CRISPR/Cas9
- Guide design and preparation
- Blastocyst assay to assess activity of guides
- Establishment of genotyping strategy
- Repair template design
- Compatible with most efficient guide(s)
- Unique restriction site for downstream screening of founders
- Pronuclear microinjection of 150 embryos
- Transfer of embryos to pseudopregnant recipients
- Optional screening of founder animals
- Transfer of founder litters at weaning
Rodent Model Generation
CRISPR/Cas9
- Loss of function alleles
- Knockout
- Large genomic deletions
- Hypomorphic
Targeted Insertions
- Patient mutations
- SNPs
- Conditional alleles
- ROSA26 insertions
- Reporter tags
Simple Transgenics
- Reporters
- Overexpression vectors
ES Cell Targeting
- Conventional
- CRISPR induced recombination
- CRISPR guides and compatible ROSA26 targeting vector
Preparation of commercially available ES cells (EUCOMM / KOMP)
Assisted Reproductive Techniques
Mouse
- Sperm
- Embryos
- Oocytes
- Ovaries
Rat
- Sperm
- Embryos
Mouse
- In vitro fertilization
- Transfer of cryopreserved embryos
- Pathogen-free
- Ovarian transplant
Rat
- Transfer of cryo-preserved embryos
Consultation
- Design of transgenic constructs
- Screening strategies
- Breeding schemes
Maintenance of Stock Lines
- Cre stains (ex. Ella)
- Reporters (ex TdTomato, GFP)
- Male/female breeders and controls (ex. C57B6J)