A co-CRISPR strategy for efficient genome editing in Caenorhabditis elegans
Ghanta, Krishna S.
Conte, Darryl Jr.
Mello, Craig C.
Student AuthorsHeesun Kim; Krishna S. Ghanta
UMass Chan AffiliationsRNA Therapeutics Institute
Program in Molecular Medicine
Document TypeJournal Article
KeywordsAnimals; Base Sequence; CRISPR-Associated Proteins; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Clustered Regularly Interspaced Short Palindromic Repeats; Deoxyribonucleases; Genetic Markers; *Genome, Helminth; Homologous Recombination; Molecular Sequence Data; Mutagenesis, Insertional; Plasmids; Sequence Analysis, DNA
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AbstractGenome editing based on CRISPR (clustered regularly interspaced short palindromic repeats)-associated nuclease (Cas9) has been successfully applied in dozens of diverse plant and animal species, including the nematode Caenorhabditis elegans. The rapid life cycle and easy access to the ovary by micro-injection make C. elegans an ideal organism both for applying CRISPR-Cas9 genome editing technology and for optimizing genome-editing protocols. Here we report efficient and straightforward CRISPR-Cas9 genome-editing methods for C. elegans, including a Co-CRISPR strategy that facilitates detection of genome-editing events. We describe methods for detecting homologous recombination (HR) events, including direct screening methods as well as new selection/counterselection strategies. Our findings reveal a surprisingly high frequency of HR-mediated gene conversion, making it possible to rapidly and precisely edit the C. elegans genome both with and without the use of co-inserted marker genes.
SourceGenetics. 2014 Aug;197(4):1069-80. doi: 10.1534/genetics.114.166389. Epub 2014 May 30. Link to article on publisher's site
Permanent Link to this Itemhttp://hdl.handle.net/20.500.14038/33369
Related ResourcesLink to Article in PubMed
Copyright © 2014 by the Genetics Society of America. Available freely online through the author-supported open access option.
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