Rapidly characterizing the fast dynamics of RNA genetic circuitry with cell-free transcription-translation (TX-TL) systems
Rapidly characterizing the fast dynamics of RNA genetic circuitry with cell-free transcription-translation (TX-TL) systems
Abstract 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 timescales 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 response 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.
Chappell James、Fall Christopher P.、Murray Richard M.、Lucks Julius B.、Spring Kevin J.、Takahashi Melissa K.、Al-Khabouri Shaima、Singhal Vipul、Sun Zachary Z.、Hayes Clarmyra A.、Noireaux Vincent
School of Chemical and Biomolecular Engineering, Cornell UniversityCSHL Course in Synthetic Biology, Cold Spring Harbor Laboratory||Department of BioEngineering, University of Illinois||Department of Computer Science, Georgetown UniversityCSHL Course in Synthetic Biology, Cold Spring Harbor Laboratory||Division of Biology and Biological Engineering, California Institute of TechnologySchool of Chemical and Biomolecular Engineering, Cornell University||CSHL Course in Synthetic Biology, Cold Spring Harbor LaboratoryCSHL Course in Synthetic Biology, Cold Spring Harbor Laboratory||Department of Integrative Biology and PharmacologySchool of Chemical and Biomolecular Engineering, Cornell University||CSHL Course in Synthetic Biology, Cold Spring Harbor LaboratoryCSHL Course in Synthetic Biology, Cold Spring Harbor Laboratory||Institute for Research in Immunology and Cancer (IRIC)CSHL Course in Synthetic Biology, Cold Spring Harbor Laboratory||Division of Biology and Biological Engineering, California Institute of TechnologyDivision of Biology and Biological Engineering, California Institute of TechnologyDivision of Biology and Biological Engineering, California Institute of TechnologySchool of Physics and Astronomy, University of Minnesota
生物科学研究方法、生物科学研究技术生物工程学分子生物学
RNA genetic circuitsTX-TLcell-freeresponse timeRNA synthetic biology
Chappell James,Fall Christopher P.,Murray Richard M.,Lucks Julius B.,Spring Kevin J.,Takahashi Melissa K.,Al-Khabouri Shaima,Singhal Vipul,Sun Zachary Z.,Hayes Clarmyra A.,Noireaux Vincent.Rapidly characterizing the fast dynamics of RNA genetic circuitry with cell-free transcription-translation (TX-TL) systems[EB/OL].(2025-03-28)[2025-05-06].https://www.biorxiv.org/content/10.1101/003335.点此复制
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