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. 2016 May 26;7(6):434–444. doi: 10.1007/s13238-016-0277-2

Figure 3.

Figure 3

CDK6 protein expression is inhibited by miR-29b via binding to the CDK6 3′-UTR in osteosarcoma cells. (A) Quantitative RT-PCR analyses of the expression levels of miR-29b in osteosarcoma cells after transfection with pre-miR-29b or pre-miR-control. The results were normalized to U6 (***P < 0.001). (B) Western blotting analyses of the expression levels of CDK6 protein in osteosarcoma cells after transfection with pre-miR-29b, pre-miR-control or nothing (mock). Left panel: representative images; right panel: quantitative analysis (***P < 0.001). (C) Quantitative RT-PCR analyses of the expression levels of CDK6 mRNA in the osteosarcoma cells after transfection. The results were normalized to GAPDH. (D and E) Direct recognition and combination of the CDK6 3′-UTR by miR-29b. Firefly luciferase reporters containing either wild-type (WT) or mutant (MUT) miR-29b binding sites in the CDK6 3′-UTR were co-transfected into osteosarcoma cells with either the pre-miR-control or pre-miR-29b. Twenty-four hours post-transfection, the cells were assayed using a luciferase assay kit, and the luciferase activities were normalized to the β-galactosidase levels of the control (***P < 0.001)