CRISPR-Cas12a system in fission yeast for multiplex genomic editing and CRISPR interference.
Journal
Nucleic acids research
ISSN: 1362-4962
Titre abrégé: Nucleic Acids Res
Pays: England
ID NLM: 0411011
Informations de publication
Date de publication:
04 06 2020
04 06 2020
Historique:
accepted:
27
04
2020
revised:
19
04
2020
received:
03
09
2019
pubmed:
7
5
2020
medline:
18
8
2020
entrez:
7
5
2020
Statut:
ppublish
Résumé
The CRISPR-Cas12a is a class II, type V clustered regularly interspaced short palindromic repeat (CRISPR) system with both RNase and DNase activity. Compared to the CRISPR-Cas9 system, it recognizes T-rich PAM sequences and has the advantage of multiplex genomic editing. Here, in fission yeast Schizosaccharomyces pombe, we successfully implemented the CRISPR-Cas12a system for versatile genomic editing and manipulation. In addition to the rrk1 promoter, we used new pol II promoters from endogenous coding genes to express crRNA for Cas12a and obtained a much higher editing efficiency. This new design expands the promoter choices for potential applications in fission yeast and other organisms. In addition, we expressed a gRNA array using a strong constitutive pol II promoter. The array transcript is processed by Cas12a itself to release multiple mature crRNAs. With this construct, multiplex genomic editing of up to three loci was achieved from a single yeast transformation. We also built a CRISPR interference system using a DNase-dead Cas12a to significantly repress endogenous gene expression. Our study provides the first CRISPR-Cas12a toolkit for efficient and rapid genomic gene editing and regulation in fission yeast.
Identifiants
pubmed: 32374858
pii: 5831192
doi: 10.1093/nar/gkaa329
pmc: PMC7261154
doi:
Substances chimiques
CRISPR-Associated Proteins
0
RNA
63231-63-0
RNA Polymerase II
EC 2.7.7.-
Deoxyribonucleases
EC 3.1.-
Ribonucleases
EC 3.1.-
Types de publication
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Pagination
5788-5798Informations de copyright
© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.
Références
Nat Commun. 2019 Apr 8;10(1):1598
pubmed: 30962441
Cell. 2015 Oct 22;163(3):759-71
pubmed: 26422227
Nat Rev Mol Cell Biol. 2016 Jan;17(1):5-15
pubmed: 26670017
Nucleic Acids Res. 2017 Dec 1;45(21):12585-12598
pubmed: 29106617
Science. 2007 Mar 23;315(5819):1709-12
pubmed: 17379808
Nat Biotechnol. 2017 Jan;35(1):31-34
pubmed: 27918548
Nucleic Acids Res. 1990 Nov 25;18(22):6686
pubmed: 2251129
Science. 2017 Mar 10;355(6329):1040-1044
pubmed: 28280199
Yeast. 2006 Feb;23(3):173-83
pubmed: 16498704
Genetics. 1994 Mar;136(3):849-56
pubmed: 8005439
Genome Biol. 2016 Jul 05;17(1):148
pubmed: 27380939
G3 (Bethesda). 2018 Mar 2;8(3):789-796
pubmed: 29321167
Nat Plants. 2017 Feb 17;3:17018
pubmed: 28211909
Science. 2001 Apr 6;292(5514):110-3
pubmed: 11283354
Methods Mol Biol. 2004;241:93-111
pubmed: 14970648
Nat Commun. 2014 Oct 29;5:5344
pubmed: 25352017
Wellcome Open Res. 2017 May 5;1:19
pubmed: 28612052
Nat Biotechnol. 2013 Sep;31(9):827-32
pubmed: 23873081
Yeast. 2018 Feb;35(2):201-211
pubmed: 28886218
Nat Methods. 2017 Jun;14(6):615-620
pubmed: 28417998
Science. 2009 Jun 26;324(5935):1716-9
pubmed: 19556509
Nature. 2016 Apr 28;532(7600):517-21
pubmed: 27096362
G3 (Bethesda). 2017 Jan 5;7(1):87-93
pubmed: 27793971
Nat Biotechnol. 2014 Dec;32(12):1262-7
pubmed: 25184501
Synth Syst Biotechnol. 2018 Nov 19;4(1):1-9
pubmed: 30505961
Nature. 2011 Sep 14;477(7365):471-6
pubmed: 21918511
Elife. 2014 Aug 19;3:
pubmed: 25139909
Nucleic Acids Res. 2013 Jan;41(2):775-89
pubmed: 23193295
Cell. 2016 May 5;165(4):949-62
pubmed: 27114038
Cell. 2013 Feb 28;152(5):1173-83
pubmed: 23452860
FEMS Yeast Res. 2018 Jun 1;18(4):
pubmed: 29726937
Nat Commun. 2017 Dec 13;8(1):2095
pubmed: 29235474
Cell. 2013 Jul 18;154(2):442-51
pubmed: 23849981
Genetics. 2016 Jun;203(2):621-9
pubmed: 27270696
Science. 2015 Apr 3;348(6230):1258699
pubmed: 25831549
Nat Biotechnol. 2018 Mar;36(3):239-241
pubmed: 29431740
Science. 2014 Apr 4;344(6179):55-8
pubmed: 24674868
Nat Methods. 2019 Sep;16(9):887-893
pubmed: 31406383
Appl Microbiol Biotechnol. 2012 Aug;95(3):577-91
pubmed: 22652839
Cell Discov. 2017 Jun 06;3:17018
pubmed: 28607761
Science. 2017 Mar 10;355(6329):
pubmed: 28280154
G3 (Bethesda). 2019 Apr 9;9(4):1153-1163
pubmed: 30755408
Biotechnol Adv. 2018 May - Jun;36(3):641-665
pubmed: 29331410
PLoS One. 2014 Jul 07;9(7):e101201
pubmed: 24999979
Science. 2012 Aug 17;337(6096):816-21
pubmed: 22745249
Genetics. 2015 Oct;201(2):403-23
pubmed: 26447128
Science. 2013 Feb 15;339(6121):823-6
pubmed: 23287722
Nat Methods. 2017 Feb;14(2):153-159
pubmed: 27992409
Nucleic Acids Res. 2013 Apr;41(7):4336-43
pubmed: 23460208
Genome Biol. 2016 Mar 08;17:45
pubmed: 26956608
G3 (Bethesda). 2018 Oct 3;8(10):3163-3171
pubmed: 30097473
Cell. 2012 Oct 26;151(3):671-83
pubmed: 23101633
Nat Methods. 2017 Dec;14(12):1163-1166
pubmed: 29083402
Cell. 2014 Jun 5;157(6):1262-78
pubmed: 24906146
Science. 2013 Jun 28;340(6140):1577-80
pubmed: 23812715
Science. 2013 Feb 15;339(6121):819-23
pubmed: 23287718
G3 (Bethesda). 2018 May 31;8(6):2067-2077
pubmed: 29703785
ACS Synth Biol. 2018 Jan 19;7(1):10-15
pubmed: 29161506
ACS Synth Biol. 2017 Jul 21;6(7):1273-1282
pubmed: 28375596
Nat Chem Biol. 2018 Jul;14(7):642-651
pubmed: 29915237
Nat Struct Biol. 2002 Nov;9(11):828-32
pubmed: 12389037