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Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq

View ORCID ProfileHans-Hermann Wessels, Alejandro Méndez-Mancilla, Efthymia Papalexi, William M Mauck, Lu Lu, John A. Morris, Eleni Mimitou, Peter Smibert, Neville E. Sanjana, Rahul Satija
doi: https://doi.org/10.1101/2022.02.02.478894
Hans-Hermann Wessels
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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  • ORCID record for Hans-Hermann Wessels
Alejandro Méndez-Mancilla
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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Efthymia Papalexi
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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William M Mauck
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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Lu Lu
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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John A. Morris
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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Eleni Mimitou
1New York Genome Center, New York, NY, USA
3Technology Innovation Lab, New York Genome Center, New York, NY, USA
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Peter Smibert
1New York Genome Center, New York, NY, USA
3Technology Innovation Lab, New York Genome Center, New York, NY, USA
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Neville E. Sanjana
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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  • For correspondence: rsatija@nygenome.org
Rahul Satija
1New York Genome Center, New York, NY, USA
2Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY, USA
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  • For correspondence: rsatija@nygenome.org
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Abstract

Pooled CRISPR screens coupled with single-cell RNA-sequencing have enabled systematic interrogation of gene function and regulatory networks. Here, we introduce Cas13 RNA Perturb-seq (CaRPool-seq) which leverages the RNA-targeting CRISPR/Cas13d system and enables efficient combinatorial perturbations alongside multimodal single-cell profiling. CaRPool-seq encodes multiple perturbations on a cleavable array which is associated with a detectable barcode sequence, allowing for the simultaneous targeting of multiple genes. We compared CaRPool-seq to existing Cas9-based methods, highlighting its unique strength to efficiently profile combinatorially perturbed cells. Finally, we apply CaRPool-seq to perform multiplexed combinatorial perturbations of myeloid differentiation regulators in an acute myeloid leukemia (AML) model system and identify extensive interactions between different chromatin regulators that can enhance or suppress AML differentiation phenotypes.

Competing Interest Statement

In the past three years, RS has worked as a consultant for Bristol-Myers Squibb, Regeneron, and Kallyope, and served as a SAB member for ImmunAI, Apollo Life Sciences GmbH, Nanostring, and the NYC Pandemic Response Lab. NES is an advisor to Vertex. PS is a co-inventor on a patent related to protein detection by sequencing as described in this work. The New York Genome Center and New York University have applied for patents relating to the work in this article.

Footnotes

  • ↵† E-mail: neville{at}sanjanalab.org

  • ↵‡ E-mail: rsatija{at}nygenome.org

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted February 02, 2022.
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Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq
Hans-Hermann Wessels, Alejandro Méndez-Mancilla, Efthymia Papalexi, William M Mauck, Lu Lu, John A. Morris, Eleni Mimitou, Peter Smibert, Neville E. Sanjana, Rahul Satija
bioRxiv 2022.02.02.478894; doi: https://doi.org/10.1101/2022.02.02.478894
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Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq
Hans-Hermann Wessels, Alejandro Méndez-Mancilla, Efthymia Papalexi, William M Mauck, Lu Lu, John A. Morris, Eleni Mimitou, Peter Smibert, Neville E. Sanjana, Rahul Satija
bioRxiv 2022.02.02.478894; doi: https://doi.org/10.1101/2022.02.02.478894

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