3 RAG1/2 introduces double-stranded breaks at non-Ig loci | 61 which is not expected to be a RAG1/2 recombinase hotspot. Substantially more binding to Jk1 and Jk2 regions was observed in Artemis-/- cells (AH2) as compared to Artemis-/-Rag2-/- cells (R2K.Art). The binding of NBS1 to the Plac8 control region was low and there was no notable difference between Artemis-/- (AH2) and Artemis-/-Rag2-/ (R2K.Art) (Figure 1C). To identify RAG off-targets on a genome-wide scale, we performed NBS1 ChIP followed by subsequent massive parallel sequencing of all immunoprecipitated material. The NBS1 ChIP-Seq reads were robustly accumulated at the Igk region on chromosome 6 (Figure 2A) in RAG1/2 proficient cells (AH2), whereas no accumulation of NBS1 reads is observed in RAG2-deficient cell line (R2K.Art). Input samples show only a background level of NBS1 reads at this region.
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