Pooled optical screens in human cells
David Feldman,David Feldman,Avtar Singh,Jonathan L. Schmid-Burgk,Anja Mezger,Anja Mezger,Anja Mezger,Anthony J. Garrity,Rebecca J. Carlson,Rebecca J. Carlson,Feng Zhang,Paul C. Blainey,Paul C. Blainey +12 more
TLDR
This work introduces an optical method to link perturbations and their phenotypic outcomes at the singlecell level in a pooled setting and applies this technology to screen a focused set of 952 genes across >3 million cells for involvement in NF-κB activation.Abstract:
Large-scale genetic screens play a key role in the systematic discovery of genes underlying cellular phenotypes. Pooling of genetic perturbations greatly increases screening throughput, but has so far been limited to screens of enrichments defined by cell fitness and flow cytometry, or to comparatively low-throughput single cell gene expression profiles. Although microscopy is a rich source of spatial and temporal information about mammalian cells, high-content imaging screens have been restricted to much less efficient arrayed formats. Here, we introduce an optical method to link perturbations and their phenotypic outcomes at the singlecell level in a pooled setting. Barcoded perturbations are read out by targeted in situ sequencing following image-based phenotyping. We apply this technology to screen a focused set of 952 genes across >3 million cells for involvement in NF-κB activation by imaging the translocation of RelA (p65) to the nucleus, recovering 20 known pathway components and 3 novel candidate positive regulators of IL-1β and TNFα-stimulated immune responses.read more
Citations
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A Multiplexed Single-Cell CRISPR Screening Platform Enables Systematic Dissection of the Unfolded Protein Response
Britt Adamson,Thomas M. Norman,Marco Jost,Min Y. Cho,James K. Nuñez,Yuwen Chen,Jacqueline E. Villalta,Luke A. Gilbert,Max A. Horlbeck,Marco Y. Hein,Ryan A. Pak,Andrew N. Gray,Carol A. Gross,Oren Parnas,Jonathan S. Weissman,Atray Dixit,Aviv Regev +16 more
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Perturb-Seq: Dissecting Molecular Circuits with Scalable Single-Cell RNA Profiling of Pooled Genetic Screens
Atray Dixit,Atray Dixit,Oren Parnas,Biyu Li,Jenny Chen,Jenny Chen,Charles P. Fulco,Charles P. Fulco,Livnat Jerby-Arnon,Nemanja D. Marjanovic,Nemanja D. Marjanovic,Danielle Dionne,Tyler Burks,Raktima Raychowdhury,Britt Adamson,Thomas M. Norman,Eric S. Lander,Eric S. Lander,Eric S. Lander,Jonathan S. Weissman,Jonathan S. Weissman,Nir Friedman,Nir Friedman,Aviv Regev,Aviv Regev,Aviv Regev +25 more
TL;DR: Perturb-seq accurately identifies individual gene targets, gene signatures, and cell states affected by individual perturbations and their genetic interactions, and posit new functions for regulators of differentiation, the anti-viral response, and mitochondrial function during immune activation.
Journal ArticleDOI
High-throughput mapping of long-range neuronal projection using in situ sequencing
Xiaoyin Chen,Yu Chi Sun,Huiqing Zhan,Justus M. Kebschull,Justus M. Kebschull,Stephan Fischer,Katherine Matho,Z. Josh Huang,Jesse Gillis,Anthony M. Zador +9 more
TL;DR: BARseq is introduced, a multiplexed method based on RNA barcoding for mapping projections of thousands of spatially resolved neurons in a single brain and relating those projections to other properties such as gene or Cre expression that can potentially uncover the organizing principles underlying the structure and formation of neural circuits.
Journal ArticleDOI
Mapping human cell phenotypes to genotypes with single-cell genomics
TL;DR: Recent advances into how single-cell genomics is being used to develop personalized phenotyping strategies that cross subcellular, cellular, and tissue scales to link the authors' genome to their cumulative cellular phenotypes are reviewed.
Journal ArticleDOI
In situ readout of DNA barcodes and single base edits facilitated by in vitro transcription.
Amjad Askary,Luis Óscar Sánchez-Guardado,James M. Linton,Duncan M. Chadly,Mark W. Budde,Long Cai,Carlos Lois,Michael B. Elowitz +7 more
TL;DR: A system for image-based readout of short (20-base-pair) DNA barcodes, called Zombie, where phage RNA polymerases transcribe engineered barcodes in fixed cells and are detected by fluorescent in situ hybridization.
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