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  • Title: [Construction and identification of adenoviral vectors expressing shRNA targeting CDK1 gene].
    Author: Zhao J, Han SX, Ma JL, Huang C, Zhang D.
    Journal: Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi; 2011 Oct; 27(10):1053-5. PubMed ID: 21968300.
    Abstract:
    AIM: To construct three adenoviral vectors harboring shRNA targeting CDK1 gene, and identify its inhibitory effect on the gene expression of CDK1 in hepatoma carcinoma HepG2 cells. METHODS: Three shRNA sequences targeting CDK1 mRNA were designed. The shRNA sequences were annealed and linked with linearized pSIREN-RetroQ-ZsGreen. The recombinants were identified by PCR and DNA sequencing. CDK1-shRNA plasmid was then transfected into the cultured HepG2 cell line with lipofectamine 2000. RT-PCR and Western blot were used to detect the mRNA and protein expression of CDK1, respectively. RESULTS: The small hair-pin RNA sequences were successfully inserted into pSIREN-RetroQ-ZsGreen vector, and the sequences were identified by DNA sequencing. Further, Realtime PCR and Western blot results validated that the three small hair-pin RNAs effectively knockdowned the expression of endogenous CDK1 in HepG2 cells. CONCLUSION: CDK1-shRNA can be effectively transfected into HepG2 cells, and induce post-transcriptional gene silencing of CDK1, which enables further functional study on CDK1.
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