Figure 1. Maps of new Cas9 and CRISPEY plasmids.
Two-plasmid systems for either galactose induction (A) or estradiol induction (B). The pCRISPEY plasmids contain single restriction enzyme sites for cloning (Gibson assembly), NotI for GAL (A) or XhoI for Z3 (B). gRNAs and their repair templates for targeted editing are synthesized as oligonucleotides and cloned into the vectors. Editing is be performed by induction from 24 – 72 hours following co-transformation of the pCRISPEY and pCAS9-Ec86 plasmids into the strain of interest.