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. 2015 Nov 2;112(49):15148–15153. doi: 10.1073/pnas.1518008112

Fig. 3.

Fig. 3.

DRIP reveals the presence of ectopic as well as endogenous HSATII hybrids whose production is affected by RT inhibition. (A) Outline of the experimental layout. Total nucleic acids (TNAs) were isolated from IVT HSATII-transfected 293T cells or SW620 tumor spheres cultured in the presence of ddC or DMSO (ddC−). TNAs were treated with DNase I digestion to remove all potential gDNA contamination. DNA/RNA hybrids were then purified by immunomagnetic pull-down using a hybrid-specific antibody, and their relative quantities were measured by HSATII-chr10 qPCR. Pretreatment of TNA samples with RNase H as indicated demonstrates abrogation of DNA/RNA hybrid detection. (B) Fold change in the enrichment of DNA/RNA hybrids in HSATII-transfected 293T cells measured by qPCR after DRIP. (C) Fold enrichment of endogenous HSATII DNA/RNA hybrids in SW620 tumor spheres analyzed by HSATII-chr10 qPCR after DRIP. Fold changes were calculated based on percent input values, and the RNase H-treated samples were set at 1. For all charts, values represent the average of three independent experiments ± SEM. *P < 0.05 (t test).