Extended Data Fig. 4: O-LIT with 36plex-5K to interrogate homolog organization.
From: 3D mapping and accelerated super-resolution imaging of the human genome using in situ sequencing
a, Minimum distances between heterologous and homologous chromosomes. All measurements represent distances between the geometric centers of chromosomes for which all six targets were imaged. Distances between a chromosome and a heterologous chromosome is the shorter of the two distances between that chromosome and the two homologous copies of the heterologous chromosome (n = 686, 668, 364, 586, 760, and 494 for Chr2, 3, 5, 16, 19, and X, respectively). Inter-homolog distances for Chr16 and 19 are less than those for Chr2, 3, and 5 (independent-samples t-test p = 4.28 ×10-37). Boxes represent the IQR (25th, 50th and 75th percentiles) and whiskers extend 1.5 times the IQR. b, Number of cells with varying numbers of homologs split by K-means clustering. The K-means algorithm was applied to 258 nuclei, individually, to cluster chromosomes into two groups based on proximity and then report the number of homolog pairs that were split by the clustering. A value of “5” indicates that the homologs from each five pairs of imaged autosomes in a single nucleus clustered into two spatially separate groups. Observed, PGP1f cells. Directed random, raw positions in Observed but with the chromosome identities of all positions randomized, with the larger chromosomes (2, 3, 5) biased towards the nuclear periphery and smaller chromosomes (16 and 19) biased towards the nuclear interior. Completely random category, randomization of the chromosome identities carried out with no spatial bias. The significance of each pair was evaluated from a two proportion z-test with n = 258 for each category with a null hypothesis of equal proportion and a significance level of 0.05. c, Density plots of homolog positions. Built by using Kernel density estimation (KDE) of nuclei projected and aligned along the x-y plane of the position of the chromosomes. d, Pie charts of total number of cells with homologs split by a virtual line along the y-axis. e, Number of aligned cells with homologs split by a virtual line parallel to the y-axis at different distances from the origin, that is, number of autosomes with one of their homologs on the left of the line and the other on the right (n = 258 for each category). Boxes represent the IQR (25th, 50th and 75th percentiles) and whiskers extend 1.5 times the IQR.
