1, 2, 4‐Triazine‐Modified 2′‐Deoxyuridine Triphosphate for Efficient Bioorthogonal Fluorescent Labeling of DNA. (7th June 2017)

• Record Type: Journal Article
• Authors: Peewasan, Krisana; Wagenknecht, Hans‐Achim
• Is Part Of: Chembiochem. Volume 18:Number 15(2017)
• Page Start: 1473
• Page End: 1476
• Publication Date: 2017-06-07
• Subjects: click chemistry -- Diels–Alder reaction -- DNA polymerase -- nucleic acids -- primer extension; Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pharmaceutical chemistry -- Periodicals; 572
• DOI: 10.1002/cbic.201700185
• Languages: English
• ISSNs: 1439-4227

Abstract

Abstract: In order to establish the Diels–Alder reaction with inverse electron demand for postsynthetic DNA modification, a 1, 2, 4‐triazine‐modified 2′‐deoxyuridine triphosphate was synthesized. The bioorthogonally reactive 1, 2, 4‐triazine group was attached at the 5‐position of 2′‐deoxyuridine by a flexible alkyl linker to facilitate its acceptance by DNA polymerases. The screening of four DNA polymerases showed successful primer extensions, using a mixture of dATP, dGTP, dCTP, and the modified 2′‐deoxyuridine triphosphate, by using KOD XL or Vent polymerase. The triazine moiety was stable under the conditions of primer extension, which was evidenced by labeling with a BCN‐modified rhodamine at room temperature in yields of up to 82 %. Two or three modified bases could be incorporated in quantitative yields when the modification sites were separated by three base pairs. These results establish the 1, 2, 4‐triazene group as a bioorthogonally reactive moiety in DNA, thereby replacing the problematic 1, 2, 4, 5‐tetrazine for postsynthetic labeling by the Diels–Alder reaction with inverse electron demand. Abstract : A better way to click : 1, 2, 4‐Triazene‐modified nucleotide triphosphates were synthesized and shown to be accepted as substrates by DNA polymerases. This new bioorthogonal modification could be an improvement over the problematic 1, 2, 4, 5‐tetrazines for postsynthetic fluorescent labeling of DNA by Diels–Alder reactions with inverse electron demand.