Figure 5. CRISPR Cas9-genome editing induces chromosome bridge formation, adding to the genome complexity from micronuclei.

(a) Evidence for genome editing-induced chromosome bridge in pair 5.5 (Extended Data Fig. 3). Scheme as in Fig. 3. CRISPR-Cas9 cut site is indicated by the dashed line and relevant segments of chr5 are indicated by letters A-C. In the first division, the DNA break on sister chromatids results in the formation of a micronucleus with the acentric portions of chr5 (segment C). At the same time, the sister centric fragments (AB) fuse, generating a dicentric bridge concomitantly with the formation of the micronucleus. Asymmetric breakage of the bridge leads to the loss of the “B” segment from the bridge chromosome in the micronucleated daughter. Faded cell inferred to contain two copies of the B segment was not followed further. DNA copy number analysis indicated that in this example the chromosome fragments in the micronucleus underwent DNA replication. This region showed no detectable rearrangements. Note that the acentric fragments of chr5 were not reincorporated into a daughter primary nucleus in the second division. A (purple): p-arm; B (black): centromere to cut site, inferred to reside in the bridge; C (teal): cut site to the telomere. Bottom: Copy number and rearrangement plots of cells from above, as in Fig. 3.

(b) Bridge formation, micronucleation, chromosome fragmentation and chromothripsis from CRISPR-Cas9 genome editing in pair 5.1 (Extended Data Fig. 3). In this sample both homologs were cleaved. The acentric arm of homolog 1 (blue allele) missegregates into a micronucleus in the first generation. The centric fragments of homolog 1 fuse, resulting in a dicentric bridge in the second cell division. After the second cell division, the cell that inherited the acentric fragment of homolog 2 (red) was found to have few SVs or copy number alterations, suggesting it was partitioned into the primary nucleus as indicated in the scheme. By contrast, the acentric segments of homolog 1 were fragmented. Bottom: copy number and rearrangement plots of cells shown above, as in Fig. 3.