Supplementary Figure 6: ChIRP of circRNA.

(a) PCR gels show that CHIRP of the indicated circRNAs could pull down the promoters of corresponding parent genes (up panel) but not the promoters of the corresponding neighbouring genes (lower panel). pEIF3J (B), the B site shown in Fig. 3d was examined; pPAIP2 (D), the D site shown in Fig. 3d was examined. Promoter sites for CTDSPL2 and MATR3 are shown in the diagram below the gel images. (b) Generally, gene bodies corresponding to parent genes could not be pulled down with CHIRP. One site for each gene body (D’ site for EIF3J; G’ site for PAIP2) showed some enrichment although to lesser degree compared with the enrichment found at the promoter (Fig. 5d). D’ for EIF3J and G’ for PAIP2 are sites in the large introns just upstream of the circRNA sequences. Sites examined in the gene body are shown below the bar figures. Error bars represent the s.e.m. * P value < 0.05, P values were determined with a two-tailed Student’s t-test. (c) ChIP results using the U1A, U1C, U2AF35, or U2AF65 antibodies for the promoter of the neighbouring genes (CTDSPL2 for EIF3J and MATR3 for PAIP2) are shown as a comparison for Fig. 3f. These results indicate that U1 snRNP (U1A & U1C) binds to promoters of genes such as EIF3J and PAIP2 but not to the promoters of CTDSPL2 and MATR3. All data were from triplicate experiments.