Fig. 6. Alternative biocatalytic method for the production of modified nucleic acids.

This isothermal biocatalytic approach based on the polymerase-mediated incorporation of modified nucleotides into a self-priming hairpin template (gray–blue). (i) Template-dependent synthesis using sugar and phosphate-modified nucleotides and the KOD polymerase; (ii) endonuclease V cleaves the second phosphodiester linkage 3’-downstream of a deoxyinosine moiety present in the template and generates the modified sequence (in red); (iii) after endonuclease-cleavage, the template is available for subsequent rounds of catalytic synthesis.