDEK and its impact on CasKi cells biological characteristics / 中国医师杂志

ABSTRACT

Objective The cell proliferation, cell cycle, apoptosis and the impact of senescence after RNAi were performed. This research provided a theoretical and experimental basis of the prevention and treatment of cervical cancer. Methods According to DEK nucleotide sequence in G enebank, 56 nt o1igonucleotide fragment containing specific target DEK sequence was designed with computer software, and the fragnent was synthesized and cloned into the eukaryotic expressed plasmid vector psiRNA-hH ineo.Then it was transfected into CaSki cells by lipofectamine. DEK mRNA and protein expression were detected to verify the gene silence effect by RT-PCR and Western blot analysis. CaSki cell proliferative inhibition rates were accessed by MTT assay at the 48th hour after DEK siRNA transfection, at the same time cell cycle and cell apoptosis were analyzed by flow cytometry, and SA - β-galactosidase enzyme cytochemiatry was used to identify cell senescence. Results DEK mRNA and protein expression was significantly reduced in psiRNA- hHDEK transfected CaSki cells(0. 28 ±0. 02). Cell proliferation was decreased, cell cycle was inhibited, and cell apoptosis and senescent cells were increased afar DEK gene was silenced. Concluslon DEK siRNA eukaryotic expression vector was successfully constructed and DEK gene expression of CaSki cells was effectively silenced. DEK gene silencing could induce CaSki cells into apoptosis and senescence.DEK gene played an important role in the occurrence and evolution of cervical cancer, which was not only a senescence suppressor gene but also an apeptosis suppressor gene.