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. 2011 Nov;21(11):1822–1832. doi: 10.1101/gr.124644.111

Figure 1.

Figure 1.

Sample replication initiation profiles obtained through massively parallel sequencing of nascent DNA strands. A chromosome map is shown at the top, and the region-of-interest is delineated by a black rectangle. The analyzed region is shown underneath the ideogram, with map coordinates indicated. The experimental tracks (MCF7 or K562 nascent strand [NS]/genome ratios) show the distribution of sequence reads (aligned with the indicated region) obtained from massively parallel sequencing of nascent strands either from MCF7 breast cancer cells or from K562 erythroleukemia cells. All data are shown as the ratio of reads obtained from a nascent strand preparation and reads obtained from a corresponding control genomic DNA preparation. For each track, the y-axis indicates the nascent strand/genomic DNA ratio. Reads were calculated as reads per kilobase per million mapped reads (RPKM); for details, see Supplemental Information. RefSeq genes are aligned under the nascent strand distribution. For RefSeq genes, thick boxes represent exons, whereas thin lines represent introns and untranslated regions. Arrows on RefSeq genes indicate the direction of transcription. Initiation at select sites was verified using real-time PCR, with primers listed in Table 2. Examples of control and nascent strand tracks are shown in Supplemental Figure 1A. (A–C) Mapping replication initiation events at previously characterized replication origins. (A) Data from the MYC locus (human chromosome 8). Replication initiation sites were mapped to the region spanning the promoter to the first exon of the MYC gene. (B) Data from the human beta globin locus (HBB) (human chromosome 11). Replication initiation sites were mapped to the region stretching from the promoter to the first intron of the HBB gene. (C) Data from the HPRT1 gene on the X chromosome. Replication initiation sites were mapped near the HPRT1 promoter. (D) Data from the CTCF locus (chromosome 16 q22.1). This region does not contain a known replication origin. Initiation from gene promoters and from the RLTPR gene region (3′ of the CTCF gene) was verified using real-time PCR on an independent preparation of nascent strands (data not shown).