{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Claire Ma"],"organism":["Mus musculus"],"software":["Salmon"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-14519"],"description":["This project explores the effects of Cdc42 gene-knock in effector cytotoxic T lymphocytes (CTLs). Effector CTLs were differentiated from OTI TCR transgenic mice for 5 days in vitro, then nucleofected with CRISPR/Cas9 RNPs targeting Cdc42 or non-targeting sequence."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Sample Collection - Cells were harvested by centrifugation into a cell pellet, and lysed with 350 μL RLT lysis buffer supplemented with 1% v/v beta-mercaptoethanol.","Library Construction - Libraries were prepared for sequencing with the TruSeq Stranded mRNA kit (Illumina).","Sequencing - Paired-end 100 base-pair sequencing was performed using an Illumina NovaSeq 6000, with all 12 libraries sequenced on 1 lane. Sequencing was performed at the Cancer Research UK Cambridge Institute by staff blinded to sample treatments.","Sample Treatment - OT-I CTLs from six mice were nucleofected with CRISPR/Cas9 RNPs on day 5 post-stimulation. 72 h later, RNA was extracted from two million CTLs.","Growth Protocol - Effector OT-I CTLs were differentiated and cultured from murine splenocytes as follows. Naïve OT-I splenocytes were stimulated with 10 nM ovalbumin peptide, SIINFEKL (OVA) for 3 days in complete mouse T cell media: RPMI 1640 medium with 10% FBS, 50 μM beta-mercaptoethanol, 10 U/ml recombinant murine IL-2, 2 mM L-Glutamine, 1 mM sodium pyruvate, 100 U/ml penicillin and 100 μg/ml streptomycin. Cells were pelleted and resuspended in fresh media from 3 days after stimulation, expanded in CTL media and nucleofected on day 5 after stimulation with CRISPR/Cas9 RNPs targeting Cdc42 or non-targeting control. Cells were harvested 72 hours after nucleofection.","Nucleic Acid Extraction - Lysates were passed through a Qiashredder and frozen. RNA was extracted using the RNEasy Mini Kit, according to the manufacturer’s protocol with on-column DNA digestion with RNase-free DNase I. RNA quality was verified by Bioanalyzer 2100 and quantity was determined using a Qubit Fluorometer (version 4)."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Data Transformation - Processed data file is a raw counts table as described in the \\\\\\\\\"high throughput sequence alignment protocol\\\\\\\\\" without further normalization or transformation.","Sequence Alignment - Reads were aligned to the mouse genome (annotation from Ensembl GRCm39 version 103) and transcript expression was quantified using Salmon (v1.9.0) (Patro et al 2017). This was performed by the Cancer Research UK Cambridge Institute bioinformatics core. Counts table from this analysis stage is also provided."],"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["Illumina NovaSeq 6000"],"study_type":["RNA-seq of coding RNA"],"species":["Mus musculus"],"pubmed_authors":["Claire Ma"],"additional_accession":[]},"is_claimable":false,"name":"RNA-seq of Cdc42 knockout murine cytotoxic T lymphocytes","description":"This project explores the effects of Cdc42 gene-knock in effector cytotoxic T lymphocytes (CTLs). Effector CTLs were differentiated from OTI TCR transgenic mice for 5 days in vitro, then nucleofected with CRISPR/Cas9 RNPs targeting Cdc42 or non-targeting sequence.","dates":{"release":"2025-07-29T00:00:00Z","modification":"2025-07-30T00:00:58.848Z","creation":"2024-10-07T20:11:12.943Z"},"accession":"E-MTAB-14519","cross_references":{"ENA":["ERP164850"],"EFO":["EFO_0002944","EFO_0004170","EFO_0003789","EFO_0004917","EFO_0005518","EFO_0003816","EFO_0003738","EFO_0004184","EFO_0003969"]}}