Tr ansient stat es during the annealing of mismatc hed and bulgedolig onucleotides

Abstract

Oligonucleotide hybridization is crucial in various biological, prebiotic and nanotechnological processes, including gene regulation, non-enzymatic primer extension and DNA nanodevice assembly. Although extensive research has focused on the thermodynamics and kinetics of nucleic acid h ybridization, the beha vior of comple x mixtures and the outcome of competition f or target binding remain less w ell understood. In this study, we in v estigate the impact of mismatches and bulges in a 12 bp DNA or RNA duplex on its association ( k on ) and dissociation ( k off ) kinetics. We find that such defects ha v e relativ ely small effects on the association kinetics, while the dissociation kinetics vary in a position-dependent manner by up to 6 orders of magnitude. Building upon this observ ation, w e e xplored a competition scenario in v olving multiple oligonucleotides, and observed a transient low specificity of probe hybridization to fully versus partially complementary targets in solution. We characterize these long-lived met ast able st ates and their e v olution to w ard equilibrium, and show that sufficiently long-lived mis-paired duple x es can serv e as substrates f or prebiotically rele v ant chemical cop ying reactions. Our results suggest that transient lo w accuracy states ma y spontaneously emerge within all complex nucleic acid systems comprising a large enough number of competing strands, with potential repercussions for gene regulation in the realm of modern biology and the prebiotic preservation of genetic information.