Single Cell Microbioreactor for observing Dynamic Gene Expression and Cell Response to Environment in Bacteria

Abstract

Biological systems have been shown to implement a low-pass filter in order to distinguish high frequency noise from a lower frequency input signal, which is essential to a cellular system to adapt or sustain fitness in fluctuating environment conditions. Gene expression has been shown to exhibit noise leading to phenotypic heterogeneity of a cell population. Many microfluidic techniques have been developed in order to study single cell inducible gene expression, however rely solely on diffusion timescales to effectively alternate between inducer concentrations. In our approach, we developed a microfluidic platform for single cell analysis that allows for dynamic control of a target cell in oscillating well-defined culture media and inducer concentrations. The hydrodynamic trap enables confinement and manipulation of single cells in free solution using the sole action of fluid flow. Automated feedback control is integrated into the device using an “on-chip” valve, which allows for precise confinement of cells in free solution. During observation, cells are confined at a fluid stagnation point generated by planar extensional flow in a cross-slot microfluidic geometry, thereby enabling non-perturbative trapping of cells for long time scales. Using this platform, we investigated the effect of small molecule inducers on gene expression in the lac operon using fluorescent reporter proteins and cell growth rates as a proxy of cellular fitness. We applied our technique to determine the cut-off frequency associated with periodic stimuli for the lac circuit in E. coli, wherein the cut-off frequency in the case of the low-pass filter is the higher frequency when response begins to attenuate. We observed that single cell gene expression depends on the correlation between growth rate and frequency of exposure to inducer concentrations.