Crystal violet staining

Cells infected with lentiviruses were plated in equal number in 12 well plates (BD falcon) with expression of Cas9 stabilized using 200 nM of Shield-1 and incubated at 37 °C for indicated times as described. Cells were rinsed twice with PBS to eliminate the floating cells and fixed in 4% Formaldehyde in PBS (V/V) for 10–15 min. Fixed cells were stained by staining solution (0.1% Crystal violet in 10% ethanol) for 20 min. Staining solution was aspirated from the wells and cells were washed with water thrice in order to remove extra stain. Stained cells were air dried and imaged using Odyssey Imaging System (LICOR). To quantify the relative cell numbers, cells were destained with 10% Acetic acid and absorbance of de-stained solution was measured at 590 nm at appropriate dilutions.