Multiplex gene editing by CRISPR-Cpf1 using a single crRNA array

Bernd Zetsche; Matthias Heidenreich; Prarthana Mohanraju; Iana Fedorova; Jeroen Kneppers; Ellen M DeGennaro; Nerges Winblad; Sourav R Choudhury; Omar O Abudayyeh; Jonathan S Gootenberg; Wen Y Wu; David A Scott; Konstantin Severinov; John van der Oost; Feng Zhang

doi:10.1038/nbt.3737

Multiplex gene editing by CRISPR-Cpf1 using a single crRNA array

Nat Biotechnol. 2017 Jan;35(1):31-34. doi: 10.1038/nbt.3737. Epub 2016 Dec 5.

Authors

Bernd Zetsche^#^{1

2

3

4

5}, Matthias Heidenreich^#^{1

2

3

4}, Prarthana Mohanraju^#⁶, Iana Fedorova^{1

2

3

4

7

8}, Jeroen Kneppers^{1

6}, Ellen M DeGennaro^{1

9}, Nerges Winblad^{1

2

3

4}, Sourav R Choudhury^{1

2

3

4}, Omar O Abudayyeh^{1

2

3

4

9}, Jonathan S Gootenberg^{1

2

3

4

10}, Wen Y Wu⁶, David A Scott^{1

2}, Konstantin Severinov^{7

11

12}, John van der Oost⁶, Feng Zhang^{1

2

3

4}

Affiliations

¹ Broad Institute of MIT and Harvard, Cambridge, MA 02142.
² McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139.
³ Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139.
⁴ Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139.
⁵ Department of Developmental Pathology, Institute of Pathology, Bonn Medical School, Sigmund Freud Street 25, 53127 Bonn, Germany.
⁶ Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Stippeneng 4, 6708 WE Wageningen, Netherlands.
⁷ Skolkovo Institute of Science and Technology, Skolkovo, 143025, Russia.
⁸ Peter the Great St.Petersburg Polytechinc University, St. Petersburg, 195251, Russia.
⁹ Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA 02139.
¹⁰ Department of Systems Biology, Harvard Medical School, Boston, MA 02115.
¹¹ Waksman Institute for Microbiology, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA.
¹² Institute of Molecular Genetics, Russian Academy of Sciences, Moscow, 123182, Russia.

^# Contributed equally.

Abstract

Targeting of multiple genomic loci with Cas9 is limited by the need for multiple or large expression constructs. Here we show that the ability of Cpf1 to process its own CRISPR RNA (crRNA) can be used to simplify multiplexed genome editing. Using a single customized CRISPR array, we edit up to four genes in mammalian cells and three in the mouse brain, simultaneously.

MeSH terms

Animals
Bacterial Proteins / genetics*
CRISPR-Associated Proteins / genetics*
CRISPR-Cas Systems / genetics*
Gene Editing / methods*
Mice
RNA, Bacterial / genetics*
Sequence Analysis, RNA / methods*

Substances

Bacterial Proteins
CRISPR-Associated Proteins
RNA, Bacterial

Abstract

MeSH terms

Substances

Grants and funding