Designing Oligos Or Primers To Genotype Transgenic Mice

1. Look for a unique junction in your transgene. For example, a transgene with a CMV promoter and a myc gene: CMV-Myc.

2. Take ~0.5-1kb of sequence from this junction between CMV and myc sequence, and paste in Primer 3.

3. Primer 3 always makes great primers for transgenic identification. Select "mispriming rodent_and_simple" to avoid mispriming in the mouse's own genome

4. Select products that identify that junction, and amplify fragments from 200-600bp. Don't make the product too large because the PCR gets harder to get.

5. Select oligos/primers of ~20 nucleotides in size (20mer) and a melting temp of 60C=Tm

6. Good place to order oligos: IDTDNA, SigmaAldrich