biostudies-arrayexpressJohannes GubatHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-14135The goal of the project is to identify deubiquitinases (DUBs) networks that could be targetable in cancer. To identify functionally interacting DUBs, we performed a pooled combinatorial knockout screen using a CRISPR/Cas12a system. Here we transduced a custom guide library targeting any 2-gene combination derived from 160 genes (including ~90 DUBs and other genes of interest) into Cas12a-expressing HCT116 cells. While the cells are subjected to puromycin selection, a reference sample (Day 4 post-transduction) was taken and sequenced. Two replicates were derived from this pool that were expanded and sequenced at Day 11 and Day 18 post-transduction.biostudies-arrayexpressNucleic Acid Extraction - Genomic DNA was isolated using QIAamp DNA Blood Maxi Kit (Qiagen) as per manufacturer's instructionsSequencing - The amplicon was sequenced on Illumina NovaSeq, reading 66 cycles Read 1 with custom primer cttgtggaaaggacgaaacaccgAAATTTCTCCTCTTGGAGAT; 10 cycles index read i7 to read the RSL, and six cycles index read i5 for the sample barcodeLibrary Construction - An amplicon containing guide and UMI sequences was amplified by PCR, also attaching adapter sequences.Growth Protocol - The cells were grown in a humidified incubator at 37C and 5% CO2 and were regularly passaged at 70-80% confluence. Cell numbers were kept at >100 million cells.Sample Collection - At the specified time point, the cells were detached with 0.25% trypsin and pellettedOrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - NGS data was analyzed with the MaGeCK software, v.0.5.6. Read counts are normalized to medianUnknownTranscriptomicsGenomicsProteomicsIllumina NovaSeq 6000DNA-seqHomo sapiensJohannes GubatPadraig D'ArcyfalseCombinatorial CRISPR loss-of-function screen for deubiquitinases and genes related to the ubiquitin proteasome system in HCT116 cellsThe goal of the project is to identify deubiquitinases (DUBs) networks that could be targetable in cancer. To identify functionally interacting DUBs, we performed a pooled combinatorial knockout screen using a CRISPR/Cas12a system. Here we transduced a custom guide library targeting any 2-gene combination derived from 160 genes (including ~90 DUBs and other genes of interest) into Cas12a-expressing HCT116 cells. While the cells are subjected to puromycin selection, a reference sample (Day 4 post-transduction) was taken and sequenced. Two replicates were derived from this pool that were expanded and sequenced at Day 11 and Day 18 post-transduction.2026-05-31T00:00:00Z2026-05-31T01:01:15.541Z2024-05-31T08:03:21.8ZE-MTAB-14135ERP160781EFO_0002944EFO_0004170EFO_0003789EFO_0002693EFO_0005518EFO_0003816EFO_0004184