TP53 promotes lineage commitment of human embryonic stem cells through ciliogenesis and sonic hedgehog signaling [CRISPR screen]

CRISPR-Cas9 screen was done to identify genes required for NPC differentiation. Sox1 is an established NPC marker. Human ESCs were infected with GecKO v2 library, selected with puromycin and then allowed to differentiate into Neural progenitor cells (NPC) using StemDiff monolayer protocol (Stem cell Technologies). Day 7 NPCs were collected and FACS sorted to obtain bottom 1% of Sox1 expressing cells. NGS was used to identify guide sequences in cell population and determine the genes required for Sox1 expression during NPC differentiation. Overall design: hESCs were infected with GecKO library v2, selected with puromycin and allowed to differentiate into NPCs by monolayer protocol. Day 7 NPCs were collected and FACS sorted to obtain bottom 1% of Sox1 expressing cells. Thus, genes required for Sox1 expression during NPC differentiation were obtained as screen hits.