Figure 1.

Donor design strategy

(A) If the site of insertion of a tag (knock-in) and the site of double strand break are not close to each other, disrupting the internal homology between these two sites increases the efficiency of precise repair. Sequence with disrupted homology should be considered as part of the insert and the homology arm should begin after the last silent mutation.

(B) Schematic design for a set of locus-specific homology arms (HA) with PCR primers which also serve as linkers between the tag and the protein of interest. For a given locus, the same set of oligos can be used to generate dsDNA donors to knock-in any tag. Plasmid containing the tag flanked by linkers is used as PCR template. Diagrams are not drawn to scale.