Figure 2

CRISPR-Cas9 screening workflow. Duplicate 96 well plates of cell clones are generated. One plate is frozen down or maintained for later use. From the other plate, genomic DNA is extracted, and the targeted region is amplified and uniquely barcoded in each well. The DNA is then pooled, adaptors ligated, and a fragment library is prepared. The sequencing library is then sequenced on the IonTorrent PGM. Reads are filtered and visualized on IGV, to identify the desired clones. The desired clones are then thawed and/or expanded for subsequent experiments.