Skip to main content
. 2017 Jan 6;13(3):554–566. doi: 10.1080/15548627.2016.1269989

Figure 1.

Figure 1.

MIR7–3HG (MIR-7) targets AMBRA1. (A) Schematics representing sequences of AMBRA1 MRE (top sequence) or its artificially mutated form (bottom sequence) cloned within the 3′-UTR of the reporter Renilla luciferase in the vector psiCHECK-2. Mutations are marked in lower case letters. MIR7–3HG seed sequence is in bold. (B) Normalized luciferase activity in lysates from HEK293 cells, co-transfected with the wild-type or mutant AMBRA1 luciferase constructs and negative control (Empty Vector) or MIR7–3HG plasmids (indicated as MIR7; mean ± SD of 3 independent experiments, *p < 0.05.) N.S., not significant. (C) Western blot analysis of cytoplasmic AMBRA1 levels 24 h after transfection with MIR7–3HG-encoding plasmids (indicated as MIR7) or negative control (Empty Vector). ACTB was used as an endogenous control. One representative western blot of 3 independent experiments is shown. The right panel shows ImageJ densitometry analysis of 3 independent experiments (mean ± SD of independent experiments,**p < 0.01). (D) AMBRA1 mRNA expression was analyzed by quantitative RT-qPCR 24 h after transfection with MIR7–3HG-encoding plasmids (indicated as MIR7) or negative control (Empty Vector). Relative quantification was measured by means of the comparative cycle threshold (ΔΔCt) method (mean ± SD of 3 independent experiments, *p < 0.05).