Figure 4.

Landscape of insertions in primary pro–B cells by TC-Seq. (A) Cartoon diagram comparing RAG1/2^core-induced translocations and insertions. In a translocation (red), RAG1/2^core introduces a single DNA break (red lightning) that recombines with the cleaved I-SceI site at Myc^I (black lightning) on chromosome 15. The resulting translocation contains Myc^I only on one side. In an insertion (blue), RAG1/2^core causes tandem DNA breaks (blue lightning), thereby excising a DNA fragment that subsequently reintegrates into the cleaved I-SceI site. The resulting insertion is flanked by Myc^I on both sides. (B) Origin of insertions by chromosome. Events were normalized per megabase to account for different chromosome sizes. (C) Profile of insertions near the I-SceI site in 5-kb intervals. Dashed lines indicate the ±20-kb region excluded from the analysis for D–F because of saturation. (D) Proportion of insertions from genic regions. (E) Frequency of insertions derived from differentially transcribed genes compared with a random model (dashed line). Asterisks indicate values significantly different from random (P < 0.01, binominal test). (F) Observed number of insertions (o, triangle) originating from ERFSs compared with the random Monte Carlo simulation (s, Tukey boxplot). Asterisks indicate significant enrichment (P < 0.0001, binominal test). For D–F, events from the saturated I-SceI region, cryptic I-SceI sites, and other portions of the genome were excluded (see Materials and methods). Data analysis was performed with pooled RAG2^core and RAG2^−/− TC-Seq libraries (two independent experiments each).