Ligase-catalyzed transcription and reverse-transcription of XNA-containing nucleic acid polymers using T3 DNA ligase

A method to enable the transliteration between various XNA-containing nucleic acids and canonical DNA is described. Using ligase-catalysed oligonucleotide polymerisation (LOOPER), we show that DNA can be used as a template to generate nucleic acids polymers comprising various levels of 2'-fluoro (2'-F), 2'-fluoro-arabinonucleic acid (FANA), 2'-<i>O</i>-methyl (2'-OMe), and Locked Nucleic Acids (LNA) in moderate yields. The fidelity and biases of the LOOPER process were studied in detail for the 2'-F system by developing a hairpin-based sequencing method, which showed fidelities exceeding 95% along with positional and sequence dependencies within the polymerised XNA-containing anticodons. Lastly, we show the ability of LOOPER to regenerate DNA from 2'-F, FANA, 2'-OMe, and LNA in moderate yield and in fidelities over 95%. Taken together, this study demonstrates the potential of LOOPER to serve as a platform for applications where the transliteration between XNA and DNA is needed, such as the <i>in vitro</i> evolution of XNA-containing nucleic acid polymers.