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© Structural Dynamics Of Macromolecules
The structure of a bacterial analog of the nicotinic receptor (one color per subunit) inserted into the cell membrane (grey and orange). A representation of the volume accessible to ions is shown in yellow.
Publication : Methods (San Diego, Calif.)

Rapid enzymatic synthesis of long RNA polymers: A simple protocol to generate RNA libraries with random sequences

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Methods (San Diego, Calif.) - 26 Mar 2019

Randrianjatovo-Gbalou I, Delarue M

Link to Pubmed [PMID] – 30926532

Methods 2019 May;161:83-90

RNA aptamers have several advantages over DNA aptamers due to their propensity to fold into three-dimensional structures. However, the synthesis of large RNA libraries remains a challenge as it requires more precautions to conserve their functional integrity, especially when such libraries are intended for aptamers or ribozymes selection. Here, we present an enzymatic method that enables the rapid synthesis of RNA polymers thanks to the efficient incorporation of ribonucleotides (NTPs) as well as chemically modified ribonucleotides by human DNA polymerase Theta (θ) mutants. These mutants have the ability to generate long single-stranded RNA polynucleotides of random sequences due to their improved template-free terminal nucleotidyltransferase activity. Here we describe the detailed protocols to produce large and diverse libraries of RNA, to make them ready to use in repeated cycles of Systematic Evolution of Ligands by Exponential enrichment (SELEX) and to synthesize C2′-modified nucleic acids with higher nuclease resistance.

https://www.ncbi.nlm.nih.gov/pubmed/30926532