High throughput single-cell detection of multiplex CRISPR-edited gene modifications

Elisa Ten Hacken; Kendell Clement; Shuqiang Li; María Hernández-Sánchez; Robert Redd; Shu Wang; David Ruff; Michaela Gruber; Kaitlyn Baranowski; Jose Jacob; James Flynn; Keith W Jones; Donna Neuberg; Kenneth J Livak; Luca Pinello; Catherine J Wu

doi:10.1186/s13059-020-02174-1

High throughput single-cell detection of multiplex CRISPR-edited gene modifications

Genome Biol. 2020 Oct 20;21(1):266. doi: 10.1186/s13059-020-02174-1.

Authors

Elisa Ten Hacken^{1

2}, Kendell Clement^{3

4

5}, Shuqiang Li^{3

6}, María Hernández-Sánchez^{1

7}, Robert Redd⁸, Shu Wang⁹, David Ruff⁹, Michaela Gruber^{1

10}, Kaitlyn Baranowski¹, Jose Jacob⁹, James Flynn⁹, Keith W Jones⁹, Donna Neuberg⁸, Kenneth J Livak⁶, Luca Pinello^{11

12

13}, Catherine J Wu^{14

15

16

17}

Affiliations

¹ Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA.
² Harvard Medical School, Boston, MA, USA.
³ Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁴ Molecular Pathology Unit, Center for Cancer Research and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, MA, USA.
⁵ Department of Pathology, Harvard Medical School, Boston, MA, USA.
⁶ Translational Immunogenomics Lab, Dana-Farber Cancer Institute, Boston, MA, USA.
⁷ IBSAL, IBMCC-Cancer Research Center, University of Salamanca, Salamanca, Spain.
⁸ Department of Data Science, Dana-Farber Cancer Institute, Boston, MA, USA.
⁹ Mission Bio, Incorporated, South San Francisco, CA, USA.
¹⁰ Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.
¹¹ Broad Institute of MIT and Harvard, Cambridge, MA, USA. lpinello@mgh.harvard.edu.
¹² Molecular Pathology Unit, Center for Cancer Research and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, MA, USA. lpinello@mgh.harvard.edu.
¹³ Department of Pathology, Harvard Medical School, Boston, MA, USA. lpinello@mgh.harvard.edu.
¹⁴ Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA. cwu@partners.org.
¹⁵ Harvard Medical School, Boston, MA, USA. cwu@partners.org.
¹⁶ Broad Institute of MIT and Harvard, Cambridge, MA, USA. cwu@partners.org.
¹⁷ Department of Medicine, Brigham and Women's Hospital, Boston, MA, USA. cwu@partners.org.

Abstract

CRISPR-Cas9 gene editing has transformed our ability to rapidly interrogate the functional impact of somatic mutations in human cancers. Droplet-based technology enables the analysis of Cas9-introduced gene edits in thousands of single cells. Using this technology, we analyze Ba/F3 cells engineered to express single or multiplexed loss-of-function mutations recurrent in chronic lymphocytic leukemia. Our approach reliably quantifies mutational co-occurrences, zygosity status, and the occurrence of Cas9 edits at single-cell resolution.

Keywords: CRISPR-Cas9; Genetics; Genome editing; Loss-of-function; Mutation; Single cell; chronic lymphocytic leukemia.

Publication types

Evaluation Study
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
CRISPR-Cas Systems*
Female
Gene Editing*
High-Throughput Screening Assays
Humans
Leukemia, Lymphocytic, Chronic, B-Cell / genetics*
Loss of Function Mutation*
Mice
Single-Cell Analysis / methods*

Abstract

Publication types

MeSH terms

Grants and funding