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Journal article · Book chapter

Genome editing: CRISPR-Cas9

By Hoof, Jakob B.1,2,3; Nødvig, Christina S.1; Mortensen, Uffe H.1,2,4

Edited by Tsang, Adrian; Grigoriev, Igor V.; de Vries, Ronald P.

From

Department of Biotechnology and Biomedicine, Technical University of Denmark1

Section for Synthetic Biology, Department of Biotechnology and Biomedicine, Technical University of Denmark2

Fungal Biomedicine and Biology, Section for Synthetic Biology, Department of Biotechnology and Biomedicine, Technical University of Denmark3

Eukaryotic Molecular Cell Biology, Section for Synthetic Biology, Department of Biotechnology and Biomedicine, Technical University of Denmark4

In the present chapter, we present the protocols and guidelines to facilitate implementation of CRISPR-Cas9 technology in fungi where few or no genetic tools are in place. Hence, we firstly explain how to identify dominant markers for genetic transformation. Secondly, we provide a guide for construction of Cas9/sgRNA episomal expression vectors.

Thirdly, we present how to mutagenize reporter genes to explore the efficiency of CRISPR-Cas9 in the relevant fungus and to ease subsequent CRISPR-mediated genetic engineering. Lastly, we describe how to make CRISPR-mediated marker-dependent and marker-free gene targeting.

Language: English
Publisher: Humana Press
Year: 2018
Edition: 2
Pages: 119-132
Series: Methods in Molecular Biology
ISBN: 1493978039 , 1493978047 , 9781493978038 and 9781493978045
ISSN: 19406029 and 10643745
Types: Journal article and Book chapter
DOI: 10.1007/978-1-4939-7804-5_11
ORCIDs: Hoof, Jakob B. and Mortensen, Uffe H.
Keywords

CRISPR Gene editing Gene targeting Mutagenesis Nonmodel fungi

Other keywords

CRISPR-Cas Systems Gene Editing Gene Targeting Genetic Engineering Genetic Vectors RNA, Guide, CRISPR-Cas Systems

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