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Title: Single-molecule fluorescence detection of a tricyclic nucleoside analogue
Fluorescent nucleobase surrogates capable of Watson–Crick hydrogen bonding are essential probes of nucleic acid structure and dynamics, but their limited brightness and short absorption and emission wavelengths have rendered them unsuitable for single-molecule detection. Aiming to improve on these properties, we designed a new tricyclic pyrimidine nucleoside analogue with a push–pull conjugated system and synthesized it in seven sequential steps. The resulting C -linked 8-(diethylamino)benzo[ b ][1,8]naphthyridin-2(1 H )-one nucleoside, which we name ABN, exhibits ε 442 = 20 000 M −1 cm −1 and Φ em,540 = 0.39 in water, increasing to Φ em = 0.50–0.53 when base paired with adenine in duplex DNA oligonucleotides. Single-molecule fluorescence measurements of ABN using both one-photon and two-photon excitation demonstrate its excellent photostability and indicate that the nucleoside is present to > 95% in a bright state with count rates of at least 15 kHz per molecule. This new fluorescent nucleobase analogue, which, in duplex DNA, is the brightest and most red-shifted known, is the first to offer robust and accessible single-molecule fluorescence detection capabilities.  more » « less
Award ID(s):
1800529 1709796
PAR ID:
10218719
Author(s) / Creator(s):
; ; ; ; ; ; ; ; ;
Date Published:
Journal Name:
Chemical Science
Volume:
12
Issue:
7
ISSN:
2041-6520
Page Range / eLocation ID:
2623 to 2628
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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