In vitro selection of an allosteric ribozyme that transduces analytes to amplicons

Abstract

We have selected an allosteric ribozyme ligase from a random sequence population that is activated up to 10,000-fold by oligonucleotide effectors. The ribozyme conforms to a classic two-state model for allostery in which the equilibrium between inactive and active conformers is dramatically altered by the presence of effector ligands. In the presence of the effector the allosteric ribozyme ligase generates templates that can subsequently be amplified using conventional amplification technologies, such as RT-PCR. Thus, the allosteric ribozyme can transduce (or convert) analytes into amplicons. We demonstrate two potential diagnostic applications of the selected allosteric ribozyme ligase: 'counting' short oligonucleotide effectors by RT-PCR, and counting a non-nucleic acid effector, ATP, by ligation.