Figure 1

Typical episomal cleavage and repair assay. (A) Map of the pRep4-HO-CAT plasmid showing key features. HO recognition cassette is cloned in just before CAT gene. Arrows indicate location of the primers. (B) Episomal cleavage-and-repair assay at the HO site DSB during a time course of Ad-HO infection. T-HO-CAT cells were infected with adeno-HO (Ad-HO) virus, cells were collected at different time points, and episomes were recovered. PCR was performed by putting primers across the break site (CAT amplicon). AMP (Ampicillin) region on the episome acts as the positive internal control and used for normalization. In addition, a genomic product was also used to ensure equal total DNA recovery from the samples. (C) qPCR analysis for kinetics of cleavage and repair of epsiomes as explained in (B). qPCR values are represented as (CAT/AMP)% over control. (D) Same as in (C) but in presence of the ATM inhibitior-KU55933 (10 μM).