High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
PLoS Genet
; 17(11): e1009755, 2021 11.
Article
em En
| MEDLINE
| ID: mdl-34748534
ABSTRACT
Gene editing in C. elegans using plasmid-based CRISPR reagents requires microinjection of many animals to produce a single edit. Germline silencing of plasmid-borne Cas9 is a major cause of inefficient editing. Here, we present a set of C. elegans strains that constitutively express Cas9 in the germline from an integrated transgene. These strains markedly improve the success rate for plasmid-based CRISPR edits. For simple, short homology arm GFP insertions, 50-100% of injected animals typically produce edited progeny, depending on the target locus. Template-guided editing from an extrachromosomal array is maintained over multiple generations. We have built strains with the Cas9 transgene on multiple chromosomes. Additionally, each Cas9 locus also contains a heatshock-driven Cre recombinase for selectable marker removal and a bright fluorescence marker for easy outcrossing. These integrated Cas9 strains greatly reduce the workload for producing individual genome edits.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Caenorhabditis elegans
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Genoma Helmíntico
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Sistemas CRISPR-Cas
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Edição de Genes
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Proteína 9 Associada à CRISPR
Limite:
Animals
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article