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Notes
This assay will NOT distinguish hemizygous from homozygous transgenic animals.
Genotyping by PCR is performed to determine the absence or presence of the transgene. This strain has some variability in repeat size, therefore Southern blotting is performed to accurately determine the size, in kilobases, of the repeat within the transgene.
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.
Expected Results
Transgene = 348 bp
Internal positive control = 168 bp
JAX Protocol
Protocol Primers
| Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
|---|---|---|---|---|---|---|
| 15967 | TGT TTG CCT GCA ATA GGC | Transgene Forward | ||||
| 15968 | TGA TGG GAA AGC TAT TAT GAC C | Transgene Reverse | ||||
| 21157 | CTC CCA ACC CCA GAG GTA GT | Internal Positive Control Forward | ||||
| 21159 | CTA TAG GGC CTG GGT CAG TG | Internal Positive Control Reverse |