Title: In vitro selection systems for TNA
Description: The origins of life on Earth and the basis for its characteristics are unclear. Based on genetic and biochemical evidence, it is believed that RNA emerged before protein and was initially a dual-functioning genetic and catalytic molecule of life in a time called the RNA world. Experiments modeling the synthesis of RNA under prebiotic settings indicate that RNA may have been difficult to make and accumulate. Furthermore, they indicate that other related molecules probably existed and some may have been easier to make than RNA. Therefore, a simpler molecule that accumulated more readily than RNA may have existed first and eventually facilitated the emergence of RNA. We have chosen to test this hypothesis. Threose Nucleic Acid (TNA) is structural analog of the natural nucleic acids containing a threose instead of a ribose sugar. TNA is a uniquely strong candidate to have preceded RNA because its synthesis may have been simpler than that of RNA and it is capable of base pairing with itself and RNA, meaning it can function genetically and even transfer genetic information to RNA. For TNA to have preceded RNA as the central molecule of life, it would have also had to have been capable of ligand-binding and catalysis. We have decided to address this question by developing an in vitro selection system to test the functional potential of TNA.

We have identified and characterized protein polymerases that perform efficient, long, and faithful DNA-dependent TNA polymerization and TNA reverse-transcription. These enzymes are active enough to enable TNA in vitro selection. We have also developed a selection system that does not require a TNA reverse-transcriptase that involves covalently linking each TNA library member to the DNA template that encodes it. Use of these systems will enable us to determine the functional potential of TNA and evaluate the possible role of simplified nucleic acids in the origin of life. We also discuss an important concept involving the fidelity and replication characteristics of primitive replicases that may have eased the pressure of error catastrophe in the origin of life.