In vivo Gene-Edited Mouse Production by CRISPR/Cas9
Responsibilities of the Investigator
- Obtain IACUC approval for your project before requesting service.
- Discuss your intended design and target sequence with the Facility Directors before initiation of the project. The Facility does not currently perform gRNA or Cas9 RNA transcription or synthesis. The Facility requests that RNAs to be injected be designed, tested, and produced according to rigorous validation procedures from approved vendors. The Facility can assist in coordinating such efforts with a relevant Core that performs this work. Please inform the Facility Directors if you intend to perform IVT in your lab or use a non-approved vendor.
- If your project design involves homology-directed repair, please inform the Facility and provide plasmid or a targeting oligo. We recommend that targeting oligos are ordered from IDT.
- Genotype the founders that result following injection of your CRISPR/Cas9 components. NOTE: Please be aware that genotyping these deletions and/or mutations is complex and may take an extended period of time. Your pups will be weaned onto your protocol and held in the Transgenic Facility for three additional weeks until your genotyping is complete, at which time the founders will be transferred to your animal room. If your prefer, sequence-based genotyping can be performed by the Facility staff for an additional fee.
- Submit a Request Form in iLabs to initiate your project.
Services Provided by the Facility
- The Transgenic Facility will quantify and dilute your RNA (and DNA) components to the concentrations appropriate for injection.
- We will use F1 hybrid FVB x C57Bl/6 mouse embryos, unless requested by the user and approved by the Facility.The Facility will collect biopsies for genotyping.
- The Facility will guarantee the generation of two founders for gene-deletion experiments only. For HDR-based experiments, the Facility will guarantee injection and transfer of at least 250 embryos and/or 30 pups born, although reasonable efforts will be made to generate the mice requested.
- NOTE: The Facility cannot guarantee germline transmission.
- The Facility will provide tattoo identification of the resulting mice you choose to have transferred to your animal room.
Nov 04, 2019