Polymerase Synthesis of Base-Modified DNA

Part of the Nucleic Acids and Molecular Biology book series (NUCLEIC, volume 31)


Enzymatic synthesis of base-modified DNA by polymerase incorporation of modified nucleotides is discussed. Modified 2′-deoxyribonucleoside triphosphates (dNTPs) are key substrates for polymerases and can be prepared either by triphosphorylation of modified nucleosides or by direct aqueous cross-coupling reactions of halogenated dNTPs with alkynes, arylboronic acids, or alkenes. The methods of polymerase synthesis include primer extension, PCR, nicking enzyme amplifications, and other methods which enable the synthesis of diverse types of long or short and double-stranded DNA or single-stranded oligonucleotides. The applications include labeling in diagnostics (labeling or coding of DNA bases) and chemical biology (bioconjugations, modulation of protein binding, etc.).


Boronic Acid Heck Reaction Suzuki Reaction Arylboronic Acid Modify Nucleoside 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


  1. 1.
    Wilner OI, Wilner I (2012) Functionalized DNA nanostructures. Chem Rev 112:2528–2556. doi: 10.1021/cr200104q PubMedCrossRefGoogle Scholar
  2. 2.
    Clever GH, Kaul C, Carell T (2007) DNA-metal base pairs. Angew Chem Int Ed 46:6226–6236. doi: 10.1002/anie.200701185 CrossRefGoogle Scholar
  3. 3.
    Famulok M, Hartig JS, Mayer G (2007) Functional aptamers and aptazymes in biotechnology, diagnostics, and therapy. Chem Rev 107:3715–3743. doi: 10.1021/cr0306743 PubMedCrossRefGoogle Scholar
  4. 4.
    Torring T, Voigt NV, Nangreave J et al (2011) DNA origami: a quantum leap for self-assembly of complex structures. Chem Soc Rev 40:5636–5646. doi: 10.1039/c1cs15057j PubMedPubMedCentralCrossRefGoogle Scholar
  5. 5.
    Caruthers MH (2013) The chemical synthesis of DNA/RNA: our gift to science. J Biol Chem 288:1420–1427. doi: 10.1074/jbc.X112.442855 PubMedPubMedCentralCrossRefGoogle Scholar
  6. 6.
    Gierlich J, Burley GA, Gramlich PME et al (2006) Click chemistry as a reliable method for the high-density postsynthetic functionalization of alkyne-modified DNA. Org Lett 8:3639–3642. doi: 10.1021/ol0610946 PubMedCrossRefGoogle Scholar
  7. 7.
    Seela F, Sirivolu VR, Chittepu P (2008) Modification of DNA with octadiynyl side chains: synthesis, base pairing, and formation of fluorescent coumarin dye conjugates of four nucleobases by the alkyne-azide “click” reaction. Bioconjugate Chem 19:211–224. doi: 10.1021/bc700300f CrossRefGoogle Scholar
  8. 8.
    Shibata T, Glynn N, McMurry TBH et al (2006) Novel synthesis of O6-alkylguanine containing oligodeoxyribonucleotides as substrates for the human DNA repair protein, O6-methylguanine DNA methyltransferase (MGMT). Nucleic Acids Res 34:1884–1891. doi: 10.1093/nar/gkl117 PubMedPubMedCentralCrossRefGoogle Scholar
  9. 9.
    Hentschel S, Alzeer J, Angelov T et al (2012) Synthesis of DNA interstrand cross-links using a photocaged nucleobase. Angew Chem Int Ed 51:3466–3469. doi: 10.1002/anie.201108018 CrossRefGoogle Scholar
  10. 10.
    Arndt S, Wagenknecht H (2014) “Photoclick” postsynthetic modification of DNA. Angew Chem 53:14580–14582. doi: 10.1002/anie.201407874 CrossRefGoogle Scholar
  11. 11.
    Gramlich PME, Wirges CT, Manetto A, Carell T (2008) Postsynthetic DNA modification through the copper-catalyzed azide-alkyne cycloaddition reaction. Angew Chem Int Ed 47:8350–8358. doi: 10.1002/anie.200802077 CrossRefGoogle Scholar
  12. 12.
    El-Sagheer AH, Brown T (2010) Click chemistry with DNA. Chem Soc Rev 39:1388–1405. doi: 10.1039/b901971p PubMedCrossRefGoogle Scholar
  13. 13.
    Rieder U, Luedtke NW (2014) Alkene-tetrazine ligation for imaging cellular DNA. Angew Chem Int Ed 53:9168–9172. doi: 10.1002/anie.201403580 CrossRefGoogle Scholar
  14. 14.
    Langer PR, Waldrop AA, Ward DC (1981) Enzymatic synthesis of biotin labeled polynucleotides: novel nucleic acid affinity probes. Proc Natl Acad Sci U S A 78:6633–6637PubMedPubMedCentralCrossRefGoogle Scholar
  15. 15.
    Kuwahara M, Sugimoto N (2010) Molecular evolution of functional nucleic acids with chemical modifications. Molecules 15:5423–5444. doi: 10.3390/molecules15085423 PubMedCrossRefGoogle Scholar
  16. 16.
    Hollenstein M (2012) Nucleoside triphosphates-building blocks for the modification of nucleic acids. Molecules 17:13569–13591. doi: 10.3390/molecules171113569 PubMedCrossRefGoogle Scholar
  17. 17.
    Hocek M (2014) Synthesis of base-modified 2′-deoxyribonucleoside triphosphates and their use in enzymatic synthesis of modified DNA for applications in bioanalysis and chemical biology. J Org Chem 79:9914–9921. doi: 10.1021/jo5020799 PubMedCrossRefGoogle Scholar
  18. 18.
    Hirao I, Kimoto M (2012) Unnatural base pair systems toward the expansion of the genetic alphabet in the central dogma. Proc Jpn Acad Ser B 88:345–367. doi: 10.2183/pjab.88.345 CrossRefGoogle Scholar
  19. 19.
    Hocek M, Fojta M (2008) Cross-coupling reactions of nucleoside triphosphates followed by polymerase incorporation. Construction and applications of base-functionalized nucleic acids. Org Biomol Chem 6:2233–2241. doi: 10.1039/b803664k PubMedCrossRefGoogle Scholar
  20. 20.
    Burgess K, Cook D (2000) Syntheses of nucleoside triphosphates. Chem Rev 100:2047–2059. doi: 10.1021/cr990045m PubMedCrossRefGoogle Scholar
  21. 21.
    Yoshikawa M, Kato T, Takenishi T (1967) A novel method for phosphorylation of nucleosides to 5′-nucleotides. Tetrahedron Lett 8:5065–5068CrossRefGoogle Scholar
  22. 22.
    Gillerman I, Fischer B (2010) An improved one-pot synthesis of nucleoside 5′-triphosphate analogues. Nucleosides Nucleotides Nucleic Acids 29:245–256. doi: 10.1080/15257771003709569 PubMedCrossRefGoogle Scholar
  23. 23.
    Kuwahara M, Nagashima JI, Hasegawa M et al (2006) Systematic characterization of 2′-deoxynucleoside-5′-triphosphate analogs as substrates for DNA polymerases by polymerase chain reaction and kinetic studies on enzymatic production of modified DNA. Nucleic Acids Res 34:5383–5394. doi: 10.1093/nar/gkl637 PubMedPubMedCentralCrossRefGoogle Scholar
  24. 24.
    Sakthivel K, Barbas CF (1998) Expanding the potential of DNA for binding and catalysis: highly functionalized dUTP derivatives that are substrates for thermostable DNA polymerases. Angew Chem Int Ed 37:2872–2875CrossRefGoogle Scholar
  25. 25.
    Thum O, Jäger S, Famulok M (2001) Functionalized DNA: a new replicable biopolymer. Angew Chem Int Ed 40:3990–3993CrossRefGoogle Scholar
  26. 26.
    Jäger S, Rasched G, Kornreich-Leshem H et al (2005) A versatile toolbox for variable DNA functionalization at high density. J Am Chem Soc 127:15071–15082. doi: 10.1021/ja051725b PubMedCrossRefGoogle Scholar
  27. 27.
    Lee SE, Sidorov A, Gourlain T et al (2001) Enhancing the catalytic repertoire of nucleic acids: a systematic study of linker length and rigidity by two modified triphosphates during PCR. Nucleic Acids Res 29:1565–1573PubMedPubMedCentralCrossRefGoogle Scholar
  28. 28.
    Seela F, Feiling E, Gross J et al (2001) Fluorescent DNA: the development of 7-deazapurine nucleoside triphosphates applicable for sequencing at the single molecule level. J Biotechnol 86:269–279. doi: 10.1016/S0168-1656(00)00418-1 PubMedCrossRefGoogle Scholar
  29. 29.
    Roychowdhury A, Illangkoon H, Hendrickson CL, Benner SA (2004) 2′-deoxycytidines carrying amino and thiol functionality: synthesis and incorporation by vent (exo-)polymerase. Org Lett 6:489–492. doi: 10.1021/ol0360290 PubMedCrossRefGoogle Scholar
  30. 30.
    Cheng Y, Dai C, Peng H et al (2011) Design, synthesis, and polymerase-catalyzed incorporation of click-modified boronic acid-TTP analogues. Chem Asian J 6:2747–2752. doi: 10.1002/asia.201100229 PubMedCrossRefGoogle Scholar
  31. 31.
    Holzberger B, Marx A (2009) Enzymatic synthesis of perfluoroalkylated DNA. Bioorg Med Chem 17:3653–3658. doi: 10.1016/j.bmc.2009.03.063 PubMedCrossRefGoogle Scholar
  32. 32.
    Wang Y, Tkachenko BA, Schreiner PR, Marx A (2011) Diamondoid-modified DNA. Org Biomol Chem 9:7482. doi: 10.1039/c1ob05929g PubMedCrossRefGoogle Scholar
  33. 33.
    Fujita H, Nakajima K, Kasahara Y et al (2015) Polymerase-mediated high-density incorporation of amphiphilic functionalities into DNA: enhancement of nuclease resistance and stability in human serum. Bioorg Med Chem Lett 25:333–336. doi: 10.1016/j.bmcl.2014.11.037 PubMedCrossRefGoogle Scholar
  34. 34.
    Dziuba D, Pohl R, Hocek M (2015) Polymerase synthesis of DNA labelled with benzylidene cyanoacetamide-based fluorescent molecular rotors: fluorescent light-up probes for DNA-binding proteins. Chem Commun 51:4880–4882. doi: 10.1039/c5cc00530b CrossRefGoogle Scholar
  35. 35.
    Baccaro A, Marx A (2010) Enzymatic synthesis of organic-polymer-grafted DNA. Chem Eur J 16:218–226. doi: 10.1002/chem.200902296 PubMedCrossRefGoogle Scholar
  36. 36.
    Bußkamp H, Batroff E (2014) Efficient labelling of enzymatically synthesized vinyl-modified DNA by an inverse-electron-demand Diels–Alder reaction. Chem Commun 50:10827–10829. doi: 10.1039/c4cc04332d CrossRefGoogle Scholar
  37. 37.
    Kuwahara M, Hanawa K, Ohsawa K et al (2006) Direct PCR amplification of various modified DNAs having amino acids: convenient preparation of DNA libraries with high-potential activities for in vitro selection. Bioorg Med Chem 14:2518–2526. doi: 10.1016/j.bmc.2005.11.030 PubMedCrossRefGoogle Scholar
  38. 38.
    Baccaro A, Steck AL, Marx A (2012) Barcoded nucleotides. Angew Chem Int Ed 51:254–257. doi: 10.1002/anie.201105717 CrossRefGoogle Scholar
  39. 39.
    Verga D, Welter M, Steck AL, Marx A (2015) DNA polymerase-catalyzed incorporation of nucleotides modified with a G-quadruplex-derived DNAzyme. Chem Commun 51:7379–7381. doi: 10.1039/c5cc01387a CrossRefGoogle Scholar
  40. 40.
    Ludwig J, Eckstein F (1989) Rapid and efficient synthesis of nucleoside 5′-O-(1-thiotriphosphates), 5′-triphosphates and 2′,3′-cyclophosphorothioates using 2-chloro-4H-1,3,2-benzodioxaphosphorin-4-one. J Org Chem 54:631–635. doi: 10.1021/jo00264a024 CrossRefGoogle Scholar
  41. 41.
    Caton-Williams J, Lin L, Smith M, Huang Z (2011) Convenient synthesis of nucleoside 5′-triphosphates for RNA transcription. Chem Commun 47:8142–8144. doi: 10.1039/c1cc12201k CrossRefGoogle Scholar
  42. 42.
    Weizman H, Tor Y (2002) Redox-active metal-containing nucleotides: synthesis, tunability, and enzymatic incorporation into DNA. J Am Chem Soc 124:1568–1569. doi: 10.1021/ja017193q PubMedCrossRefGoogle Scholar
  43. 43.
    Hollenstein M (2013) Deoxynucleoside triphosphates bearing histamine, carboxylic acid, and hydroxyl residues—synthesis and biochemical characterization. Org Biomol Chem 11:5162–5172. doi: 10.1039/c3ob40842f PubMedCrossRefGoogle Scholar
  44. 44.
    Hollenstein M (2012) Synthesis of deoxynucleoside triphosphates that include proline, urea, or sulfonamide groups and their polymerase incorporation into DNA. Chem Eur J 18:13320–13330. doi: 10.1002/chem.201201662 PubMedCrossRefGoogle Scholar
  45. 45.
    Obeid S, Yulikov M, Jeschke G, Marx A (2008) Enzymatic synthesis of multiple spin-labeled DNA. Angew Chem Int Ed 47:6782–6785. doi: 10.1002/anie.200802314 CrossRefGoogle Scholar
  46. 46.
    Genet JP, Savignac M (1999) Recent developments of palladium(0) catalyzed reactions in aqueous medium. J Organomet Chem 576:305–317CrossRefGoogle Scholar
  47. 47.
    Shaughnessy KH (2009) Hydrophilic ligands and their application in aqueous-phase metal-catalyzed reactions. Chem Rev 109:643–710. doi: 10.1021/cr800403r PubMedCrossRefGoogle Scholar
  48. 48.
    Western EC, Daft JR, Johnson EM et al (2003) Efficient one-step Suzuki arylation of unprotected halonucleosides, using water-soluble palladium catalysts. J Org Chem 68:6767–6774. doi: 10.1021/jo034289p PubMedCrossRefGoogle Scholar
  49. 49.
    Thoresen LH, Jiao G-S, Haaland WC et al (2003) Rigid, conjugated, fluoresceinated thymidine triphosphates: syntheses and polymerase mediated incorporation into DNA analogues. Chem Eur J 9:4603–4610. doi: 10.1002/chem.200304944 PubMedCrossRefGoogle Scholar
  50. 50.
    Čapek P, Cahová H, Pohl R et al (2007) An efficient method for the construction of functionalized DNA bearing amino acid groups through cross-coupling reactions of nucleoside triphosphates followed by primer extension or PCR. Chem Eur J 13:6196–6203. doi: 10.1002/chem.200700220 PubMedCrossRefGoogle Scholar
  51. 51.
    Vrábel M, Horáková P, Pivoňková H et al (2009) Base-modified DNA labeled by [Ru(bpy)3]2+ and [Os(bpy)3]2+ complexes: construction by polymerase incorporation of modified nucleoside triphosphates, electrochemical and luminescent properties, and applications. Chem Eur J 15:1144–1154. doi: 10.1002/chem.200801538 PubMedCrossRefGoogle Scholar
  52. 52.
    Kielkowski P, Macíčková-Cahová H, Pohl R, Hocek M (2011) Transient and switchable (triethylsilyl)ethynyl protection of DNA against cleavage by restriction endonucleases. Angew Chem Int Ed 50:8727–8730. doi: 10.1002/anie.201102898 CrossRefGoogle Scholar
  53. 53.
    Riedl J, Ménová P, Pohl R et al (2012) GFP-like fluorophores as DNA labels for studying DNA-protein interactions. J Org Chem 77:8287–8293. doi: 10.1021/jo301684b PubMedCrossRefGoogle Scholar
  54. 54.
    Dadová J, Orság P, Pohl R et al (2013) Vinylsulfonamide and acrylamide modification of DNA for cross-linking with proteins. Angew Chem Int Ed 52:10515–10518. doi: 10.1002/anie.201303577 CrossRefGoogle Scholar
  55. 55.
    Balintová J, Pohl R, Horáková P et al (2011) Anthraquinone as a redox label for DNA: synthesis, enzymatic incorporation, and electrochemistry of anthraquinone-modified nucleosides, nucleotides, and DNA. Chem Eur J 17:14063–14073. doi: 10.1002/chem.201101883 PubMedCrossRefGoogle Scholar
  56. 56.
    Hervé G, Sartori G, Enderlin G et al (2014) Palladium-catalyzed Suzuki reaction in aqueous solvents applied to unprotected nucleosides and nucleotides. RSC Adv 4:18558–18594. doi: 10.1039/c3ra47911k CrossRefGoogle Scholar
  57. 57.
    Omumi A, Beach DG, Baker M et al (2011) Postsynthetic guanine arylation of DNA by Suzuki-Miyaura cross-coupling. J Am Chem Soc 133:42–50. doi: 10.1021/ja106158b PubMedCrossRefGoogle Scholar
  58. 58.
    Cahová H, Jäschke A (2013) Nucleoside-based diarylethene photoswitches and their facile incorporation into photoswitchable DNA. Angew Chem Int Ed 52:3186–3190. doi: 10.1002/anie.201209943 CrossRefGoogle Scholar
  59. 59.
    Lercher L, McGouran JF, Kessler BM et al (2013) DNA modification under mild conditions by Suzuki-Miyaura cross-coupling for the generation of functional probes. Angew Chem Int Ed 52:10553–10558. doi: 10.1002/ange.201304038 CrossRefGoogle Scholar
  60. 60.
    Čapek P, Pohl R, Hocek M (2006) Cross-coupling reactions of unprotected halopurine bases, nucleosides, nucleotides and nucleoside triphosphates with 4-borono-phenylalanine in water. Synthesis of (purin-8-yl)- and (purin-6-yl)phenylalanines. Org Biomol Chem 4:2278–2284. doi: 10.1039/b604010a PubMedCrossRefGoogle Scholar
  61. 61.
    Collier A, Wagner G (2006) A facile two-step synthesis of 8-arylated guanosine mono- and triphosphates (8-aryl GXPs). Org Biomol Chem 4:4526–4532. doi: 10.1039/b614477b PubMedCrossRefGoogle Scholar
  62. 62.
    Western EC, Shaughnessy KH (2005) Inhibitory effects of the guanine moiety on Suzuki couplings of unprotected halonucleosides in aqueous media. J Org Chem 70:6378–6388. doi: 10.1021/jo050832l PubMedCrossRefGoogle Scholar
  63. 63.
    Cahová H, Havran L, Brázdilová P et al (2008) Aminophenyl- and nitrophenyl-labeled nucleoside triphosphates: synthesis, enzymatic incorporation, and electrochemical detection. Angew Chem Int Ed 47:2059–2062. doi: 10.1002/anie.200705088 CrossRefGoogle Scholar
  64. 64.
    Horáková P, Macíčková-Cahová H, Pivoňková H et al (2011) Tail-labelling of DNA probes using modified deoxynucleotide triphosphates and terminal deoxynucleotidyl tranferase. Application in electrochemical DNA hybridization and protein-DNA binding assays. Org Biomol Chem 9:1366–1371. doi: 10.1039/c3cc41438h PubMedCrossRefGoogle Scholar
  65. 65.
    Ménová P, Cahová H, Plucnara M et al (2013) Polymerase synthesis of oligonucleotides containing a single chemically modified nucleobase for site-specific redox labelling. Chem Commun 49:4652–4654. doi: 10.1039/c3cc41438h CrossRefGoogle Scholar
  66. 66.
    Raindlová V, Pohl R, Sanda M, Hocek M (2010) Direct polymerase synthesis of reactive aldehyde-functionalized DNA and its conjugation and staining with hydrazines. Angew Chem Int Ed 49:1064–1066. doi: 10.1002/anie.200905556 CrossRefGoogle Scholar
  67. 67.
    Riedl J, Pohl R, Rulíšek L, Hocek M (2012) Synthesis and photophysical properties of biaryl-substituted nucleos(t)ides. Polymerase synthesis of DNA probes bearing solvatochromic and pH-sensitive dual fluorescent and 19F NMR labels. J Org Chem 77:1026–1044. doi: 10.1021/jo202321g PubMedCrossRefGoogle Scholar
  68. 68.
    Balintová J, Plucnara M, Vidláková P et al (2013) Benzofurazane as a new redox label for electrochemical detection of DNA: towards multipotential redox coding of DNA bases. Chem Eur J 19:12720–12731. doi: 10.1002/chem.201301868 PubMedCrossRefGoogle Scholar
  69. 69.
    Mačková M, Pohl R, Hocek M (2014) Polymerase synthesis of DNAs bearing vinyl groups in the major groove and their cleavage by restriction endonucleases. ChemBioChem 15:2306–2312. doi: 10.1002/cbic.201402319 PubMedCrossRefGoogle Scholar
  70. 70.
    Heck RF (1979) Palladium-catalyzed reactions of organic halides with olefins. Acc Chem Res 12:146–151CrossRefGoogle Scholar
  71. 71.
    Beletskaya IP, Cheprakov AV (2000) The Heck reaction as a sharpening stone of palladium catalysis. Chem Rev 100:3009–3066. doi: 10.1021/cr9903048 PubMedCrossRefGoogle Scholar
  72. 72.
    Agrofoglio LA, Gillaizeau I, Saito Y (2003) Palladium-assisted routes to nucleosides. Chem Rev 103:1875–1916. doi: 10.1021/cr010374q PubMedCrossRefGoogle Scholar
  73. 73.
    Tobrman T, Dvořák D (2008) Heck reactions of 6- and 2-halopurines. Eur J Org Chem 17:2923–2928. doi: 10.1002/ejoc.200800091 CrossRefGoogle Scholar
  74. 74.
    Kore AR, Shanmugasundaram M (2012) Highly stereoselective palladium-catalyzed Heck coupling of 5-iodouridine-5′-triphosphates with allylamine: a new efficient method for the synthesis of (E)-5-aminoallyl-uridine-5′-triphosphates. Tetrahedron Lett 53:2530–2532. doi: 10.1016/j.tetlet.2012.03.018 CrossRefGoogle Scholar
  75. 75.
    Dadova J, Pohl R, Fojta M, Hocek M (2013) Aqueous Heck cross-coupling preparation of acrylate-modified nucleotides and nucleoside triphosphates for polymerase synthesis of acrylate-labeled DNA. J Org Chem 78:9627–9637. doi: 10.1021/jo4011574 PubMedCrossRefGoogle Scholar
  76. 76.
    Cahová H, Pohl R, Bednárová L, Nováková K (2008) Synthesis of 8-bromo-, 8-methyl-and 8-phenyl-dATP and their polymerase incorporation into DNA. Org Biomol Chem 6:3657–3660. doi: 10.1039/B811935J PubMedCrossRefGoogle Scholar
  77. 77.
    Siegmund V, Diederichsen U, Marx A (2012) Enzymatic synthesis of 8-vinyl-and 8-styryl-2′-deoxyguanosine modified DNA—novel fluorescent molecular probes. Bioorg Med Chem Lett 22:3136–3139. doi: 10.1016/j.bmcl.2012.03.056 PubMedCrossRefGoogle Scholar
  78. 78.
    Pavlov YI, Minnick DT, Izuta S, Kunkel TA (1994) DNA replication fidelity with 8-oxodeoxyguanosine triphosphate. Biochemistry 33:4695–4701. doi: 10.1021/bi00181a029 PubMedCrossRefGoogle Scholar
  79. 79.
    Morgues S, Trzcionka J, Vasseur JJ, Pratviel G, Meunier B (2008) Incorporation of oxidized guanine nucleoside 5′-triphosphates in DNA with DNA polymerases and preparation of single-lesion carrying DNA. Biochemistry 47:4788–4799. doi: 10.1021/bi7022199 CrossRefGoogle Scholar
  80. 80.
    Kamath-Loeb AS, Hizi A, Kasai H, Loeb LA (1997) Incorporation of the guanosine triphosphate analogs 8-oxo-dGTP and 8-NH2-dGTP by reverse transcriptases and mammalian DNA polymerases. J Biol Chem 47:4788–4799. doi: 10.1074/jbc.272.9.5892 Google Scholar
  81. 81.
    Lam C, Hipolito C, Perrin DM (2008) Synthesis and enzymatic incorporation of modified deoxyadenosine triphosphates. Eur J Org Chem 4915–4923. doi: 10.1002/ejoc.200800381 Google Scholar
  82. 82.
    Hollenstein M, Hipolito CJ, Lam CH (2009) A self-cleaving DNA enzyme modified with amines, guanidines and imidazoles operates independently of divalent metal cations (M2+). Nucleic Acids Res 5:1638–1649. doi: 10.1093/nar/gkn1070 CrossRefGoogle Scholar
  83. 83.
    Hipolito CJ, Hollenstein M, Lam CH (2011) Protein-inspired modified DNAzymes: dramatic effects of shortening side-chain length of 8-imidazolyl modified deoxyadenosines in selecting RNaseA mimicking DNAzymes. Org Biomol Chem 9:2266–2273. doi: 10.1039/c1ob05359k PubMedCrossRefGoogle Scholar
  84. 84.
    Ito J, Braithwaite DK (1991) Compilation and alignment of DNA polymerase sequences. Nucleic Acids Res 19:4045–4049. doi: 10.1093/nar/19.15.4045 PubMedPubMedCentralCrossRefGoogle Scholar
  85. 85.
    Filée J, Forterre P, Sen-Lin T, Laurent J (2002) Evolution of DNA polymerase families: evidences for multiple gene exchange between cellular and viral proteins. J Mol Evol 54:763–773. doi: 10.1007/s00239-001-0078-x PubMedCrossRefGoogle Scholar
  86. 86.
    Steitz TA (1999) DNA polymerases: structural diversity and common mechanisms. J Biol Chem 274:17395–17398. doi: 10.1074/jbc.274.25.17395 PubMedCrossRefGoogle Scholar
  87. 87.
    Ramsay N, Jemth A-S, Brown A et al (2010) CyDNA: synthesis and replication of highly Cy-dye substituted DNA by an evolved polymerase. J Am Chem Soc 132:5096–5104. doi: 10.1021/ja909180c PubMedPubMedCentralCrossRefGoogle Scholar
  88. 88.
    Staiger N, Marx A (2010) A DNA polymerase with increased reactivity for ribonucleotides and C5‐modified deoxyribonucleotides. ChemBioChem 11:1963–1966. doi: 10.1002/cbic.201000384 PubMedCrossRefGoogle Scholar
  89. 89.
    Brázdilová P, Vrábel M, Pohl R, Pivoňková H et al (2007) Ferrocenylethynyl derivatives of nucleoside triphosphates: synthesis, incorporation, electrochemistry, and bioanalytical applications. Chem Eur J 13:9527–9533. doi: 10.1002/chem.200701249 PubMedCrossRefGoogle Scholar
  90. 90.
    Li H, Peng X, Seela F (2004) Fluorescence quenching of parallel-stranded DNA bound ethidium bromide: the effect of 7-deaza-2′-deoxyisoguanosine and 7-halogenated derivatives. Bioorg Med Chem Lett 14:6031–6034. doi: 10.1016/j.bmcl.2004.09.071 PubMedCrossRefGoogle Scholar
  91. 91.
    Seela F, Sirivolu VR, Chittepu P (2007) Modification of DNA with octadiynyl side chains: synthesis, base pairing, and formation of fluorescent coumarin dye conjugates of four nucleobases by the alkyne-azide “click” reaction. Bioconjugate Chem 19:211–224. doi: 10.1021/bc700300f CrossRefGoogle Scholar
  92. 92.
    Ménová P, Dziuba D, Güixens-Gallardo P et al (2015) Fluorescence quenching in oligonucleotides containing 7-substituted 7-deazaguanine bases prepared by the nicking enzyme amplification reaction. Bioconjugate Chem 26:361–366. doi: 10.1021/acs.bioconjchem.5b00006 CrossRefGoogle Scholar
  93. 93.
    Ménová P, Hocek M (2012) Preparation of short cytosine-modified oligonucleotides by nicking enzyme amplification reaction. Chem Commun 48:6921–6923. doi: 10.1039/C2CC32930A CrossRefGoogle Scholar
  94. 94.
    Ménová P, Raindlová V, Hocek M (2013) Scope and limitations of the nicking enzyme amplification reaction for the synthesis of base-modified oligonucleotides and primers for PCR. Bioconjugate Chem 24:1081–1093. doi: 10.1021/bc400149q CrossRefGoogle Scholar
  95. 95.
    Dziuba D, Pohl R, Hocek M (2014) Bodipy-labeled nucleoside triphosphates for polymerase synthesis of fluorescent DNA. Bioconjugate Chem 25:1984–1995. doi: 10.1021/bc5003554 CrossRefGoogle Scholar
  96. 96.
    Jäger S, Famulok M (2004) Generation and enzymatic amplification of high‐density functionalized DNA double strands. Angew Chem Int Ed 43:3337–3340. doi: 10.1002/anie.200453926 CrossRefGoogle Scholar
  97. 97.
    Kielkowski P, Fanfrlík J, Hocek M (2014) 7‐Aryl‐7‐deazaadenine 2′‐deoxyribonucleoside triphosphates (dNTPs): better substrates for DNA polymerases than dATP in competitive incorporations. Angew Chem Int Ed 53:7552–7555. doi: 10.1002/anie.201404742 CrossRefGoogle Scholar
  98. 98.
    Sinkeldam RW, Greco NJ, Tor Y (2010) Fluorescent analogs of biomolecular building blocks: design, properties, and applications. Chem Rev 110:2579–2619. doi: 10.1021/cr900301e PubMedPubMedCentralCrossRefGoogle Scholar
  99. 99.
    Tanpure AA, Pawar MG, Srivatsan SG (2013) Fluorescent nucleoside analogs: probes for investigating nucleic acid structure and function. Isr J Chem 53:366–378. doi: 10.1002/ijch.201300010 CrossRefGoogle Scholar
  100. 100.
    Palecek E, Bartosik M (2012) Electrochemistry of nucleic acids. Chem Rev 112:3427–3481. doi: 10.1021/cr200303p PubMedCrossRefGoogle Scholar
  101. 101.
    Balintová J, Špaček J, Pohl R, Brázdová M et al (2015) Azidophenyl as a click-transformable redox label of DNA suitable for electrochemical detection of DNA–protein interactions. Chem Sci 6:575–587. doi: 10.1039/C4SC01906G CrossRefGoogle Scholar
  102. 102.
    Prober JM, Trainor GL, Dam RJ, Hobbs FW et al (1987) A system for rapid DNA sequencing with fluorescent chain-terminating dideoxynucleotides. Science 238:336–341. doi: 10.1126/science.2443975 PubMedCrossRefGoogle Scholar
  103. 103.
    Turcatti G, Romieu A, Fedurco M, Tairi AP (2008) A new class of cleavable fluorescent nucleotides: synthesis and optimization as reversible terminators for DNA sequencing by synthesis. Nucleic Acids Res 36:e25. doi: 10.1093/nar/gkn021 PubMedPubMedCentralCrossRefGoogle Scholar
  104. 104.
    Riedl J, Pohl R, Ernsting NP et al (2012) Labelling of nucleosides and oligonucleotides by solvatochromic 4-aminophthalimide fluorophore for studying DNA–protein interactions. Chem Sci 3:2797–2806. doi: 10.1039/C2SC20404E CrossRefGoogle Scholar
  105. 105.
    Gramlich P, Warncke S, Gierlich J, Carell T (2008) Click–click–click: single to triple modification of DNA. Angew Chem Int Ed 47:3442–3444. doi: 10.1002/anie.200705664 CrossRefGoogle Scholar
  106. 106.
    Seela F, Sirivolu VR (2008) Pyrrolo-dC oligonucleotides bearing alkynyl side chains with terminal triple bonds: synthesis, base pairing and fluorescent dye conjugates prepared by the azide–alkyne “click” reaction. Org Biomol Chem 6:1674–1687. doi: 10.1039/b719459e PubMedCrossRefGoogle Scholar
  107. 107.
    Weisbrod SH, Marx A (2007) A nucleoside triphosphate for site-specific labelling of DNA by the Staudinger ligation. Chem Commun 1828–1830. doi: 10.1039/b809528k
  108. 108.
    Baccaro A, Weisbrod SH, Marx A (2007) DNA conjugation by the Staudinger Ligation: new thymidine analogues. Synthesis 1949–1954. doi: 10.1055/s-2007-983728 Google Scholar
  109. 109.
    Borsenberger V, Howorka S (2009) Diene-modified nucleotides for the Diels–Alder-mediated functional tagging of DNA. Nucleic Acids Res 37:1477–1485. doi: 10.1093/nar/gkn1066 PubMedPubMedCentralCrossRefGoogle Scholar
  110. 110.
    Schoch J, Wiessler M, Jäschke A (2010) Post-synthetic modification of DNA by inverse-electron-demand Diels–Alder reaction. J Am Chem Soc 132:8846–8847. doi: 10.1021/ja102871p PubMedCrossRefGoogle Scholar
  111. 111.
    Schoch J, Staudt M, Samanta A, Wiessler M, Jäschke A (2012) Site-specific one-pot dual labeling of DNA by orthogonal cycloaddition chemistry. Bioconjugate Chem 23:1382–1385. doi: 10.1021/bc300181n CrossRefGoogle Scholar
  112. 112.
    Raindlová V, Pohl R, Hocek M (2012) Synthesis of aldehyde‐linked nucleotides and DNA and their bioconjugations with lysine and peptides through reductive amination. Chem Eur J 18:4080–4087. doi: 10.1002/chem.201103270 PubMedCrossRefGoogle Scholar
  113. 113.
    Gourlain T, Sidorov A, Mignet N, Thorpe SJ et al (2001) Enhancing the catalytic repertoire of nucleic acids. II. Simultaneous incorporation of amino and imidazolyl functionalities by two modified triphosphates during PCR. Nucleic Acids Res 29:1898–1905. doi: 10.1093/nar/29.9.1898 PubMedPubMedCentralCrossRefGoogle Scholar
  114. 114.
    Imaizumi Y, Kasahara Y, Fujita H, Kitadume S et al (2013) Efficacy of base-modification on target binding of small molecule DNA aptamers. J Am Chem Soc 135:9412–9419. doi: 10.1021/ja4012222 PubMedCrossRefGoogle Scholar
  115. 115.
    Macíčková-Cahová H, Hocek M (2009) Cleavage of adenine-modified functionalized DNA by type II restriction endonucleases. Nucleic Acids Res 37:7612–7622. doi: 10.1093/nar/gkp845 PubMedPubMedCentralCrossRefGoogle Scholar
  116. 116.
    Macíčková‐Cahová H, Pohl R, Hocek M (2011) Cleavage of functionalized DNA containing 5‐modified pyrimidines by type II restriction endonucleases. ChemBioChem 12:431–438. doi: 10.1002/cbic.201000644 PubMedCrossRefGoogle Scholar
  117. 117.
    Mačková M, Boháčová S, Perlíková P et al (2015) Polymerase synthesis and restriction enzyme cleavage of DNA containing 7-substituted 7-deazaguanine. ChemBioChem 16:2225–2236. doi: 10.1002/cbic.201500315 PubMedCrossRefGoogle Scholar
  118. 118.
    Vaníková Z, Hocek M (2014) Polymerase synthesis of photocaged DNA resistant against cleavage by restriction endonucleases. Angew Chem Int Ed 53:6734–6737. doi: 10.1002/anie.201402370 CrossRefGoogle Scholar

Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  1. 1.Gilead Sciences & IOCB Research CenterInstitute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech RepublicPrague 6Czech Republic
  2. 2.Department of Organic Chemistry, Faculty of ScienceCharles University in PraguePrague 2Czech Republic

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