Constructing an NF-κB Oscillator for Dual-Input Control in Yeast

Interactions between biological oscillators and environmental periodic signals are crucial for the correct regulation of cellular behaviors. Most synthetic biological oscillators only respond to one signal that initiates the oscillation, limiting the study of the interaction between the biological oscillator and multiple environmental cues. Here, we present a detailed protocol for constructing a synthetic oscillatory circuit that can respond to both α-factor and ethanol inputs. Based on the mammalian NF-κB negative feedback loop, this circuit produces sustained and tunable oscillation in the budding yeast and can be operated by dual periodic inputs.