Design strategies for countering the effect of fluorophore-quencher labelling on DNA hairpin thermodynamics.

We report the impact of fluorophore-quencher labelling on the thermodynamics of hairpin opening by testing five fluorophores and two quenchers labelled at the end and/or internal positions. Two counter strategies were introduced, label the hairpin probe at an internal position or append an external hairpin stem on the trigger strand to promote coaxial stacking hybridization. The observations remained valid for complex hairpin opening operations such as hybridization chain reaction.