Buscar Close Search
Page Menu

Using Cas12a in zebrafish

Cas12a (also referred to as Cpf1) is a recently described CRISPR-associated (Cas) protein that can efficiently introduce targeted DNA breaks in mammalian cells and zebrafish embryos. In collaboration with the Wolfe Lab, we have recently identified optimized variants of Cas12a and its crRNA and applied them in zebrafish, where they are highly active. Below is our current protocol for generating Cas12a crRNAs for injection in zebrafish.
Note - we no longer use the "GC" swap for the crRNA backbone. Across many more targets that we have tested in zebrafish since the NAR paper, we find that the extended "dr"-crRNA without GC works best. This is reflected in the PDF here. 

Generation of crRNA templates, crRNA, and injection (updated 8/28/20): PDF.

For Cas12a protein, you can generate your own using the pET plasmid from Addgene for recombinant expression, or purchase LbCas12a from New England Biolabs or IDT