Reprogramming Microbes for the Remote Detection of Environmental Threats
Abstract
Riboswitches are sequences of RNA that control gene expression via RNA-ligand interactions, without the need for accessory proteins. Riboswitches consist of an aptamer that recognizes the ligand and an expression platform that couples ligand binding to a change in gene expression. Using in vitro selection, it is possible to screen large (~10^13 member) libraries of RNA sequences to discover new aptamers. However, limitations in bacterial transformation efficiency make screening such large libraries for riboswitch function in intact cells impractical. We established that an E. coli S30 extract that couples transcription/translation is sufficient to recapitulate the function of riboswitches expressed in intact cells. We established that a family of synthetic riboswitches acts through a kinetic trapping mechanism, rather than an equilibrium mechanism. Finally, we established CIS-Select as an in vitro system to perform selections for novel synthetic riboswitches. We anticipate that with optimization, CIS-Select will enable direct selection for riboswitch activity from very large RNA libraries, while bypassing the limitations of bacterial transformation efficiency.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 15, 2013
- Accession Number
- ADA587112
Entities
People
- Justin Gallivan
Organizations
- Emory University