FINE-TUNED ULTRASPECIFIC NUCLEIC ACID HYBRIDIZATION PROBES

摘要

Compositions and methods for highly specific nucleic acid probes and primers are provided. The probe system comprises a complement strand and a protector stand that form a partially double- stranded probe. The reaction standard free energy of hybridization between the probe and target nucleic acid as determined by Expression 1 (?Go rxn = ?Go t-TC - ?Go nh-PC + (?Go v-TC - ?Go h-PC)) is from about -4 kcal/mol to about +4 kcal/mol. Alternatively, the reaction standard free energy of hybridization between the probe and target nucleic acid is determined by Expression 1 to be within 5 kcal/mol of the standard free energy as determined by Expression 2 (-R?ln(([P]0 - [C]0)/[C]0)]), where the [P]0 term of Expression 2 equals the concentration of the protector strand and the [C]0 term of Expression 2 equals the concentration of the complement strand. In addition, a method for on-the-fly fine tuning of a reaction using the present probe is provided.