Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida

CRISPR/Cas9 systems are nowadays applied extensively to effect genome editing in various organisms including plants. CRISPR from Prevotella and Francisella 1 (Cpf1) is a newly characterized RNA-guided endonuclease that has two distinct features as compared to Cas9. First, Cpf1 utilizes a thymidine-r...

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Main Authors:	Endo, Akira, Masafumi, Mikami, Kaya, Hidetaka, Toki, Seiichi
Format:	Online
Language:	English
Published:	Nature Publishing Group 2016
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5131344/

id	pubmed-5131344
recordtype	oai_dc
spelling	pubmed-51313442016-12-15 Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida Endo, Akira Masafumi, Mikami Kaya, Hidetaka Toki, Seiichi Article CRISPR/Cas9 systems are nowadays applied extensively to effect genome editing in various organisms including plants. CRISPR from Prevotella and Francisella 1 (Cpf1) is a newly characterized RNA-guided endonuclease that has two distinct features as compared to Cas9. First, Cpf1 utilizes a thymidine-rich protospacer adjacent motif (PAM) while Cas9 prefers a guanidine-rich PAM. Cpf1 could be used as a sequence-specific nuclease to target AT-rich regions of a genome that Cas9 had difficulty accessing. Second, Cpf1 generates DNA ends with a 5′ overhang, whereas Cas9 creates blunt DNA ends after cleavage. “Sticky” DNA ends should increase the efficiency of insertion of a desired DNA fragment into the Cpf1-cleaved site using complementary DNA ends. Therefore, Cpf1 could be a potent tool for precise genome engineering. To evaluate whether Cpf1 can be applied to plant genome editing, we selected Cpf1 from Francisella novicida (FnCpf1), which recognizes a shorter PAM (TTN) within known Cpf1 proteins, and applied it to targeted mutagenesis in tobacco and rice. Our results show that targeted mutagenesis had occurred in transgenic plants expressing FnCpf1 with crRNA. Deletions of the targeted region were the most frequently observed mutations. Our results demonstrate that FnCpf1 can be applied successfully to genome engineering in plants. Nature Publishing Group 2016-12-01 /pmc/articles/PMC5131344/ /pubmed/27905529 http://dx.doi.org/10.1038/srep38169 Text en Copyright © 2016, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Endo, Akira Masafumi, Mikami Kaya, Hidetaka Toki, Seiichi
spellingShingle	Endo, Akira Masafumi, Mikami Kaya, Hidetaka Toki, Seiichi Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
author_facet	Endo, Akira Masafumi, Mikami Kaya, Hidetaka Toki, Seiichi
author_sort	Endo, Akira
title	Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
title_short	Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
title_full	Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
title_fullStr	Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
title_full_unstemmed	Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida
title_sort	efficient targeted mutagenesis of rice and tobacco genomes using cpf1 from francisella novicida
description	CRISPR/Cas9 systems are nowadays applied extensively to effect genome editing in various organisms including plants. CRISPR from Prevotella and Francisella 1 (Cpf1) is a newly characterized RNA-guided endonuclease that has two distinct features as compared to Cas9. First, Cpf1 utilizes a thymidine-rich protospacer adjacent motif (PAM) while Cas9 prefers a guanidine-rich PAM. Cpf1 could be used as a sequence-specific nuclease to target AT-rich regions of a genome that Cas9 had difficulty accessing. Second, Cpf1 generates DNA ends with a 5′ overhang, whereas Cas9 creates blunt DNA ends after cleavage. “Sticky” DNA ends should increase the efficiency of insertion of a desired DNA fragment into the Cpf1-cleaved site using complementary DNA ends. Therefore, Cpf1 could be a potent tool for precise genome engineering. To evaluate whether Cpf1 can be applied to plant genome editing, we selected Cpf1 from Francisella novicida (FnCpf1), which recognizes a shorter PAM (TTN) within known Cpf1 proteins, and applied it to targeted mutagenesis in tobacco and rice. Our results show that targeted mutagenesis had occurred in transgenic plants expressing FnCpf1 with crRNA. Deletions of the targeted region were the most frequently observed mutations. Our results demonstrate that FnCpf1 can be applied successfully to genome engineering in plants.
publisher	Nature Publishing Group
publishDate	2016
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5131344/
_version_	1613749796121608192

Efficient targeted mutagenesis of rice and tobacco genomes using Cpf1 from Francisella novicida

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