title: Synthesis and enzymatic incorporation of photolabile dUTP analogues into DNA and their applications for DNA labeling.
abstract: Two novel photolabile nucleotide triphosphate (NTP) analogues were synthesized through Sonogashira coupling and their enzymatic incorporation into DNA was evaluated with three different DNA polymerases (Taq, Vent exo- and T4) by polymerase chain reaction. Both nucleotide triphosphate analogues were recognized by these DNA polymerases as substrates for primer extension. Light irradiation of PCR products removed the photolabile group and released the amino and carboxyl moieties. Further site-specific dual-labeling for oligodeoxynucleotides (ODNs) and random labeling for a long DNA construct with fluorophores were successfully achieved with incorporation of the photolabile amine modified deoxyuridine triphosphate (dUnTP).
authors.affiliation: State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, No. 38 Xueyuan Rd., Beijing 100191, China.
authors.forename: Junzhou
authors.initials: J
authors.lastname: Wu
authors.forename: Xinjing
authors.initials: X
authors.lastname: Tang
mesh_terms: D004247:DNA / Q000737:chemistry*; D003856:Deoxyuracil Nucleotides / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D055668:Photochemical Processes
chemical_list: D003856:Deoxyuracil Nucleotides; D005456:Fluorescent Dyes; C027078:deoxyuridine triphosphate; D004247:DNA
keywords: Enzymatic incorporation; Fluorescence labeling; Nucleotide analogues; Photoactivation; Photolabile nucleotides