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Why is the Tm provided on your oligonucleotide specification sheets different from the calculated Tm I obtained when I designed my oligos?
Oligonucleotides designed using our online SciTools® programs (e.g., the PrimerQuest Primer Design Tool, RealTime PCR Design Tool or the OligoAnlayzer® Tool) are ordered for a wide variety of applications, and not solely for PCR. The Tm shown on the spec sheets is calculated based on default conditions of 50 mM Na+ without taking into consideration Mg2+ and dNTP concentrations, both which influence Tm. To determine the correct Tm for your oligo under your specific reaction conditions, enter the sequence into the online OligoAnalyzer® Tool, making sure to change the defaults (32 mM Na+, 3 mM Mg2+, and 0.8 mM dNTPs) to your reactions conditions.