Figure 2: NOE promotes error-prone DNA repair events that differ between cell types.

(a) NOE stimulates insertions and deletions in HEK293T cells. The allele frequency of deletions (left of plot) and insertions (right of plot) are shown for nucleofections performed with (dark blue, NOE) or without (light blue, RNP) N-oligo. Editing is summarized for each clone in Supplementary Fig. 3. Raw data is available in Supplementary Data 3. (b) Inserted sequences are derived from single- and double-stranded heterologous DNA. Wildtype EMX1 sequence is presented at top with the protospacer (bold), PAM (bold underline), and cut site (triangle) diagrammed. Four example alleles are presented below with sgRNA template (green) and/or non-homologous oligonucleotide (grey) sequence inserted in both orientations. Complete sequencing alignments are available in Supplementary Data 3. (c) NOE stimulates deletions in U2OS cells. Multiple sequence reads from edited cell populations are presented as described in Fig. 2a. (d) Insertion of non-homologous DNA primarily occurs in HEK293T and K562 cells. DNA harvested from unedited (-), RNP treated (R), or NOE treated (N) cell populations was evaluated using a panel of PCR reactions (diagrammed at top). Primers N1 and N2 anneal to the N-oligo sequence; primers T1 and T2 anneal to residual sgRNA template. Open arrow—position of the 700 bp ladder band.