Genome-Editing Custom Project General
CRISPR-Cas9 technology has emerged as a rapid, highly-precise
method to generate knock-out and knock-in mouse models. We can
perform your genome editing project for you from strategy design
through founder genotyping. We successfully created 34 new mouse
models at a 100% technical success rate in 2018. These included
seven gene deletions, fourteen point mutations, six epitope tags or
STOP codon insertions, four conditional alleles (floxed), one
fluorescent reporter knock-in, and two site-directed transgenes
inserted into the Rosa26 locus.
Basic steps for VGER mouse genome editing
- Contact email@example.com
to schedule an advisory meeting. Following the meeting, the
investigator completes the "VGER Gene Editing Service Form" (see
attachment below) and is provided with an estimate.
- A genome editing strategy is devised and reviewed with the
investigator. We now have four classes of CRISPR/Cas9 editing
strategies based on the donor DNA format and length of the desired
gene edit. See the attachment below for efficiency data fon each
type of project.
- ctRNAs are ordered and tested in vitro in a
ribonucleoprotein format with tracrRNA and Cas9 protein for
on-target efficiency at the desired genetic locus. See
the "MilliporeSigma 2018 Pricing" attachment below for
current reagent pricing.
- A donor ssDNA is designed for homology directed repair projects
and reviewed with the investigator.
- Typically, Cas9 protein, ctRNA(s), and a ssDNA oligo or dsDNA
are injected into mouse zygotes at the one cell stage.
- F0 pups are biopsied for genotyping and screened by a
VGER-developed PCR based assay.
- F0 Founder(s) are analyzed by Sanger Sequencing to identify
those containing the predicted desired gene edit.
- F0 Founder(s) is transferred to investigator for breeding
to WT and validation of the desired genome edit in the N1
- Breeding and screening of the N1 generation is also available
by estimate upon request.
here to contact the core