Figure 1. An instantaneous transcription rate is calculated from single-cell fluorescence time series.

(A) In the 3-color diploid strain, tet-Trans-Activator (tTA) expressed from P_MYO2 activates P_7xtetO and is repressed by doxycycline. Homologous copies of P_7xtetO drive expression of either CFP or YFP while P_PGK1 drives RFP expression constitutively. (B) Monitoring growth of single cells in microfluidic culture. The segmented mother cell, daughter, and contiguous whole cell regions are outlined in blue, green, and red, respectively. For the mother and its daughters, (C) raw volume and (D) CFP concentration time series were smoothed to remove measurement noise; (E) integrated CFP fluorescence was calculated as their product (corresponding regions shown in B). The sum of bud and mother values until division constitutes a whole cell trace, which when extended past division, yields a running total trace that is fit to a differentiable smoothing spline. (F) Calculated relative mRNA levels and (G) instantaneous transcription rate.