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Chen and Wente Lab CRISPR Plasmids Available from Addgene


We have synthesized a Cas9 coding sequence with nuclear localization signals (nls) at both its amino and carboxyl termini and codons optimized for zebrafish expression (nls-zCas9-nls). Injection of in vitro transcribed nls-zCas9-nls mRNA with a gene-specific chimeric guide RNA consistently resulted in high mutagenic efficiency in both somatic and germline cells. Injected fish often exhibit loss-of-function phenotype, indicating prevalent biallelic inactivation. The injected fish produces predominantly mutation-carrying progeny. In addition, multiple loci can be efficiently targeted simultaneously in the same fish.

Chen-Wente-lab-plasmid-cloning-cartoon

Resources:

A protocol for synthesizing gRNAs: Icon gRNA plasmid construction protocol (66.8 KB)

These plasmids are descirbed in:

Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system. Jao LE, Wente SR, Chen W. Proc Natl Acad Sci . 2013 Aug 25. PubMed .

Individual plasmids can be ordered via the links below:

ID Plasmid
46761 pT7tyrgRNA : A gRNA targeting Tyr. Add to Cart
46760 pT7EGFPgRNA : A gRNA targeting EGFP. Add to Cart
46757 pT3TS-nCas9n : A Cas9 expression plasmid. Add to Cart
46759 pT7-gRNA : A gRNA core. Add to Cart
47929 pCS2-nCas9n : For expression of an optimized Cas9 for genome-editing in zebrafish. Add to Cart
47930 pT7goldRNA : A plasmid for in vitro transcription of golden gRNA. Add to Cart
47931 pT7mitfagRNA :A plasmid for in vitro transcription of mitfa gRNA. Add to Cart
47932 pT7ddx19gRNA : A plasmid for in vitro transcription of ddx19 gRNA. Add to Cart