An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>

ABSTRACT Candida albicans is the most common fungal pathogen of humans. Historically, molecular genetic analysis of this important pathogen has been hampered by the lack of stable plasmids or meiotic cell division, limited selectable markers, and inefficient methods for generating gene knockouts. Th...

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Autores principales:	Namkha Nguyen, Morgan M. F. Quail, Aaron D. Hernday
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	CRISPR Candida albicans Cas9 gene addback gene knockout genetics Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/f4dc70938e6b4a3f9c720f58bed2963e
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spelling	oai:doaj.org-article:f4dc70938e6b4a3f9c720f58bed2963e2021-11-15T15:21:46ZAn Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>10.1128/mSphereDirect.00149-172379-5042https://doaj.org/article/f4dc70938e6b4a3f9c720f58bed2963e2017-04-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphereDirect.00149-17https://doaj.org/toc/2379-5042ABSTRACT Candida albicans is the most common fungal pathogen of humans. Historically, molecular genetic analysis of this important pathogen has been hampered by the lack of stable plasmids or meiotic cell division, limited selectable markers, and inefficient methods for generating gene knockouts. The recent development of clustered regularly interspaced short palindromic repeat(s) (CRISPR)-based tools for use with C. albicans has opened the door to more efficient genome editing; however, previously reported systems have specific limitations. We report the development of an optimized CRISPR-based genome editing system for use with C. albicans. Our system is highly efficient, does not require molecular cloning, does not leave permanent markers in the genome, and supports rapid, precise genome editing in C. albicans. We also demonstrate the utility of our system for generating two independent homozygous gene knockouts in a single transformation and present a method for generating homozygous wild-type gene addbacks at the native locus. Furthermore, each step of our protocol is compatible with high-throughput strain engineering approaches, thus opening the door to the generation of a complete C. albicans gene knockout library. IMPORTANCE Candida albicans is the major fungal pathogen of humans and is the subject of intense biomedical and discovery research. Until recently, the pace of research in this field has been hampered by the lack of efficient methods for genome editing. We report the development of a highly efficient and flexible genome editing system for use with C. albicans. This system improves upon previously published C. albicans CRISPR systems and enables rapid, precise genome editing without the use of permanent markers. This new tool kit promises to expedite the pace of research on this important fungal pathogen.Namkha NguyenMorgan M. F. QuailAaron D. HerndayAmerican Society for MicrobiologyarticleCRISPRCandida albicansCas9gene addbackgene knockoutgeneticsMicrobiologyQR1-502ENmSphere, Vol 2, Iss 2 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	CRISPR Candida albicans Cas9 gene addback gene knockout genetics Microbiology QR1-502
spellingShingle	CRISPR Candida albicans Cas9 gene addback gene knockout genetics Microbiology QR1-502 Namkha Nguyen Morgan M. F. Quail Aaron D. Hernday An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
description	ABSTRACT Candida albicans is the most common fungal pathogen of humans. Historically, molecular genetic analysis of this important pathogen has been hampered by the lack of stable plasmids or meiotic cell division, limited selectable markers, and inefficient methods for generating gene knockouts. The recent development of clustered regularly interspaced short palindromic repeat(s) (CRISPR)-based tools for use with C. albicans has opened the door to more efficient genome editing; however, previously reported systems have specific limitations. We report the development of an optimized CRISPR-based genome editing system for use with C. albicans. Our system is highly efficient, does not require molecular cloning, does not leave permanent markers in the genome, and supports rapid, precise genome editing in C. albicans. We also demonstrate the utility of our system for generating two independent homozygous gene knockouts in a single transformation and present a method for generating homozygous wild-type gene addbacks at the native locus. Furthermore, each step of our protocol is compatible with high-throughput strain engineering approaches, thus opening the door to the generation of a complete C. albicans gene knockout library. IMPORTANCE Candida albicans is the major fungal pathogen of humans and is the subject of intense biomedical and discovery research. Until recently, the pace of research in this field has been hampered by the lack of efficient methods for genome editing. We report the development of a highly efficient and flexible genome editing system for use with C. albicans. This system improves upon previously published C. albicans CRISPR systems and enables rapid, precise genome editing without the use of permanent markers. This new tool kit promises to expedite the pace of research on this important fungal pathogen.
format	article
author	Namkha Nguyen Morgan M. F. Quail Aaron D. Hernday
author_facet	Namkha Nguyen Morgan M. F. Quail Aaron D. Hernday
author_sort	Namkha Nguyen
title	An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
title_short	An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
title_full	An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
title_fullStr	An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
title_full_unstemmed	An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>
title_sort	efficient, rapid, and recyclable system for crispr-mediated genome editing in <named-content content-type="genus-species">candida albicans</named-content>
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/f4dc70938e6b4a3f9c720f58bed2963e
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An Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in <named-content content-type="genus-species">Candida albicans</named-content>

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