A method of predicting the melting temperature of an LNA-modified fluorescence-quenched oligonucleotide comprising: (a) a computer system receiving a data input from a user, the computer comprising a processor, and instructions executable by the processor; (b) responsive to the data input from a user, the computer system calculating a T m value using the equation: T m = Δ H o Δ S o + R ln C 1 − C 2 / 2 , wherein Δ H o = Δ H DNA o + ΔΔ H LNA o , ΔΔ H LNA o = ∑ i , j = A , C , G , T N ij n − n ΔΔ H ij n − n and Δ S o = Δ S DNA o + ΔΔ S LNA o , ΔΔ S LNA o = ∑ i , j = A , C , G , T N ij n − n ΔΔ S ij n − n wherein: T m is the melting temperature; C 1 and C 2 are concentrations of nucleic acid single strands that anneal to form a duplex, Δ H o and Δ S o are the transition enthalpy and entropy values, respectively, for duplex formation, Δ H o DNA and Δ S o DNA are transition enthalpies and entropies, respectively, for duplex formation of unmodified, perfectly matched DNA sequences, ΔΔ H o LNA and ΔΔ S o LNA are enthalpy and entropy adjustment values, respectively, that account for the thermodynamic effects of Locked Nucleic Acid (LNA) modifications and mismatches on duplex formation, and N ij n − n is the number of times the particular n-n doublet ij appears in the duplex; and (c) providing an output to a display of the calculated T m , Δ H o , and/or Δ S o values, wherein the oligonucleotide comprises at least two Locked Nucleic Acid (LNA) modifications, wherein the Δ H o and Δ S o values are determined using nearest neighbor parameters, and wherein the oligonucleotide is used to distinguish single nucleotide polymorphisms (SNPs). (a) a computer system receiving a data input from a user, the computer comprising a processor, and instructions executable by the processor; (b) responsive to the data input from a user, the computer system calculating a T m value using the equation: T m = Δ H o Δ S o + R ln C 1 − C 2 / 2 , wherein Δ H o = Δ H DNA o + ΔΔ H LNA o , ΔΔ H LNA o = ∑ i , j = A , C , G , T N ij n − n ΔΔ H ij n − n and Δ S o = Δ S DNA o + ΔΔ S LNA o , ΔΔ S LNA o = ∑ i , j = A , C , G , T N ij n − n ΔΔ S ij n − n wherein: T m is the melting temperature; C 1 and C 2 are concentrations of nucleic acid single strands that anneal to form a duplex, Δ H o and Δ S o are the transition enthalpy and entropy values, respectively, for duplex formation, Δ H o DNA and Δ S o DNA are transition enthalpies and entropies, respectively, for duplex formation of unmodified, perfectly matched DNA sequences, ΔΔ H o LNA and ΔΔ S o LNA are enthalpy and entropy adjustment values, respectively, that account for the thermodynamic effects of Locked Nucleic Acid (LNA) modifications and mismatches on duplex formation, and N ij n − n is the number of times the particular n-n doublet ij appears in the duplex; and T m is the melting temperature; C 1 and C 2 are concentrations of nucleic acid single strands that anneal to form a duplex, Δ H o and Δ S o are the transition enthalpy and entropy values, respectively, for duplex formation, Δ H o DNA and Δ S o DNA are transition enthalpies and entropies, respectively, for duplex formation of unmodified, perfectly matched DNA sequences, ΔΔ H o LNA and ΔΔ S o LNA are enthalpy and entropy adjustment values, respectively, that account for the thermodynamic effects of Locked Nucleic Acid (LNA) modifications and mismatches on duplex formation, and N ij n − n is the number of times the particular n-n doublet ij appears in the duplex; and (c) providing an output to a display of the calculated T m , Δ H o , and/or Δ S o values, <EOS>