Role of a non-natural beta-C-nucleotide unit in DNA as a template for DNA and RNA syntheses and as a substrate for nucleolytic digestion.
A non-natural beta-C-nucleoside bearing a 3,4-dibenzyloxyphenyl group as a nucleobase (X) was synthesized and incorporated into a 34-mer oligomer with the sequence 5'-dTTTTTAAAAAAXATATAGCAGCGACATGTCACCG-3'. This synthetic oligonucleotide was examined for template activity in the enzymatic syntheses of DNA by the Klenow fragments of Escherichia coli DNA polymerase I and the recombinant DNA polymerase I, and in the synthesis of RNA by the E. coli RNA polymerase core enzyme. As a result, the template-directed polymerization of both DNA and RNA was precisely terminated at the position of X. The X-containing oligonucleotide was also tested for digestion by an exonuclease, Exo III nuclease (Exo III), and an endonuclease, Mung Bean nuclease (MB). The results indicate that the artificial nucleobase X acts as a terminator for digestion by Exo III, whereas the site X becomes susceptible to digestion by MB. These findings provide a useful tool for the size control of products in the synthesis and degradation of nucleic acids.
D001483:Base Sequence; D004247:DNA / Q000138:chemical synthesis*; D004256:DNA Polymerase I / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry; D005090:Exodeoxyribonucleases / Q000737:chemistry; D009841:Oligonucleotides / Q000737:chemistry*; D012313:RNA / Q000138:chemical synthesis*; D012318:RNA Polymerase I / Q000737:chemistry; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000737:chemistry
D017931:DNA Primers; D009841:Oligonucleotides; D012313:RNA; D004247:DNA; D012318:RNA Polymerase I; D004256:DNA Polymerase I; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III; D015719:Single-Strand Specific DNA and RNA Endonucleases