UCSC In-Silico PCR
  
>chrI:9966375-9967175 801bp CTCACCACGCACAACTTGAC ATTTACGGCCGCTGTGATAC
CTCACCACGCACAACTTGACatgaagaaaaagtaatgaacaacagcaaga
gcactacatccaacattttctcctccttgcgtaacaagtgttgttggagt
agatccatacgttccacaagcgaatccatcttcaccgactacgcctgtag
aaatacatttttagtttctaaaaaatctgatttaaaaaaaacagaaaatt
caactcacctatcatataaaccgtagaaattgctagataacagaacgcga
gatacaaaatcggtcgaactggatacgcgaatcgtgataaatcaacaaga
aatgtgacaagagtgaataaagagcacacgaaacatgccacagaccatgc
tgctgtccagattcgaagaattcgtcgaacacgtccatcaaacatcattg
gtacttgattgtcagcttcacatgttaatgaacattcaggaaggcgtcca
ctgagaagaggcaatgaaaaatgagatccgcttgttgtatgcatatgatg
agggcatttttttggagaaaggacaacttcgttggcaattgttgaaacac
cgaatgtgacgtcattactacttctgtaacttgaaatattttatatttcg
atctacggaaacttgttttattgttcgataaaacaaagaatttagaaatc
tcgatgtacatttttttcggaaaatttctaaattcaaaaactttgataaa
aaactatttgacgctttggagtaggagtttttatggggctcaaaaattcg
gaataatacttaaattcagccaaacttgtcaGTATCACAGCGGCCGTAAA
T

Primer Melting Temperatures
  Forward: 60.4 C ctcaccacgcacaacttgac
Reverse: 60.0 C atttacggccgctgtgatac
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