GSC-like cells have a distinct replication timing program from CySC-like and cultured Drosophila cells. (A) Hierarchical comparison of replication timing indicates that the GSC-like cell replication program is dissimilar to the other three replication programs. On the y-axis, tree branches are shown at the root mean squared Z-score difference of all genomic windows (Euclidean distance), and cell–cell distances at each tree branch are combined using the average method. (B) The mean replication timing Z-score for 11 defined chromosomal regions and the Z-score of the rDNA unit indicate how this summary of the genome hierarchically explains the cell types’ epigenomes. (C) Plot of replication timing for each cell type across the 8 kb transcribed rDNA unit. Early replication timing for this genomic location only occurs in GSC-like cells. (D) Pairwise comparison of replication timing differences for Chromosome 2, in which domains (reaching a size at least 20 kb) are evaluated against the null hypothesis of having equal replication timing. Shown are the location and identification of regions that are replicating earlier in the indicated comparison. (E) The sum total genomic size of pairwise comparisons that are replicating earlier in the specified cell comparison for the left and right arms of Chromosome 2. Statistical significance for total genomic size was calculated using a Fisher's exact test with category aggregation of chromosomes (Supplemental Table S6). The total genomic size was divided by the length of each chromosome arm to calculate the fraction (in percentage) of the arm replicating earlier in that cell type. These values are shown above each bar. (F) Replication timing line plots for each cell type within the pericentromeric region of Chromosome 2 demonstrates the locations where the GSC-like cells replicate early in comparison to the other cell types. A quantification of the total genomic size at which GSC-like cells replicate earlier is presented on the right with the fraction of pericentromeric region that is earlier replicating presented above each bar. (G) The fourth chromosome has more genomic regions that replicate earlier in GSC-like cells compared with the other cell types. Three of these genomic windows are consistently earlier replicating in GSC-like cells compared with the other three cell types and are marked with an arrowhead. Above each bar is the fraction of the fourth chromosome that is earlier replicating in that cell type. (H) Comparison of replication timing between male cell types for the Y Chromosome shows the generally late replication timing of this chromosome. The fraction of the Y Chromosome that is earlier replicating in each cell type is presented above each bar. (I) Compared with female Kc cells, all male cells have a greater total genomic size of earlier replicating DNA on the X Chromosome. These early-replicating domains are located across the entirety of the X Chromosome. The fraction of the X Chromosome that is earlier replicating in each cell type is shown above each corresponding bar.