Analysis of retroviral integration sites in the RUNX1 locus. (A) MLV and HIV integration sites and clusters are displayed on the locus as described in the legend of Figure 6A. Translocation breakpoints associated to hematopoietic malignancies are indicated with green bars and were retrieved from the TICdb database of translocation breakpoints in cancer.²⁷  The +23 enhancer (orange bar) was functionally defined in the murine and human genome.³⁰  The H3K4me3 track is that determined by ChIP sequencing in the genome of human CD34⁺/CD133⁺ HPCs.²⁴  P1 and P2 indicate the distal and the proximal RUNX1 promoters. (B) Schematic representation (left) and experimental results (right) of a reverse transcription PCR on RNA samples retrotranscribed from CD34⁺ cells from 3 different donors (dn1-dn3). R (5′-CGACAAACCTGAGGTCATT-3′) and F1 (5′-AGCCTGGCAGTGTCAGAAGT-3′) primers identify the longest RUNX1 isoform (NM_001754) together with its splicing variants, indicated by multiple bands (P1) on the agarose gel. The R and F2 (5′-GAGCTGCTTGCTGAAGATCC-3′) primers specifically amplify the P2 transcript. The RNA of the housekeeping glyceraldehyde-3-phosphate dehydrogenase gene (GAPDH) was amplified as a positive control. (C) Zoom-in of the 2 regions of the RUNX1 locus indicated by dotted squares in panel A. H3K4me3 and Pol II tracks are those determined by ChIP sequencing in the genome of human CD34⁺/CD133⁺ HPCs.²⁴  A CpG island (CpG 31; green box), marking the +23 enhancer (orange box), is also indicated. The UCSC conservation track at the bottom shows multiple alignments and measurements of evolutionary conservation among all placental mammals and between 7 selected vertebrates (rhesus monkey, mouse, dog, chicken, Xenopus tropicalis, Fugu, and zebrafish; green bars).