Journal
ACS SYNTHETIC BIOLOGY
Volume 4, Issue 5, Pages 503-515Publisher
AMER CHEMICAL SOC
DOI: 10.1021/sb400206c
Keywords
RNA genetic circuits; TX-TL; cell-free; response time; RNA synthetic biology
Categories
Funding
- National Science Foundation Graduate Research Fellowship Program [DGE-1144153]
- Defense Advanced Research Projects Agency Young Faculty Award (DARPA YFA) [N66001-12-1-4254]
- Office of Naval Research Young Investigators Program Award (ONR YIP) [N00014-13-1-0531]
- Defense Advanced Research Projects Agency (DARPA/MTO) Living Foundries program [HR0011-12-C-0065]
- Howard Hughes Medical Institute
- Office of Naval Research
- Div Of Chem, Bioeng, Env, & Transp Sys
- Directorate For Engineering [1402843] Funding Source: National Science Foundation
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RNA regulators are emerging as powerful tools to engineer synthetic genetic networks or rewire existing ones. A potential strength of RNA networks is that they may be able to propagate signals on time scales that are set by the fast degradation rates of RNAs. However, a current bottleneck to verifying this potential is the slow design-build-test cycle of evaluating these networks in vivo. Here, we adapt an Escherichia coli-based cell-free transcription-translation (TX-TL) system for rapidly prototyping RNA networks. We used this system to measure the response time of an RNA transcription cascade to be approximately five minutes per step of the cascade. We also show that this response time can be adjusted with temperature and regulator threshold tuning. Finally, we use TX-TL to prototype a new RNA network, an RNA single input module, and show that this network temporally stages the expression of two genes in vivo.
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