CHOPCHOP v2: Improved online CRISPR/Cas9 target prediction tool

Bill Reid, Ph.D. Postdoctoral Researcher, Institute for Bioscience and Biotechnology Research, University of Maryland College Park

Bill Reid, Ph.D. Postdoctoral Researcher, Mosquito and Fly Research Unit, USDA/ARS, Center for Agricultural, Medical and Veterinary Entomology, Gainesville, Florida MORE ABOUT THE AUTHOR

In a paper published in Nucleic Acids Research, Labun et al. (2016), the developers of CHOPCHOP, report on their updated version: CHOPCHOPv2 .

The first version of CHOPCHOP was released by a team of researchers from Harvard University a little over two years ago and quickly gained popularity as the tool to design guides for CRISPR/Cas9.

ChopChopThe program was simple to use, generated visual output, and allowed users to tailor their analysis for off-target stringency, and designed primers flanking the target sites to aid in the downstream analysis of Cas9 activity.


spCas9 and Cpf1 differ in their PAM preference and in the DNA breaks generated (image from Sontheimer and Wolfe, 2015)

The CHOPCHOP team also supported the requests of multiple researchers requesting the addition of genomes and presently includes 37 genomes among 32 species.

Now hosted at the University of Bergen, Norway, there are several advances in the program including new scanning for Cas9 and Cpf1 nickases, and the ability to use scan for custom user-defined PAMs. The ‘options’ available allow for new efficiency scoring results, based off of efficiency models from five different guide efficiency prediction algorithms, along with the more conservative approach to score based on a ‘G’ at position 20.

Screenshot of CHOPCHOPv2 output showing the results of predicted cleavage sites in the A/T rich genome of Plasmodium falciparum. The upper panel shows the predicted sites for spCas9 cleavage (PAM = NGG) while the lower panel shows the predicted cleavage sites for Cpf1 (PAM = TTN).

The program also allows for the targeting of promoter (or upstream) regions, and users can also now design guides of various length, ranging from 14 to 30 nucleotides.

To further improve on finding suitable guides, CHOPCHOPv2 can now also scans for self-complementarity of the predicted guide as part of a sgRNA, to identify possible internal gRNA interactions that could reduce the ability of the Cas9-sgRNA complex to cleave.

Taken together, these updates to CHOPCHOP incorporate the current best practices for guide RNAs, while still retaining the transparency of the models used for the assessment of guide efficiency in a clear, easy-to-use interface.
Labun K, Montague TG, Gagnon JA, Thyme SB, Valen E. 2016. CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering. Nucl Acids Res. doi: 10.1093/nar/gkw398.

Sontheimer EJ, Wolfe SA. 2015. Cas9 gets a classmate. Nat Biotechnol. doi:10.1038/nbt.3426.


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