Figure 3. Schematic of the genomic DNA modifications demonstrated by the no-SCAR system.

The donor and target DNA are on the left, the chimeric genomic DNA produced after integration through a fully single-stranded DNA intermediate is in the middle, and the segregated chimeric DNA that results in populations of mutant and wild-type are on the right. (a,b) 21 bp of the targeted region for making point mutations into the rpoB and ack genes. The PAM site is shown in red, mutations are shown in lowercased blue text, and mutation target in lowercased black text. (c) Deletion of 1095 bp of ack using a 73mer oligo. (d) Insertion of 79 bp at the C-terminus of pfkA using dsDNA. Red × indicates the site of Cas9 targeting. (e) Insertion of degeneracies outside of the protospacer sequence.