Skip to main content
This website uses cookies to ensure you get the best experience. By continuing to use this site, you agree to the use of cookies.

Please note: Your browser does not support the features used on Addgene's website. You may not be able to create an account or request plasmids through this website until you upgrade your browser. Learn more

Please note: Your browser does not fully support some of the features used on Addgene's website. If you run into any problems registering, depositing, or ordering please contact us at [email protected] Learn more

Efficient multiplex biallelic zebrafish genome editing using a CRISPR nuclease system.
Jao LE, Wente SR, Chen W
Proc Natl Acad Sci U S A. 2013 Aug 5.
PubMed Article

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 lab plasmid cloning figure

Resources:

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

Plasmids from Article

ID Plasmid Purpose  
46757pT3TS-nCas9n
Loading...
46759pT7-gRNAgRNA cloning vector for in vitro transcription of target gRNA
Loading...
46760pT7EGFPgRNA
Loading...
46761pT7tyrgRNA
Loading...
47929pCS2-nCas9nexpression of an optimized Cas9 for genome-editing in zebrafish
Loading...
47930pT7goldRNAIn vitro transcription of golden gRNA
Loading...
47931pT7mitfagRNAin vitro transcription of mitfa gRNA
Loading...
47932pT7ddx19gRNAin vitro transcription of ddx19 gRNA
Loading...