Efficient genome editing in Fusarium oxysporum based on CRISPR/Cas9 ribonucleoprotein complexes

Fungal Genetics and Biology : FG & B
Qiang WangJeffrey J Coleman

Abstract

The Fusarium oxysporum species complex (FOSC) is an economically important group of pathogenic filamentous fungi that are able to infect both animals and plants. Reverse genetic techniques, including gene disruption/deletion methods, to study these fungi are available although limitations exist resulting in decreased efficiency. Herein we describe a gene editing system developed using a F. oxysporum-optimized Cas9 ribonucleoprotein (RNP) and protoplast transformation method. The Cas9 protein and sgRNA were assembled to form a stable RNP in vitro and this complex was transferred into fungal protoplasts for gene editing with PEG-mediated transformation. In order to determine if the Cas9 RNP system is functional in the FOSC protoplasts and assess the efficacy of the system, two genes, URA5 and URA3, were selected for targeted disruption generating uracil auxotroph mutants that are resistant to 5-fluoroorotic acid, 5-FOA. In addition, a gene in a secondary metabolite biosynthetic cluster, the ortholog of BIK1, was mutated using this system and the maximum efficiency of this gene disruption was about 50%. Further analysis of the bik1 mutant confirmed that this polyketide synthase was involved in the synthesis of the red pigment, bik...Continue Reading

References

Feb 1, 1989·Molecular & General Genetics : MGG·J de MontignyF Lacroute
Jun 1, 1997·Fungal Genetics and Biology : FG & B·A Aleksenko, A J Clutterbuck
Feb 20, 1999·Protein Expression and Purification·P SheffieldZ Derewenda
Apr 20, 2001·The Journal of Cell Biology·N MosammaparastL F Pemberton
Aug 24, 2006·JAMA : the Journal of the American Medical Association·Douglas C ChangUNKNOWN Fusarium Keratitis Investigation Team
Oct 16, 2007·Clinical Microbiology Reviews·Marcio Nucci, Elias Anaissie
Apr 30, 2009·Molecular Microbiology·Philipp WiemannBettina Tudzynski
Apr 30, 2009·Molecular Plant Pathology·Caroline B Michielse, Martijn Rep
Jul 1, 2009·BMC Bioinformatics·Alex N Nguyen BaAlan M Moses
Oct 13, 2011·Molecular Systems Biology·Fabian SieversDesmond G Higgins
Nov 10, 2011·Molecular Plant-microbe Interactions : MPMI·Jeffrey J ColemanHans D VanEtten
Mar 2, 2012·International Journal of Food Microbiology·Frederik T HansenRasmus J N Frandsen
Mar 6, 2013·Nucleic Acids Research·James E DiCarloGeorge M Church
Jul 6, 2013·Cell Reports·Andrew R BassettJi-Long Liu
Oct 23, 2013·Medicine·Maged MuhammedEleftherios Mylonakis
Oct 26, 2013·Nature Protocols·F Ann RanFeng Zhang
Dec 18, 2013·Science·Ophir ShalemFeng Zhang
Sep 30, 2014·Cell·Randall J PlattFeng Zhang
Oct 30, 2014·Nature Communications·Jake Z JacobsMikel Zaratiegui
Nov 29, 2014·Science·Jennifer A Doudna, Emmanuelle Charpentier
Jul 16, 2015·PloS One·Christina S NødvigUffe H Mortensen
Oct 20, 2015·Nature Biotechnology·Je Wook WooJin-Soo Kim
Apr 14, 2016·ACS Synthetic Biology·Carsten PohlYvonne Nygård
Sep 25, 2016·Applied and Environmental Microbiology·Véronique Edel-HermannChristian Steinberg
Nov 12, 2016·Trends in Genetics : TIG·Mingyu LiWenbiao Chen
Jan 10, 2017·Frontiers in Plant Science·Mickael MalnoyChidananda Nagamangala Kanchiswamy
Mar 14, 2017·Fungal Genetics and Biology : FG & B·Maximilian WenderothReinhard Fischer
Jul 15, 2015·Fungal biology and biotechnology·Toru Matsu-UraChristian Hong

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Citations

Mar 10, 2019·The New Phytologist·Jérôme CollemareMarc-Henri Lebrun
Aug 21, 2019·Microorganisms·Rahul Mahadev ShelakeJae-Yean Kim
May 28, 2019·Frontiers in Plant Science·Muntazir MushtaqRomesh Kumar Salgotra
Jun 23, 2019·Applied Microbiology and Biotechnology·Yan-Zhen MeiChuan-Chao Dai
Aug 15, 2020·Frontiers in Plant Science·Erika N DortRichard C Hamelin
Jul 25, 2019·Applied Microbiology and Biotechnology·Runjie SongHao Lu
Jan 11, 2019·BMC Biotechnology·Zhenzhen Hao, Xiaoyun Su
Apr 8, 2020·Fungal biology and biotechnology·Haseena KhanPeter S Solomon
Dec 29, 2019·Scientific Reports·Massimo FerraraGiuseppina Mulè
Mar 5, 2019·Frontiers in Plant Science·Isabel Vicente MuñozGiovanni Vannacci
Apr 16, 2020·Frontiers in Bioengineering and Biotechnology·Yu-Ke CenYu-Guo Zheng
Jan 8, 2020·Critical Reviews in Biotechnology·Huaxiang DengYujie Cai
Oct 21, 2020·GM Crops & Food·Swati TyagiPratyoosh Shukla
Oct 24, 2020·Current Genomics·Seenichamy Rathinam PrabhukarthikeyanSamantaray Sanghamitra
Sep 24, 2019·Journal of Biotechnology·Gabriela Pannunzio Carmignotto, Adriano Rodrigues Azzoni
Jan 21, 2020·Fungal Genetics and Biology : FG & B·Andi M WilsonBrenda D Wingfield
Apr 22, 2019·Current Opinion in Biotechnology·Casey A HookerKevin V Solomon
Feb 10, 2019·Fungal Genetics and Biology : FG & B·Qiang Wang, Jeffrey J Coleman
Jul 18, 2019·Computational and Structural Biotechnology Journal·Qiang Wang, Jeffrey J Coleman
Apr 2, 2021·Frontiers in Bioengineering and Biotechnology·Rosa SagitaKristina Haslinger
Apr 4, 2021·Journal of Fungi·Sanaz NargesiMohammad Taghi Hedayati
Apr 9, 2021·Biochemistry. Biokhimii︠a︡·Aleksandra M Rozhkova, Valeriy Yu Kislitsin
May 25, 2021·Fungal Genetics and Biology : FG & B·Parthasarathy SanthanamRichard R Bélanger
Jun 14, 2021·Fungal Genetics and Biology : FG & B·Chenggang Wang, Jeffrey A Rollins
Nov 28, 2019·ACS Applied Materials & Interfaces·Shuojun LiHeyou Han
Oct 21, 2020·Microbiological Research·Rajdeep JaswalT R Sharma

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