Template-directed synthesis of complementary strands is pivotal for life. Nature employs polymerases for this reaction, leaving the ability of DNA itself to direct the incorporation of individual nucleotides at the end of a growing primer difficult to assess. Using 64 sequences, we now find that any of the four nucleobases, in combination with any neighboring residue, support enzyme-free primer extension when primer and mononucleotide are sufficiently reactive, with ≥93% primer extension for all sequences. Between the 64 possible base triplets, the rate of extension for the poorest template, CAG, with A as templating base, and the most efficient template, TCT, with C as templating base, differs by less than two orders of magnitude. Further, primer extension with a balanced mixture of monomers shows ≥72% of the correct extension product in all cases, and ≥90% incorporation of the correct base for 46 out of 64 triplets in the presence of a downstream-binding strand. A mechanism is proposed with a binding equilibrium for the monomer, deprotonation of the primer, and two chemical steps, the first of which is most strongly modulated by the sequence. Overall, rates show a surprisingly smooth reactivity landscape, with similar incorporation on strongly and weakly templating sequences. These results help to clarify the substrate contribution to copying, as found in polymerase-catalyzed replication, and show an important feature of DNA as genetic material.

Rapid replication: Non‐enzymatic primer extension has previously been studied in the context of prebiotic chemistry, but not for practical applications. Reactions with primers featuring a 3′‐amino‐2′,3′‐dideoxynucleotide can be rapid and selective for all four templating nucleobases (see scheme). On a chip with immobilized capture strands, 500 fmol of template suffice for single‐nucleotide determinations within 2.7 h.

Rasche Replikationsschritte: Die nichtenzymatische Primerverlängerung wurde bislang nur bei Fragen der präbiotischen Chemie, nicht aber für praktische Anwendungen eingesetzt. Reaktionen mit Primern, die ein 3′‐Amino‐2′,3′‐didesoxynucleotid enthalten, können für alle vier als Template dienenden Nucleobasen schnell und selektiv sein (siehe Schema). Auf einem Chip mit immobilisierten DNA‐Sonden genügen 500 fmol Templat für Einzelnucleotidbestimmungen innerhalb von 2.7 h.

Rapid extention: Catalysis by pyridine accelerates template‐directed chemical primer extension to t1/2<1 min for all four nucleotides (A/C/G/T). For C and G, greater than 99 % conversion is observed after 30 s (see picture). These sequence‐selective reactions open up new avenues for the interrogation of DNA sequences and chemical replication.

Rascher Einbau: Eine Pyridinkatalyse beschleunigt die templatgerichtete chemische Primerverlängerung mit allen vier Nucleotiden (A/C/G/T) auf t1/2<1 min. Mit C und G wird >99 % Umsatz nach 30 s beobachtet (siehe Bild). Diese sequenzselektiven Reaktionen bieten neue Möglichkeiten für die DNA‐Sequenzanalyse und die chemische Replikation.