title: Real-Time Detection of a Self-Replicating RNA Enzyme.
abstract: A system was developed to detect the self-replication of an RNA enzyme in real time. The enzyme is an RNA ligase that undergoes exponential amplification at a constant temperature and can be made to operate in a ligand-dependent manner. The real-time system is based on a fluorimetric readout that directly couples the ligation event to an increase in florescence signal that can be monitored using standard instrumentation. The real-time system can also operate entirely with l-RNA, which is not susceptible to degradation by ribonucleases that are present in biological samples. The system is analogous to real-time PCR, but with the potential to detect small molecules, proteins, and other targets that can be recognized by a suitable aptamer. The ligand-dependent self-replication of RNA has potential applications in molecular diagnostics and biosensing that benefit from the rapid, precise, and real-time detection of various target molecules.
authors.affiliation: Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA. colea@scripps.edu.
authors.forename: Charles
authors.initials: C
authors.lastname: Olea
authors.affiliation: Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA. gjoyce@scripps.edu.
authors.forename: Gerald F
authors.initials: GF
authors.lastname: Joyce
mesh_terms: D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D011118:Polynucleotide Ligases / Q000737:chemistry*; D012313:RNA / Q000138:chemical synthesis* / Q000737:chemistry*; D016337:RNA, Catalytic / Q000737:chemistry*
chemical_list: D016337:RNA, Catalytic; D012313:RNA; D011118:Polynucleotide Ligases
keywords: RNA enzyme; RNA ligation; aptazyme; exponential amplification; fluorescence detection; in vitro selection; self-replication