Enzyme-Free Replication with Two or Four Bases.

All known forms of life encode their genetic information in a sequence of bases of a genetic polymer and produce copies through replication. How this process started before polymerase enzymes had evolved is unclear. Enzyme-free copying of short stretches of DNA or RNA has been demonstrated using activated nucleotides, but not replication. We have developed a method for enzyme-free replication. It involves extension with reversible termination, enzyme-free ligation, and strand capture. We monitored nucleotide incorporation for a full helical turn of DNA, during both a first and a second round of copying, by using mass spectrometry. With all four bases (A/C/G/T), an "error catastrophe" occurred, with the correct sequence being "overwhelmed" by incorrect ones. When only C and G were used, approximately half of the daughter strands had the mass of the correct sequence after 20 copying steps. We conclude that enzyme-free replication is more likely to be successful with just the two strongly pairing bases than with all four bases of the genetic alphabet.