Real-time Imaging of Nascent DNA in Live Cells by Monitoring the Fluorescence Lifetime of DNA-Incorporated Thiazole Orange-Modified Nucleotides

被引：14

作者：

Kuba, Miroslav ^{[1
,2
]}

Khoroshyy, Petro ^{[1
]}

Lepsik, Martin ^{[1
]}

Kuzmova, Erika ^{[1
]}

Kodr, David ^{[1
]}

Kraus, Tomas ^{[1
]}

Hocek, Michal ^{[1
,2
]}

机构：

[1] Czech Acad Sci, Inst Organ Chem & Biochem, Flemingovo nam 2, Prague 6, Czech Republic

[2] Charles Univ Prague, Fac Sci, Dept Organ Chem, Hlavova 8, Prague 2, Czech Republic

来源：

ANGEWANDTE CHEMIE-INTERNATIONAL EDITION | 2023年 / 62卷 / 38期

关键词：

DNA Synthesis; Fluorescence Lifetime Imaging Microscopy; Fluorophores; Metabolic Labelling; Nucleotides; HYBRIDIZATION; DYNAMICS;

D O I：

10.1002/anie.202307548

中图分类号：

O6 [化学];

学科分类号：

0703 ;

摘要：

A modified 2 & PRIME;-deoxycytidine triphosphate derivative (dC(TO)TP) bearing a thiazole orange moiety tethered via an oligoethylene glycol linker was designed and synthesized. The nucleotide was incorporated into DNA by DNA polymerases in vitro as well as in live cells. Upon incorporation of dC(TO)TP into DNA, the thiazole orange moiety exhibited a fluorescence lifetime that differed significantly from the non-incorporated (i.e. free and non-covalently intercalated) forms of dC(TO)TP. When dC(TO)TP was delivered into live U-2 OS cells using a synthetic nucleoside triphosphate transporter, it allowed us to distinguish and monitor cells that were actively synthesizing DNA in real time, from the very first moments after the treatment. We anticipate that this probe could be used to study chromatin organization and dynamics.

引用

页数：5

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