The design and implementation of synthetic circuits that operate robustly in the cellular context is fundamental for the advancement of synthetic biology. However, their practical implementation presents challenges due to low predictability of synthetic circuit design and time-intensive troubleshooting. Here, we present the Cyberloop, a testing framework to accelerate the design process and implementation of biomolecular controllers. Cellular fluorescence measurements are sent in real-time to a computer simulating candidate stochastic controllers, which in turn compute the control inputs and feed them back to the controlled cells via light stimulation. Applying this framework to yeast cells engineered with optogenetic tools, we examine and characterize different biomolecular controllers, test the impact of non-ideal circuit behaviors such as dilution on their operation, and qualitatively demonstrate improvements in controller function with certain network modifications. From this analysis, we derive conditions for desirable biomolecular controller performance, thereby avoiding pitfalls during its biological implementation.

在细胞环境中稳定运行的合成电路的设计和实现是合成生物学进步的基础。然而，由于合成电路设计的可预测性低和故障排除耗时，它们的实际实施面临着挑战。在这里，我们介绍了 Cyber​​loop，这是一个加速生物分子控制器的设计过程和实现的测试框架。细胞荧光测量结果被实时发送到模拟候选随机控制器的计算机，计算机依次计算控制输入并通过光刺激将其反馈给受控细胞。将该框架应用于用光遗传学工具工程化的酵母细胞，我们检查和表征不同的生物分子控制器，测试非理想电路行为（例如稀释）对其操作的影响，并定性地证明通过某些网络修改对控制器功能的改进。从这一分析中，我们得出了理想的生物分子控制器性能的条件，从而避免了其生物实施过程中的陷阱。