Incubate 8 µl restriction digest with 4 µl PCR product for a minimum of 3 hours at 37 C prior to running on gel.
The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX).
To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility.
Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.
Mutant = 280 bp
Heterozygote = 140 bp and 280 bp
Wild type = 140 bp
Separate digested products on a 3.0% agarose gel.**