Niederholtmeyer Henrike, Sun Zachary Z, Hori Yutaka, Yeung Enoch, Verpoorte Amanda, Murray Richard M, Maerkl Sebastian J
Institute of Bioengineering, School of Engineering, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States.
Elife. 2015 Oct 5;4:e09771. doi: 10.7554/eLife.09771.
While complex dynamic biological networks control gene expression in all living organisms, the forward engineering of comparable synthetic networks remains challenging. The current paradigm of characterizing synthetic networks in cells results in lengthy design-build-test cycles, minimal data collection, and poor quantitative characterization. Cell-free systems are appealing alternative environments, but it remains questionable whether biological networks behave similarly in cell-free systems and in cells. We characterized in a cell-free system the 'repressilator', a three-node synthetic oscillator. We then engineered novel three, four, and five-gene ring architectures, from characterization of circuit components to rapid analysis of complete networks. When implemented in cells, our novel 3-node networks produced population-wide oscillations and 95% of 5-node oscillator cells oscillated for up to 72 hr. Oscillation periods in cells matched the cell-free system results for all networks tested. An alternate forward engineering paradigm using cell-free systems can thus accurately capture cellular behavior.
尽管复杂的动态生物网络控制着所有生物体中的基因表达，但可比的合成网络的正向工程仍然具有挑战性。目前在细胞中表征合成网络的模式导致设计-构建-测试周期漫长、数据收集极少且定量表征不佳。无细胞系统是有吸引力的替代环境，但生物网络在无细胞系统和细胞中的行为是否相似仍值得怀疑。我们在无细胞系统中表征了“抑制振荡电路”，这是一种三节点合成振荡器。然后，我们设计了新颖的三基因、四基因和五基因环形架构，从电路组件的表征到完整网络的快速分析。当在细胞中实施时，我们新颖的三节点网络产生了全群体振荡，并且95%的五节点振荡器细胞振荡长达72小时。细胞中的振荡周期与所有测试网络的无细胞系统结果相匹配。因此，使用无细胞系统的另一种正向工程模式可以准确地捕捉细胞行为。
