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Goldstein Lab CRISPR Plasmids Available from Addgene


Our Cas9-sgRNA plasmids contain both the Cas9 gene and an sgRNA gene. The inclusion of both components on a single plasmid facilitates archival and distribution, and makes it simpler to prepare injection mixes. The Cas9 coding sequence is codon-optimized and includes synthetic introns for efficient expression in C. elegans , and is driven by the eft-3 promoter for germline expression. The sgRNA is driven by the R07E5.16 U6 RNA promoter. Compared to the C. elegans CRISPR plasmids from John Calarco's lab, which are also available on Addgene, the biggest differences are 1) Inclusion of both components on the same plasmid, and 2) Use of a different U6 promoter to drive sgRNA expression. We have not compared the two systems side by side but would expect them to be similar in terms of efficiency.

plasmid-cartoon-overview

Additional Resources:

The Supplementary Information in the publication contains a protocol for using the plasmids. A website has been created for sharing protocols and information: http://wormcas9hr.weebly.com/

These plasmids are described in:

Engineering the Caenorhabditis elegans genome using Cas9-triggered homologous recombination. Dickinson DJ, Ward JD, Reiner DJ, Goldstein B. Nat. Methods . 2013 Sept 1. PubMed . PMC .).

Individual plasmids can be ordered via the links below:

ID Plasmid
47550 pDD122 (Peft-3::Cas9 + ttTi5605 sgRNA) : Cas9 + sgRNA plasmid that is targeted to a genomic site near the ttTi5605 Mos1 insertion allele. Add to Cart
47549 pDD162 (Peft-3::Cas9 + Empty sgRNA) : Cas9 + sgRNA plasmid that can be modified to cleave any Cas9 target site in the C. elegans genome. Add to Cart
47551 pDD104 (Peft-3::Cre) : Cre recombinase expression plasmid for removal of floxed selectable markers in the C. elegans germline. Add to Cart

Please note: eft-3 has officially been changed to eef-1A.1 Please see the eef-1A.1 WormBase entry for details.