You will need the following materials:
1. Purified Cas9-NLS protein, 40 µM in 1x Cas9 buffer
2. Purified sgRNA from in vitro transcription, >48 µM or synthetic
3. Single-stranded DNA HDR donor, 100 µM (as an IDT Ultramer) (optional)
4. Lonza 4D Nucleofector with X Unit
5. Lonza kit: electroporation solution and 16 reaction small-sized cuvettes (solution specific to your cell type)
6. 1x Cas9 buffer (20 mM HEPES-KOH pH 7.5, 150 mM KCl, 10% glycerol, 1 mM TCEP) and
2x Cas9 buffer (40 mM HEPES-KOH pH 7.5, 300 mM KCl, 20% glycerol, 2 mM TCEP)