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. Author manuscript; available in PMC: 2017 Dec 5.
Published in final edited form as: Nat Biotechnol. 2017 Jun 5;35(8):789–792. doi: 10.1038/nbt.3900

Figure 1.

Figure 1

A bacterial interference-based negative selection screen identifies amino acid substitutions of AsCpf1 conferring activity at non-canonical PAMs. (a) Crystal structure of AsCpf1 (PDB ID: 5B43) in complex with crRNA and target DNA, highlighting the PAM nucleotides (magenta), and PAM-proximal residues selected for mutagenesis (blue). (b) Schematic of bacterial interference assay used to identify variants with altered PAM specificity. (c) Sensitivity of wild-type AsCpf1 to substitution mutations in the PAM as measured by bacterial interference. Bars show mean ± s.e.m. of n = 3 plated transformations. (d) Scatter plots of screen readout, highlighting depleted variants. Each dot represents a distinct wild-type or mutant codon. The dashed line indicates 15-fold depletion.