A user-friendly CRISPR/Cas9 system for mutagenesis of Neurospora crassa.
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
03 09 2024
03 09 2024
Historique:
received:
20
12
2023
accepted:
28
08
2024
medline:
4
9
2024
pubmed:
4
9
2024
entrez:
3
9
2024
Statut:
epublish
Résumé
As a widely used eukaryotic model organism, Neurospora crassa offers advantages in genetic studies due to its diverse biology and rapid growth. Traditional genetic manipulation methods, such as homologous recombination, require a considerable amount of time and effort. In this study, we present an easy-to-use CRIPSR/Cas9 system for N. crassa, in which the cas9 sequence is incorporated into the fungal genome and naked guide RNA is introduced via electroporation. Our approach eliminates the need for constructing multiple vectors, speeding up the mutagenesis process. Using cyclosporin-resistant-1 (csr-1) as a selectable marker gene, we achieved 100% editing efficiency under selection conditions. Furthermore, we successfully edited the non-selectable gene N-acylethanolamine amidohydrolase-2 (naa-2), demonstrating the versatility of the system. Combining gRNAs targeting csr-1 and naa-2 simultaneously increased the probability of finding mutants carrying the non-selectable mutation. The system is not only user-friendly but also effective, providing a rapid and efficient method for generating loss-of-function mutants in N. crassa compared to traditional methods.
Identifiants
pubmed: 39227671
doi: 10.1038/s41598-024-71540-x
pii: 10.1038/s41598-024-71540-x
doi:
Substances chimiques
Fungal Proteins
0
RNA, Guide, CRISPR-Cas Systems
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
20469Informations de copyright
© 2024. The Author(s).
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