title: Molecular Requirements of High-Fidelity Replication-Competent DNA Backbones for Orthogonal Chemical Ligation.
abstract: The molecular properties of the phosphodiester backbone that made it the evolutionary choice for the enzymatic replication of genetic information are not well understood. To address this, and to develop new chemical ligation strategies for assembly of biocompatible modified DNA, we have synthesized oligonucleotides containing several structurally and electronically varied artificial linkages. This has yielded a new highly promising ligation method based on amide backbone formation that is chemically orthogonal to CuAAC "click" ligation. A study of kinetics and fidelity of replication through these artificial linkages by primer extension, PCR, and deep sequencing reveals that a subtle interplay between backbone flexibility, steric factors, and ability to hydrogen bond to the polymerase modulates rapid and accurate information decoding. Even minor phosphorothioate modifications can impair the copying process, yet some radical triazole and amide DNA backbones perform surprisingly well, indicating that the phosphate group is not essential. These findings have implications in the field of synthetic biology.
authors.affiliation: Department of Chemistry, Chemistry Research Laboratory, University of Oxford , 12 Mansfield Road, Oxford OX1 3TA, U.K.
authors.forename: Arun
authors.initials: A
authors.lastname: Shivalingam
authors.affiliation: Department of Chemistry, Chemistry Research Laboratory, University of Oxford , 12 Mansfield Road, Oxford OX1 3TA, U.K.
authors.forename: Agnes E S
authors.initials: AE
authors.lastname: Tyburn
authors.affiliation: Department of Chemistry, Chemistry Research Laboratory, University of Oxford , 12 Mansfield Road, Oxford OX1 3TA, U.K.|Chemistry Branch, Department of Science and Mathematics, Faculty of Petroleum and Mining Engineering, Suez University , Suez 43721, Egypt.
authors.forename: Afaf H
authors.initials: AH
authors.lastname: El-Sagheer
authors.affiliation: Department of Chemistry, Chemistry Research Laboratory, University of Oxford , 12 Mansfield Road, Oxford OX1 3TA, U.K.
authors.forename: Tom
authors.identifier: 0000-0002-6538-3036
authors.initials: T
authors.lastname: Brown
mesh_terms: D004247:DNA / Q000737:chemistry* / Q000235:genetics; D004261:DNA Replication; D004583:Electrons; D004952:Esters / Q000737:chemistry*; D007700:Kinetics; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D014230:Triazoles / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D004952:Esters; D009841:Oligonucleotides; D014230:Triazoles; D004247:DNA