Enzymatic synthesis of hypermodified DNA polymers for sequence-specific display of four different hydrophobic groups

0537186 - ÚOCHB 2021 RIV GB eng J - Journal Article
Ondruš, Marek - Sýkorová, Veronika - Bednárová, Lucie - Pohl, Radek - Hocek, Michal
Enzymatic synthesis of hypermodified DNA polymers for sequence-specific display of four different hydrophobic groups.
Nucleic Acids Research. Roč. 48, č. 21 (2020), s. 11982-11993. ISSN 0305-1048. E-ISSN 1362-4962
R&D Projects: GA ČR(CZ) GX20-00885X
Grant - others:AV ČR(CZ) AP1501
Program: Akademická prémie - Praemium Academiae
Institutional support: RVO:61388963
Keywords : nucleotides * DNA * functionalized polymers
OECD category: Biochemistry and molecular biology
Impact factor: 16.971, year: 2020
Method of publishing: Open access
https://doi.org/10.1093/nar/gkaa999

A set of modified 2'-deoxyribonucleoside triphosphates (dNTPs) bearing a linear or branched alkane, indole or phenyl group linked through ethynyl or alkyl spacer were synthesized and used as substrates for polymerase synthesis of hypermodified DNA by primer extension (PEX). Using the alkyl-linked dNTPs, the polymerase synthesized up to 22-mer fully modified oligonucleotide (ON), whereas using the ethynyl-linked dNTPs, the enzyme was able to synthesize even long sequences of >100 modified nucleotides in a row. In PCR, the combinations of all four modified dNTPs showed only linear amplification. Asymmetric PCR or PEX with separation or digestion of the template strand can be used for synthesis of hypermodified single-stranded ONs, which are monodispersed polymers displaying four different substituents on DNA backbone in sequence-specific manner. The fully modified ONs hybridized with complementary strands and modified DNA duplexes were found to exist in B-type conformation (B- or C-DNA) according to CD spectral analysis. The modified DNA can be replicated with high fidelity to natural DNA through PCR and sequenced. Therefore, this approach has a promising potential in generation and selection of hypermodified aptamers and other functional polymers.
Permanent Link: http://hdl.handle.net/11104/0316289