Single nucleotide editing without DNA cleavage using CRISPR/Cas9-deaminase in the sea urchin embryo.
Dev Dyn
; 246(12): 1036-1046, 2017 12.
Article
em En
| MEDLINE
| ID: mdl-28857338
ABSTRACT
BACKGROUND:
A single base pair mutation in the genome can result in many congenital disorders in humans. The recent gene editing approach using CRISPR/Cas9 has rapidly become a powerful tool to replicate or repair such mutations in the genome. These approaches rely on cleaving DNA, while presenting unexpected risks.RESULTS:
In this study, we demonstrate a modified CRISPR/Cas9 system fused to cytosine deaminase (Cas9-DA), which induces a single nucleotide conversion in the genome. Cas9-DA was introduced into sea urchin eggs with sgRNAs targeted for SpAlx1, SpDsh, or SpPks, each of which is critical for skeletogenesis, embryonic axis formation, or pigment formation, respectively. We found that both Cas9 and Cas9-DA edit the genome, and cause predicted phenotypic changes at a similar efficiency. Cas9, however, resulted in significant deletions in the genome centered on the gRNA target sequence, whereas Cas9-DA resulted in single or double nucleotide editing of C to T conversions within the gRNA target sequence.CONCLUSIONS:
These results suggest that the Cas9-DA approach may be useful for manipulating gene activity with decreased risks of genomic aberrations. Developmental Dynamics 2461036-1046, 2017. © 2017 Wiley Periodicals, Inc.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Strongylocentrotus purpuratus
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Embrião não Mamífero
/
Sistemas CRISPR-Cas
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Edição de Genes
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article