Sorting, Cloning, and Colony Propagation for CRISPR/Cas9 Genome Editing Using the CellRaft System

Genome editing, particularly via CRISPR/Cas9 is a powerful method for investigating the role of a given gene in cellular physiology. The method relies on: 1) Reliable introduction (i.e. transfection) of genetic material into cells; 2) Identifying transfection-positive cells; 3) Selecting clones capable of growth and 4) Propagating colonies for analysis of genome edits and downstream functional assays. Here, we present optimized methods using the Cell Microsystems CytoSort Array, a microwell array with releasable single cell isolation features called CellRafts. Using the CytoSort Array, combined with the manual CellRaft System for use with inverted microscopes, we have developed a protocol that represents a more rapid and flexible approach to post-transfection cloning compared to limiting dilution or fluorescence-assisted cell sorting (FACS).