UCSC In-Silico PCR
 
>chrIII:5048076-5048635 560bp GTCCTGGTTCAGCTTGCTTC TTCAGCATAAAAATGACCCAAG
GTCCTGGTTCAGCTTGCTTCacaatatacatatcgccactatcagtgatt
gtcatggaatttactaaaaaaaaatatattttaaaaaatgttttatacca
gaaaacttacataatccttcgataaacgactcatgaacaacatctccatc
aagtgaaattactcttaaaattgtcgttcgatcagagaattcagctgatg
tgtagattatctgtaaatttgaataattaataacttaagtgaaaataatt
tattgtaccctgtctttgtacgaaattacttgtggaaatggggccatttt
cttgaagaatttcttgtcttcgtgcagagttcggagagtttttccgttga
atttatataaaccgccttttgaagctacctgaaaaatgaataatttttaa
ttgttgttgtttaattataatatacccatatattgttgtcttggtcgtaa
acaacgttataaggaactcttgcccctcttggctccaaatgaagaagcat
ttcatcaaatcgttgataatcactggtgtctgacggttCTTGGGTCATTT
TTATGCTGAA

Primer Melting Temperatures
  Forward: 60.0 C gtcctggttcagcttgcttc
Reverse: 59.1 C ttcagcataaaaatgacccaag
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