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

共 37 条

[11] DNA Stains as Surrogate Nucleobases in Fluorogenic Hybridization Probes [J].

Hovelmann, Felix ;

Seitz, Oliver .

ACCOUNTS OF CHEMICAL RESEARCH, 2016, 49 (04) :714-723

[12] Sequence dependence of fluorescence emission and quenching of doubly thiazole orange labeled DNA: Effective design of a hybridization-sensitive probe [J].

Ikeda, Shuji ;

Kubota, Takeshi ;

Kino, Katsuhito ;

Okamoto, Akimitsu .

BIOCONJUGATE CHEMISTRY, 2008, 19 (08) :1719-1725

[13] Exciton-Controlled Hybridization-Sensitive Fluorescent Probes: Multicolor Detection of Nucleic Acids [J].

Ikeda, Shuji ;

Kubota, Takeshi ;

Yuki, Mizue ;

Okamoto, Akimitsu .

ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, 2009, 48 (35) :6480-6484

[14] Design of thiazole orange oligonucleotide probes for detection of DNA and RNA by fluorescence and duplex melting [J].

Klimkowski, Piotr ;

De Ornellas, Sara ;

Singleton, Daniel ;

El-Sagheer, Afaf H. ;

Brown, Tom .

ORGANIC & BIOMOLECULAR CHEMISTRY, 2019, 17 (24) :5943-5950

[15] Nucleotides Bearing Red Viscosity-Sensitive Dimethoxy-Bodipy Fluorophore for Enzymatic Incorporation and DNA Labeling [J].

Kuba, Miroslav ;

Pohl, Radek ;

Kraus, Tomas ;

Hocek, Michal .

BIOCONJUGATE CHEMISTRY, 2023, 34 (01) :133-139

[16] Nucleotide-Bearing Benzylidene-Tetrahydroxanthylium Near-IR Fluorophore for Sensing DNA Replication, Secondary Structures and Interactions [J].

Kuba, Miroslav ;

Kraus, Tomas ;

Pohl, Radek ;

Hocek, Michal .

CHEMISTRY-A EUROPEAN JOURNAL, 2020, 26 (52) :11950-11954

[17] Hybridization-Sensitive Fluorescent Probe for Long-Term Monitoring of Intracellular RNA [J].

Kubota, Takeshi ;

Ikeda, Shuji ;

Yanagisawa, Hiroyuki ;

Yuki, Mizue ;

Okamoto, Akimitsu .

BIOCONJUGATE CHEMISTRY, 2009, 20 (06) :1256-1261

[18] Flow cytometric determination of cell cycle progression via direct labeling of replicated DNA [J].

Kuzmova, Erika ;

Zawada, Zbigniew ;

Navratil, Michal ;

Gunterova, Jana ;

Kraus, Tomas .

ANALYTICAL BIOCHEMISTRY, 2021, 614

[19] A Kinetic and Fluorogenic Enhancement Strategy for Labeling of Nucleic Acids [J].

Loehr, Morten O. ;

Luedtke, Nathan W. .

ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, 2022, 61 (22)

[20] Direct imaging of DNA in living cells reveals the dynamics of chromosome formation [J].

Manders, EMM ;

Kimura, H ;

Cook, PR .

JOURNAL OF CELL BIOLOGY, 1999, 144 (05) :813-821

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