CRISPR/Cas9 services and off-target analysis
- gRNA design and production — Guide RNAs (gRNAs) are designed and analyzed using state-of-the-art analysis software to determine potential off-target sites and secondary structure.
- gRNA in vitro activity test — Transfection or electroporation of Cas protein and gRNA into appropriate cell lines or embryos (along with repair template for knock-ins) is performed and activity assessed with PCR, Cel-I, restriction enzyme digests and/or sequencing as needed.
- Repair template design and production for knock-ins — Repair templates (ssODN, long ssDNA, AAV-packaged or donor plasmids) are generated along with homology arms (when appropriate) to be used when making knock-in animals or cell lines.
- Off-target analysis — Potential off-target sites are determined via standard software analysis tools and are empirically evaluated via PCR and DNA sequencing.
General molecular genetics and cloning services
- Cloning services — General cloning services such as making plasmids for qPCR standards, TA cloning for sequencing of mixed PCR products or complex cloning.
- Generation of targeting vectors/repair templates — Design and production of custom templates, including double-stranded plasmids (dsDNA), single-stranded oligonucleotides (ssODN), long single-stranded templates (lssDNA) or AAV-packaged templates.
Genotyping
- Assay development — Custom primer design and assay optimization.
- Genotyping analysis — DNA extraction from tail snips, ear punches or other tissue samples followed by endpoint PCR, restriction fragment length polymorphism (RFLP) or high-resolution melt (HRM) analysis. Droplet digital PCR (ddPCR) services are also available for determining transgene copy number.
Colony management
Organized and experienced management of rodent colonies is performed at an hourly rate. Services include setting up matings, weaning, animal identification, tissue biopsies for genotyping and comprehensive record keeping.
Model characterization
A wide variety of services are available for animal model characterization. Services are customized to the needs of the investigator and include histology, qPCR, imaging, tissue collection, behavioral analysis and other phenotyping assays. Droplet digital PCR (ddPCR) services are available to detect very subtle alterations in gene expression.
Custom projects
The AMC is available to discuss custom projects in which our molecular biology and animal modeling experience can be utilized. We have access to a wide range of equipment and have experience with many different techniques that may be beneficial to your research. Please contact us to set up a free consultation to determine how we can best assist you.