Figure 2. In vitro selection results.

The fraction of sequences surviving selection (green) and before selection (black) are shown for CCR5A TALENs (A) and ATM TALENs (B) with EL/KK FokI domains as a function of the number of mutations in both half-sites (left and right half-sites combined excluding the spacer). (C) Specificity scores for the CCR5A TALENs at all positions in the target half-sites plus a single flanking position. The colors range from dark blue (maximum specificity score of 1.0) to white (no specificity, score of 0) to dark red (maximum negative score of −1.0); see the main text for details. Boxed bases represent the intended target base. Note for the right half-site, the R18 TALENs, the sense strand is shown. (D) Same as (C) for the ATM TALENs. For (A), (B), (C) and (D) sample statistics (sample sizes, means, standard deviations, and P-values) are given in Supplementary Table S2 and S3. (E) Enrichment values from the selection of L13+R13 CCR5B TALEN for 16 mutant DNA sequences (mutations in red) relative to on-target DNA (OnB). (F) Correspondence between discrete in vitro TALEN cleavage efficiency (cleaved DNA as a fraction of total DNA) for the sequences listed in (E) normalized to on-target cleavage (= 1) versus their enrichment values in the selection normalized to the on-target enrichment value (= 1). The Pearson’s r coefficient of correlation between normalized cleavage efficiency and normalized enrichment value is 0.90. (G) Discrete assays of on-target and off-target sequences used in (F) as analyzed by PAGE.