Targeted CRISPR-Cas9-based gene knockouts in the model brown alga Ectocarpus.
Ectocarpus
CRISPR
Cas9
brown alga
reverse genetics
transformation
Journal
The New phytologist
ISSN: 1469-8137
Titre abrégé: New Phytol
Pays: England
ID NLM: 9882884
Informations de publication
Date de publication:
09 2021
09 2021
Historique:
received:
02
12
2020
accepted:
20
05
2021
pubmed:
3
6
2021
medline:
13
8
2021
entrez:
2
6
2021
Statut:
ppublish
Résumé
Brown algae are an important group of multicellular eukaryotes, phylogenetically distinct from both the animal and land plant lineages. Ectocarpus has emerged as a model organism to study diverse aspects of brown algal biology, but this system currently lacks an effective reverse genetics methodology to analyse the functions of selected target genes. Here, we report that mutations at specific target sites are generated following the introduction of CRISPR-Cas9 ribonucleoproteins into Ectocarpus cells, using either biolistics or microinjection as the delivery method. Individuals with mutations affecting the ADENINE PHOSPHORIBOSYL TRANSFERASE (APT) gene were isolated following treatment with 2-fluoroadenine, and this selection system was used to isolate individuals in which mutations had been introduced simultaneously at APT and at a second gene. This double mutation approach could potentially be used to isolate mutants affecting any Ectocarpus gene, providing an effective reverse genetics tool for this model organism. The availability of this tool will significantly enhance the utility of Ectocarpus as a model organism for this ecologically and economically important group of marine organisms. Moreover, the methodology described here should be readily transferable to other brown algal species.
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
2077-2091Informations de copyright
© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.
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