Table 3.

Plasmid DNA amounts for co-transfection in epegRNA library selection experiment, related to Step 8

Plasmid	DNA amount per 1 million HAP1 cells
epegRNA library	451 ng
Nicking gRNA (in BPK1520)	180 ng
pCMV-PEmax-P2A-GFP	1579 ng
pEF1a-hMLH1dn	789 ng

We use the Neon electroporation system which routinely yields 15–30% GFP+ cells. If substituting transfection methods, we recommend optimizing to achieve a minimum of 15% GFP+ cells. Others have reported successful transfection of HAP1s with TurboFectin 8.0 (OriGene #TF81005), although we typically observe significantly reduced efficiency compared to electroporation. Anecdotally, Xfect (Takara; Cat #631317) works well for transfecting small plasmids into HAP1s, but we have not tested this transfection reagent with larger plasmids like the pCMV-PEmax-P2A-GFP.