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CONTACT JAX micetech@jax.org 1.800.422.6423 (US) 1.207.288.5845 (International) LEARN MORE ABOUT JAX MICE AND SERVICES JAX.ORG/JMCRS Frequently Asked Questions How large of an insertion can JAX make with CRISPR/Cas9? We can insert up to 4-6 Kb using CRISPR/Cas9 in mouse zygotes. If a larger insertion is needed, JAX has other methods such as Bxb1 or transgenics that we can consider. What is the risk of off-target effects from CRISPR? When we design a CRISPR project, we evaluate and select guide RNA that are predicted to have low off-target risks. If there is a high risk of off-target effects, we can perform PCR analysis on those sites for an additional cost. In addition, the mosaic founder mice are backcrossed to breed away from any potential off-target modifications. What is the purpose of breeding to the N1 generation? All CRISPR projects include the breeding to N1 generation. By breeding to N1 and re-genotyping the mice, we can guarantee that the genetic modification is germline transmissible. In addition, founders are mosaic, and backcrossing resolves alleles. How does JAX sequence the genetic modification? We use a combination of short-range and long-range sequencing techniques to ensure that the modification is intact and in the expected genomic location. What is included in the free feasibility assessment? Each assessment may be different depending on the model, but JAX will identify suitable guide RNA(s) that target the region with low risk of off-target effects, assess risk of adverse phenotypes that may impact viability or fertility, and determine if the proposed design will meet the intended scientific goal. If complications are discovered, JAX will communicate the risks and discuss potential alternatives to mitigate risks. Will JAX provide a genotyping protocol for the mutation? JAX will provide primer sequences for you to use in your current genotyping platform, as well as sequencing if you would like to design your own primers. Can JAX provide homozygous mice? Yes, we can add on a custom breeding project to breed to homozygosity. We can also breed to other strains (a cre- expressing strain, for example) or produce study-ready cohorts. How do I initiate a new project? Your JAX sales representative will ask for additional details about the model that you would like to generate. Our study directors will then perform a feasibility assessment, and generate a Statement of Work (SOW), which will typically be provided to you within 1 week. If you would like to proceed, you must sign the SOW and provide a PO to initiate the project. Crispr/Cas9 In Zygotes Model Generation

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