Clustering of integration sites. (A) Clustering in the SCID gene-corrected samples is greater than for a gammaretroviral vector in tissue culture. Clustering was analyzed by comparing the distribution of distances between integration sites (x-axis). That is, the lengths of chromosomal segments between integration sites is measured for all pairs and tabulated. Enrichment for short distances between pairs (left side of x-axis) indicates relatively greater clustering. The probability of encountering distances of the indicated lengths by chance (Prob close sites, y-axis) was normalized for the number of sites in each set. To obtain enough control gammaretroviral integration sites for comparison, sites from various studies were pooled.^19,24,27  The dataset for gammaretroviral vector integration in CD34⁺ cells²⁵  is smaller than the others, so the uncertainty is greater (larger error bars) because of the smaller sample size. The blue horizontal line (random) represents the probability expected for random control sites. The SCID sites were significantly more clustered than those of Moloney murine leukemia virus in HeLa cells. (B) Clustering is greater for frequently isolated SCID-X1 integration sites, reflecting selective expansion of cell clones with integration sites in clusters. The distance between integration sites is shown on the x-axis, and the probability of integration site distance is shown on the y-axis. The population of unique integration sites was annotated for the frequency of sequence reads for each, then the more abundant half (green) was compared with the less abundant half (red). The more abundant sites were significantly more clustered (P ≪ .05).