in vivo CRISPR screening of ANKL-PDX targeting to iron-requirement molecules

ANKL cell proliferates predominantly in liver sinusoid depending on transferrin. However, little is known about molecular mechanism of iron-dependency of ANKL. To investigate, we performed in vivo CRISPR screening of liver resident ANKL cells targeting to genes encoding 482 iron-requirement molecules. Overall design: ANKL1 cells, an primary ANKL cells harvested from ANKL-patient-derived xenograft mouse model (ANKL-PDX) previously reported, were transduced with Cas9- and small guide RNA (sgRNA) library-expressive plasmids via lentivirus vector, and PDX were established with them. Fourteen days after, ANKL cells were re-harvested from liver of established PDX, followed by genomic DNA extraction and integrated sgRNA amplification. After deep sequencing, dropped-out guide sgRNAs were extracted using MAGeCK program.