UCSC In-Silico PCR
  
>chrV:10129085-10129897 813bp GGCGGCATCATACAATTTTC CACATGAATAAGTCCGTGCG
GGCGGCATCATACAATTTTCtttataatttgaaccgcaatgtgaaacttt
tcatcgtcgaggaagatgttctatcgaaaaatgattcggaaatcgaagag
tatttgactaaattcacgttggttgttgttctcgacgagagttacgaacg
tactgcaaaagtgaacaatatctgccgtaaacaccacatcagatttattt
ctggagcaatctacggatggattggctacgcatttttcgatttcgatggc
catgcttatttagtgtaagttaaatcgttttaaaagtctcatcccaatat
ttcttttaaagaaaagctaaaagcccggattgcttgaatgaagaagaatc
ggaaaccggaaagacgagcacagtggttacggtagatgaagaattcgtgc
ttgaaacgttttcatatccatcatttgtcgaaacgctcaactctgatttc
acggcgaagaaaattgtgagaaagtgcaaacggatcgttccaacatcata
tttcctcgtgaaaagtatgctccgcgcttccagcgaaaataaacttaccg
gagtcactgaaaacgatattgagaaattaattccaatctggaacgaggag
gtggccgctggaaatcataccatcgatatgcaaccagttcaacccgatcg
atttgatcagtgagtattattcgcacgaaaattataaaatgaatcaataa
aaaacatagaatgtattatctttttaattatttaaaaaaacaagttaaaa
atgtatcgtaatttcaaaaaacggtagcattccgtcttaaagaCGCACGG
ACTTATTCATGTG

Primer Melting Temperatures
  Forward: 60.3 C ggcggcatcatacaattttc
Reverse: 60.1 C cacatgaataagtccgtgcg
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