UCSC In-Silico PCR
 
>chrIV:10487423-10488158 736bp GTAGATGGGAAGGCATTTCG TGATTTGCTCAATGAAAGCG
GTAGATGGGAAGGCATTTCGattgagtaaatctcgaatcatttgaaccat
ttccatgagaaaatcacgtttatcaatatccgatgatcgctcttccacat
attcactaaacatttgggcggacattttgtctaataatgccaatattaca
gagaaaaagtgaccacgagcatcatcatcagtgtgcttgtcgtcgtggat
tactcgaatcattgcttgaacaagtggacgataaaccattaaaatgaatg
atgtcaattctctctcagcgttaccttcaccacttttctgtaaaatatga
aatacttgtcatgtacgaatgataatctagtaccgcgtcagcgaatagtc
tctccagaatagcagcaataatactggcacactcagcggcacgatctgca
aattctcctttttcggttgccatattgtcgatttgtattatgtcaagagc
gatttgaagacatggaatcagaagttgttctcggcagattgaaagctgaa
catttatttaattcggatattttgtaatttttgcatgaagacaaattatt
gggaactatctaaaataatgtatctaaagatatttccttaccgaaaagaa
ccgagtttccacaatctgtgcaataaaattgagccgctcacgagtaacaa
tatttgatccaaaattattcatcaaatcaacaataaatccacaaagattc
tctggagataccgaatCGCTTTCATTGAGCAAATCA

Primer Melting Temperatures
  Forward: 59.5 C gtagatgggaaggcatttcg
Reverse: 60.0 C tgatttgctcaatgaaagcg
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