- Carefully design your sgRNA. The sgRNA cleavage site should be close to the mutation site and it is best to be within 15 bp of each other.
- Optimize the transfection efficiency before transfection of gRNA/Cas9 and donor template.
- Use FACS sorter to enrich the transfected cells.
- Use drug or marker to select KI clones if feasible (e.g tag a puromycin resistance gene at C-termini of insertion gene for selection).
- Add HDR enhancer when doing KI experiment.
- For more information, please view GenScript’s webinar on Strategies to Efficiently Generate CRISPR KO/KI Cell Lines.