Fig. 2: SPRINTER improves S-phase identification and enables accurate clone assignment of S-phase cells.

a, The proportion of correctly identified G1/G2- and S-phase cells (y axis) was computed for CCC (blue), MAPD (orange), rtMAPD (MAPD extended with replication timing, red) and SPRINTER (green) across cell cycle phases (x axis) for 100 cell subpopulations (dots), each formed by sampling 500 cells from the diploid (left) or tetraploid (right) ground truth datasets. b, ROC curves (false-positive rates versus true-positive rates) measure the performance in distinguishing G1-phase cells from actively replicating cells using the classification scores computed by existing methods (blue, orange and red) or combining SPRINTER’s S- and G2-phase P values (using the minimum, green) by bootstrapping 300 diploid (top) or tetraploid (bottom) cells for 100 repeats (each curve). c, A binomial process was used to generate cell subpopulation pairs with the same (top) or different (bottom) true underlying fractions of replicating cells (that is, proliferation). The figure is created with BioRender.com. d, The proliferation accuracy was computed for all methods (colors) considering 600 pairs of clones generated as described in c by sampling varying numbers of diploid (left) and tetraploid (right) cells per clone (x axis) with varying S and G2 fractions (20–30% ± 30–50%) for 50 repeats (dots). e, Top, RDRs across 50 kb bins (x axis) for an S-phase cell are affected by replication-induced fluctuations (early- and late-replicating bins in magenta and green, respectively) preventing accurate CNA identification (scattered black lines for expected CNAs). Bottom, instead, SPRINTER’s replication-corrected RDRs are similar to CNA expectations (black lines). f, The absolute error rate (x axis) between true and expected fractions of S-phase cells assigned to a clone was calculated per cell using all methods (colors) in 30 populations of 300 tetraploid cells each, altogether comprising 389 clones. The proportion of clones for which the assigned true S fraction was compatible with the expected S fraction was computed using a binomial test (pie charts). In d and f, box plots show the median and IQR with whiskers denoting values within 1.5 times the IQR from the first and third quartiles. AUC, area under the curve; ROC, receiver operating characteristic; IQR, interquartile range.