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. 2016 Aug 19;7:12429. doi: 10.1038/ncomms12429

Figure 4. CircANRIL does not regulate 9p21 protein-coding genes and lacks miRNA sponging activity.

Figure 4

(a) Overexpression of circANRIL in HEK-293 cells does not modulate endogenous linANRIL RNA abundance as measured by qPCR (left panel). Expression of p16INK4a and p14ARF, encoded by CDKN2A, and p15INK4b, encoded by CDKN2B, in circANRIL-overexpressing cells (right panel) (pool of three biological replicates, quadruplicate qPCR measurements). (b) Prediction of miRNA-binding sites in circANRIL using miRanda and TargetSpy algorithms. (c) miRNA expression normalized to miR-27b as measured by qPCR (four biological replicates, quadruplicate measurements). (d) RNA immunoprecipitation (RIP) of AGO2. Analysis of precipitated RNAs: CDR1as, MALAT1, positive controls; U1, negative control (RIP was performed in a pool of three biological replicates, quadruplicate qPCR measurements). ***P<0.001. NS, not significantly different. (analysis of variance, and Tukey as post hoc test). Data are given as mean±s.e.m.