UCSC In-Silico PCR
 
>chrIII:1081624-1081758 135bp GCCATTTTTCACGCAATTTT GCATCAACGACGTTCAACAG
GCCATTTTTCACGCAATTTTtcttgaaatcctcggtatttccatgttcta
accctgtcgaaaagctgatcacacagtggacagaggagttccgagctcaa
atcgcgtcgaaccggCTGTTGAACGTCGTTGATGC

Primer Melting Temperatures
  Forward: 59.9 C gccatttttcacgcaatttt
Reverse: 60.3 C gcatcaacgacgttcaacag
The temperature calculations are done assuming 50 mM salt and 50 nM annealing oligo concentration. The code to calculate the melting temp comes from Primer3, the formula by Rychlik W, Spencer WJ and Rhoads RE NAR 1990, which can be activated in Primer3 with PRIMER_TM_FORMULA=0.

Help
  What is chr_alt & chr_fix?
Replicating in-Silico PCR results on local machine