Figure 1

Ploidy-Seq approach. Outline of the approach for isolating and enriching tumor subpopulation by differences in total DNA content prior to next-generation sequencing. Images and cytometric data are displayed from the triple-negative breast tumor case. (a) Two regions (R1 and R2) were sampled and nuclear suspensions were prepared and stained with DAPI. (b) The nuclei were flow-sorted and different distributions of ploidy were gated. (c) Cells from each ploidy peak were isolated into separate tubes and the DNA was isolated and used to prepare libraries for next-generation sequencing. (d) Somatic mutations from each subpopulation were detected and set theory operations were performed to identify early intermediate or late mutations. (e) The composition of the tumor subpopulations relative to the total mass were calculated from the number of events in the cytometric data.