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Differentiating a Least-Stable Single Nucleotide Mismatch in DNA Via Metal Ion-Mediated Base Pairing and Using Thioflavin T as an Extrinsic Fluorophore

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posted on 2021-03-08, 21:06 authored by Srikrishna Pramanik, Laxmikanta Khamari, Saptarshi Mukherjee
Monitoring the DNA dynamics in solution has great potential to develop new nucleic acid-based sensors and devices. With spectroscopic approaches, both at the ensemble average and single-molecule resolution, this study is directed to differentiate a single nucleotide mismatch (SNM) via a metal ion-stabilized mismatched base-pairing (C–Ag+–C/C–Cu2+–T) (C = cytosine, T = thymine) and site-selective extrinsic fluorophore, specifically, Thioflavin T (ThT). This is the first approach of its kind where dynamic quantities like molecular diffusion coefficients and diffusion times have been utilized to distinguish the least-stable SNM (CC & CT) formed by the most discriminating nucleobase, specifically, cytosine in a 20-mer duplex DNA. Additionally, this work also quantifies metal ions (Ag+ and Cu2+) at lower concentrations using fluorescence correlation spectroscopy. Our results can provide greater molecular-level insights into the mismatch-dependent metal–DNA interactions and also illuminate ThT as a new fluorophore to monitor the dynamics involved in DNA–metal composites.

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