期刊

ACS Synthetic Biology

标题

Two-Layered Microfluidic Devices for High-Throughput Dynamic Analysis of Synthetic Gene Circuits in E. coli

作者

Yanhong Sun, Fengyu Zhang, Lusi Li, Kaiyue Chen, Shujing Wang, Qi Ouyang, and Chunxiong Luo

摘要

Escherichia coli is a common chassis for synthetic gene circuit studies. In addition to the dose–response of synthetic gene circuits, the analysis of dynamic responses is also an important part of the future design of more complicated synthetic systems. Recently, microfluidic-based methods have been widely used for the analysis of gene expression dynamics. Here, we established a two-layered microfluidic platform for the systematic characterization of synthetic gene circuits (eight strains in eight different culture environments could be observed simultaneously with a 5 min time resolution). With this platform, both dose responses and dynamic responses with a high temporal resolution could be easily derived for further analysis. A controlled environment ensures the stability of the bacterial growth rate, excluding changes in gene expression dynamics caused by changes of the growth dilution rate. The precise environmental switch and automatic micrograph shooting ensured that there was nearly no time lag between the inducer addition and the data recording. We studied four four-node incoherent-feedforward-loop (IFFL) networks with different operators using this device. The experimental results showed that as the effect of inhibition increased, two of the IFFL networks generated pulselike dynamic gene expressions in the range of the inducer concentrations, which was different from the dynamics of the two other circuits with only a simple pattern of rising to the platform. Through fitting the dose–response curves and the dynamic response curves, corresponding parameters were derived and introduced to a simple model that could qualitatively explain the generation of pulse dynamics.

原文链接

https://pubs.acs.org/doi/10.1021/acssynbio.2c00307