FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae

Michal A. Swiat, Sofia Dashko, Maxime den Ridder, Melanie Wijsman, John van der Oost, Jean Marc Daran, Pascale Daran-Lapujade

Research output: Contribution to journalArticleAcademicpeer-review

27 Citations (Scopus)

Abstract

Cpf1 is a new class II family of CRISPR-Cas RNA-programmable endonucleases with unique features that make it a very attractive alternative or complement to Cas9 for genome engineering. Using constitutively expressed Cpf1 from Francisella novicida, the present study demonstrates that FnCpf1 can mediate RNA-guided DNA cleavage at targeted genomic loci in the popular model and industrial yeast Saccharomyces cerevisiae. FnCpf1 very efficiently and precisely promoted repair DNA recombination with efficiencies up to 100%. Furthermore, FnCpf1 was shown to introduce point mutations with high fidelity. While editing multiple loci with Cas9 is hampered by the need for multiple or complex expression constructs, processing itself a customized CRISPR array FnCpf1 was able to edit four genes simultaneously in yeast with a 100% efficiency. A remarkable observation was the unexpected, strong preference of FnCpf1 to cleave DNA at target sites harbouring 5'-TTTV-3' PAM sequences, a motif reported to be favoured by Cpf1 homologs of Acidaminococcus and Lachnospiraceae. The present study supplies several experimentally tested guidelines for crRNA design, as well as plasmids for FnCpf1 expression and easy construction of crRNA expression cassettes in S. cerevisiae. FnCpf1 proves to be a powerful addition to S. cerevisiae CRISPR toolbox.
Original languageEnglish
Pages (from-to)12585-12598
JournalNucleic acids research
Volume45
Issue number21
DOIs
Publication statusPublished - 1 Dec 2017

Fingerprint

Clustered Regularly Interspaced Short Palindromic Repeats
Saccharomyces cerevisiae
Acidaminococcus
Francisella
Yeasts
RNA
Recombinational DNA Repair
DNA Cleavage
Endonucleases
Point Mutation
DNA Repair
Plasmids
Observation
Genome
Guidelines
DNA
Genes
Gene Editing

Cite this

Swiat, M. A., Dashko, S., den Ridder, M., Wijsman, M., van der Oost, J., Daran, J. M., & Daran-Lapujade, P. (2017). FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae. Nucleic acids research, 45(21), 12585-12598. https://doi.org/10.1093/nar/gkx1007
Swiat, Michal A. ; Dashko, Sofia ; den Ridder, Maxime ; Wijsman, Melanie ; van der Oost, John ; Daran, Jean Marc ; Daran-Lapujade, Pascale. / FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae. In: Nucleic acids research. 2017 ; Vol. 45, No. 21. pp. 12585-12598.
@article{a0a8b97336c34ab2b0315b7f6edd9db0,
title = "FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae",
abstract = "Cpf1 is a new class II family of CRISPR-Cas RNA-programmable endonucleases with unique features that make it a very attractive alternative or complement to Cas9 for genome engineering. Using constitutively expressed Cpf1 from Francisella novicida, the present study demonstrates that FnCpf1 can mediate RNA-guided DNA cleavage at targeted genomic loci in the popular model and industrial yeast Saccharomyces cerevisiae. FnCpf1 very efficiently and precisely promoted repair DNA recombination with efficiencies up to 100{\%}. Furthermore, FnCpf1 was shown to introduce point mutations with high fidelity. While editing multiple loci with Cas9 is hampered by the need for multiple or complex expression constructs, processing itself a customized CRISPR array FnCpf1 was able to edit four genes simultaneously in yeast with a 100{\%} efficiency. A remarkable observation was the unexpected, strong preference of FnCpf1 to cleave DNA at target sites harbouring 5'-TTTV-3' PAM sequences, a motif reported to be favoured by Cpf1 homologs of Acidaminococcus and Lachnospiraceae. The present study supplies several experimentally tested guidelines for crRNA design, as well as plasmids for FnCpf1 expression and easy construction of crRNA expression cassettes in S. cerevisiae. FnCpf1 proves to be a powerful addition to S. cerevisiae CRISPR toolbox.",
author = "Swiat, {Michal A.} and Sofia Dashko and {den Ridder}, Maxime and Melanie Wijsman and {van der Oost}, John and Daran, {Jean Marc} and Pascale Daran-Lapujade",
year = "2017",
month = "12",
day = "1",
doi = "10.1093/nar/gkx1007",
language = "English",
volume = "45",
pages = "12585--12598",
journal = "Nucleic acids research",
issn = "0305-1048",
publisher = "Oxford University Press",
number = "21",

}

Swiat, MA, Dashko, S, den Ridder, M, Wijsman, M, van der Oost, J, Daran, JM & Daran-Lapujade, P 2017, 'FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae', Nucleic acids research, vol. 45, no. 21, pp. 12585-12598. https://doi.org/10.1093/nar/gkx1007

FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae. / Swiat, Michal A.; Dashko, Sofia; den Ridder, Maxime; Wijsman, Melanie; van der Oost, John; Daran, Jean Marc; Daran-Lapujade, Pascale.

In: Nucleic acids research, Vol. 45, No. 21, 01.12.2017, p. 12585-12598.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae

AU - Swiat, Michal A.

AU - Dashko, Sofia

AU - den Ridder, Maxime

AU - Wijsman, Melanie

AU - van der Oost, John

AU - Daran, Jean Marc

AU - Daran-Lapujade, Pascale

PY - 2017/12/1

Y1 - 2017/12/1

N2 - Cpf1 is a new class II family of CRISPR-Cas RNA-programmable endonucleases with unique features that make it a very attractive alternative or complement to Cas9 for genome engineering. Using constitutively expressed Cpf1 from Francisella novicida, the present study demonstrates that FnCpf1 can mediate RNA-guided DNA cleavage at targeted genomic loci in the popular model and industrial yeast Saccharomyces cerevisiae. FnCpf1 very efficiently and precisely promoted repair DNA recombination with efficiencies up to 100%. Furthermore, FnCpf1 was shown to introduce point mutations with high fidelity. While editing multiple loci with Cas9 is hampered by the need for multiple or complex expression constructs, processing itself a customized CRISPR array FnCpf1 was able to edit four genes simultaneously in yeast with a 100% efficiency. A remarkable observation was the unexpected, strong preference of FnCpf1 to cleave DNA at target sites harbouring 5'-TTTV-3' PAM sequences, a motif reported to be favoured by Cpf1 homologs of Acidaminococcus and Lachnospiraceae. The present study supplies several experimentally tested guidelines for crRNA design, as well as plasmids for FnCpf1 expression and easy construction of crRNA expression cassettes in S. cerevisiae. FnCpf1 proves to be a powerful addition to S. cerevisiae CRISPR toolbox.

AB - Cpf1 is a new class II family of CRISPR-Cas RNA-programmable endonucleases with unique features that make it a very attractive alternative or complement to Cas9 for genome engineering. Using constitutively expressed Cpf1 from Francisella novicida, the present study demonstrates that FnCpf1 can mediate RNA-guided DNA cleavage at targeted genomic loci in the popular model and industrial yeast Saccharomyces cerevisiae. FnCpf1 very efficiently and precisely promoted repair DNA recombination with efficiencies up to 100%. Furthermore, FnCpf1 was shown to introduce point mutations with high fidelity. While editing multiple loci with Cas9 is hampered by the need for multiple or complex expression constructs, processing itself a customized CRISPR array FnCpf1 was able to edit four genes simultaneously in yeast with a 100% efficiency. A remarkable observation was the unexpected, strong preference of FnCpf1 to cleave DNA at target sites harbouring 5'-TTTV-3' PAM sequences, a motif reported to be favoured by Cpf1 homologs of Acidaminococcus and Lachnospiraceae. The present study supplies several experimentally tested guidelines for crRNA design, as well as plasmids for FnCpf1 expression and easy construction of crRNA expression cassettes in S. cerevisiae. FnCpf1 proves to be a powerful addition to S. cerevisiae CRISPR toolbox.

U2 - 10.1093/nar/gkx1007

DO - 10.1093/nar/gkx1007

M3 - Article

VL - 45

SP - 12585

EP - 12598

JO - Nucleic acids research

JF - Nucleic acids research

SN - 0305-1048

IS - 21

ER -