CRISPR/Cas9-mediated tissue-specific knockout and cDNA rescue using sgRNAs that target exon-intron junctions in Drosophila melanogaster.
STAR Protoc
; 3(3): 101465, 2022 09 16.
Article
em En
| MEDLINE
| ID: mdl-35719725
ABSTRACT
In this protocol, we take CRISPR/Cas9 and Gal4/UAS approaches to achieve tissue-specific knockout in parallel with rescue of the knockout by cDNA expression in Drosophila. We demonstrate that guide RNAs targeting the exon-intron junction of target genes cleave the genomic locus of the genes, but not UAS-cDNA transgenes, in a tissue where Gal4 drives Cas9 expression. The efficiency of this approach enables the determination of pathogenicity of disease-associated variants in human genes in a tissue-specific manner in Drosophila. For complete details on the use and execution of this protocol, please refer to Yap et al. (2021).
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Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Drosophila melanogaster
/
Sistemas CRISPR-Cas
Limite:
Animals
Idioma:
En
Revista:
STAR Protoc
Ano de publicação:
2022
Tipo de documento:
Article
País de afiliação:
Estados Unidos