biostudies-arrayexpressHomo sapiensCell Ranger v8.8.0https://www.ebi.ac.uk/biostudies/studies/E-MTAB-14567Using the Direct Capture Perturb-seq technology (Replogle, J.M. et al. Nat Biotechnol 38, 954–961 (2020)), we created a pooled of guides that target sites of promoter usage within 50 gene candidates on a stable MCF-7 KRAB dCas9 cell line. Single-cell sequencing 10x5' with CRISPR library and transcriptome. Sequenced on a Nova-Seq.biostudies-arrayexpressSample Collection - Gene expression (GEX) and sgRNA libraries were prepared following 10X Genomics Chromium Single Cell 5' Kit User Guide v2 (Dual Index) with Feature Barcode technology for CRISPR Screening (CG000510 Rev B).Nucleic Acid Extraction - After 6 days, the cells were harvested for total RNA extraction using the RNeasy Mini Plus Kit (74134 - Qiagen) according to the manufacturer’s instructions. cDNA was synthesized using RevertAid RT Reverse Transcription kit (K1691 - Thermo Fisher Scientific) following the manufacturer's protocol.Growth Protocol - Media was replaced 24 hours after transfection, and 48 hours later, supernatants containing viruses were collected by centrifugation and filtration through a 0.45-μm PVDF filter (Millex Millipore - 0890). The viruses were either used immediately or stored in aliquots at −80 °C.Library Construction - By the 8th day post-transduction, more than 70% of the MCF-7 KRAB dCas9 population expressed the library (BFP+), and a final cell sorting (Aria II Cell Sorter - BD Biosciences) was conducted prior single-cell direct capture Perturb-Seq. PromCRISPRi cells with more than 90% viability were prepared as single-cell suspension and loaded into droplet emulsions to two lanes of Chromium Single Cell Chip K, aiming to recover ~20,000 cells per GEM group = 40,000 in total.Sequencing - Sequencing was performed on a NovaSeq 6000 (Illumina) according to the 10x Genomics User Guide.MINSEQE ScoreAssays and DataProcessed DataorganisationMAGE-TAB FilesSequence Alignment - Using Cell Ranger v8.8.0, the transcriptomic library was aligned to reference GRCh38 v77 using “count”, and the feature library was aligned to the supplied list of guides.Data Transformation - Count matrices were filtered for genes with a minimum of 20 cells and genes with a minimum of 1000 reads—highly variable genes assigned to top 2000 with Seurat_v3 then normalized with log-scaled transformation.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsNAIllumina NovaSeq 6000RNA-seq of coding RNA from single cellsHomo sapiensERP165596Helaine Graziele VieiraHelen KingRobert WeatherittfalsePerturb-Seq scRNA-seq for Alternative Promote Usage SitesUsing the Direct Capture Perturb-seq technology (Replogle, J.M. et al. Nat Biotechnol 38, 954–961 (2020)), we created a pooled of guides that target sites of promoter usage within 50 gene candidates on a stable MCF-7 KRAB dCas9 cell line. Single-cell sequencing 10x5' with CRISPR library and transcriptome. Sequenced on a Nova-Seq.2025-11-19T00:00:00Z2026-05-27T13:36:38.221Z2024-10-28T13:52:15.315ZE-MTAB-14567ERP165596EFO_0002944EFO_0004170EFO_0003789EFO_0005684EFO_0004917EFO_0005518EFO_0003816EFO_0004184