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idr0071-study.txt
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# STUDY DESCRIPTION SECTION
# Section with generic information about the study including title, description, publication details (if applicable) and contact details
Comment[IDR Study Accession] idr0071
Study Title Optical pooled screens in human cells
Study Type in situ sequencing
Study Type Term Source REF EFO
Study Type Term Accession EFO_0008989
Study Description Genetic screens are critical for the systematic identification of genes underlying cellular phenotypes. Pooling gene perturbations greatly improves scalability, but is not compatible with imaging of complex and dynamic cellular phenotypes. Here, we introduce a pooled approach for optical genetic screens in mammalian cells. We use targeted in situ sequencing to demultiplex a library of genetic perturbations following image-based phenotyping. We screened a set of 952 genes across millions of cells for involvement in NF-κB signaling by imaging the translocation of RelA (p65) to the nucleus. Screening at a single time point across 3 cell lines recovered 15 known pathway components, while repeating the screen with live-cell imaging revealed a novel role for Mediator complex subunits in regulating the duration of p65 nuclear retention. These results establish a new approach to perform highly multiplexed, image-based screens of spatially and temporally defined phenotypes with pooled libraries.
Study Key Words CRISPR functional genomics high-content screening in situ sequencing optical pooled screen pooled screen
Study Organism Homo sapiens
Study Organism Term Source REF NCBITaxon
Study Organism Term Accession 9606
Study Experiments Number 6
Study External URL
Study BioImage Archive Accession
Study Public Release Date 2021-01-27
# Study Publication
Study PubMed ID 31626775
Study Publication Title Optical pooled screens in human cells
Study Author List Feldman D, Singh A, Schmid-Burgk JL, Carlson RJ, Mezger A, Garrity AJ, Zhang F, Blainey P
Study PMC ID PMC6886477
Study DOI 10.1016/j.cell.2019.09.016
# Study Contacts
Study Person Last Name Blainey
Study Person First Name Paul
Study Person Email [email protected]
Study Person Address Blainey Lab, Broad Institute, Cambridge, Massachusetts
Study Person ORCID https://orcid.org/0000-0002-4889-8783
Study Person Roles submitter
# Study License and Data DOI
Study License CC BY 4.0
Study License URL https://creativecommons.org/licenses/by/4.0/
Study Copyright Feldman et al
Study Data Publisher University of Dundee
Study Data DOI
Term Source Name NCBITaxon EFO CMPO FBbi
Term Source URI http://purl.obolibrary.org/obo/ http://www.ebi.ac.uk/efo/ http://www.ebi.ac.uk/cmpo/ http://purl.obolibrary.org/obo/
# EXPERIMENT SECTION
# Experiment Section containing all information relative to each experiment in the study including materials used, protocols names and description, phenotype names and description. For multiple experiments this section should be repeated. Copy and paste the whole section below and fill out for the next experiment"
Experiment Number 1
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentA
Experiment Sample Type cell
Experiment Description Optical pooled CRISPR knockout screen in HeLa cells for regulators of p65 translocation upon stimulation by IL-1b or TNF-a, detected by imaging a p65-mNeonGreen fluorescent reporter. CRISPR sgRNAs are detected by sequencing proxy barcodes in cells transduced with the lentiGuide-BC vector. Included cycles of sequencing-by-synthesis (SBS): 1, 2, 3, 4, 5, 6, 7, 8, 9.
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File
Experiment Number 2
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentB
Experiment Sample Type cell
Experiment Description Optical pooled CRISPR knockout screen in HeLa cells for regulators of p65 translocation upon stimulation by IL-1b or TNF-a, detected by indirect immunofluorescence of endogenous p65. CRISPR sgRNAs are detected by sequencing from the 5' end of the sgRNA sequence in cells transduced with the CROPseq-Puro vector. Included cycles of sequencing by synthesis (SBS): 1, 2, 3, 5, 6, 7, 8, 9, 10
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File
Experiment Number 3
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentC
Experiment Sample Type cell
Experiment Description Optical pooled CRISPR knockout screen in A549 cells for regulators of p65 translocation upon stimulation by IL-1b or TNF-a, detected by indirect immunofluorescence of endogenous p65. CRISPR sgRNAs are detected by sequencing from the 5' end of the sgRNA sequence in cells transduced with the CROPseq-Puro vector. Included cycles of sequencing-by-synthesis (SBS): 1, 2, 3, 4, 5, 7, 8, 9, 10.
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File
Experiment Number 4
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentD
Experiment Sample Type cell
Experiment Description Optical pooled CRISPR knockout screen in HCT-116 cells for regulators of p65 translocation upon stimulation by IL-1b or TNF-a, detected by indirect immunofluorescence of endogenous p65. CRISPR sgRNAs are detected by sequencing from the 5' end of the sgRNA sequence in cells transduced with the CROPseq-Puro vector. Included cycles of sequencing-by-synthesis (SBS): 1, 2, 3, 4, 5, 6, 7, 8, 9.
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File
Experiment Number 5
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentE
Experiment Sample Type cell
Experiment Description Twelve cycles of in situ sequencing in HeLa cells expressing barcodes from the lentiGuide-BC lentiviral vector. Included cycles of sequencing-by-synthesis (SBS): 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12.
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File
Experiment Number 6
Comment[IDR Experiment Name] idr0071-feldman-crisprko/experimentF
Experiment Sample Type cell
Experiment Description Combinatorial detection of multiple CRISPR sgRNAs per cell in HeLa cells serially transduced with the CROPseq-Puro and CROPseq-Zeo vectors. Included cycles of sequencing-by-synthesis (SBS): 1, 2, 3, 4, 5, 6.
Experiment Size 5D Images: Average Image Dimension (XYZCT): Total Tb:
Experiment Example Images
Experiment Imaging Method fluorescence microscopy
Experiment Imaging Method Term Source REF Fbbi
Experiment Imaging Method Term Accession Fbbi_00000246
Experiment Comments widefield fluorescence microscopy
# assay files
Experiment Assay File
Experiment Assay File Format tab-delimited text
Assay Experimental Conditions
Assay Experimental Conditions Term Source REF
Assay Experimental Conditions Term Accession
Quality Control Description
# Protocols
Protocol Name growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type growth protocol treatment protocol image acquisition and feature extraction protocol data analysis protocol
Protocol Type Term Source REF EFO EFO
Protocol Type Term Accession EFO_0003789 EFO_0003969
Protocol Description
# Phenotypes
Phenotype Name
Phenotype Description
Phenotype Score Type
Phenotype Term Source REF CMPO
Phenotype Term Name
Phenotype Term Accession
# Feature Level Data Files (give individual file details unless there is one file per well)
Feature Level Data File Name
Feature Level Data File Format
Feature Level Data File Description
Feature Level Data Column Name
Feature Level Data Column Description
Feature Level Data Column Description
# Processed Data Files
Processed Data File Name
Processed Data File Format tab-delimited text
Processed Data File Description
Processed Data Column Name
Processed Data Column Type
Processed Data Column Annotation Level
Processed Data Column Description
Processed Data Column Link To Assay File