Fig. 3
mEos2-TFs exhibit two dissociation rate constants. a Scheme of time-lapse microscopy. Red spheres indicate a detected mEos2-TF molecule. τon : time laser is on, τoff : time laser is off, τtl : time-lapse time. b Schematic decay of visible fluorescence over time under short (blue), intermediate (magenta) and long (red) time-lapse intervals indicated by the horizontal lines. Inset: Measured mEos2-TBP molecules still bound after 4 short (left), 2 intermediate (middle) and 2 long (right) time-lapse intervals demonstrating effects of photobleaching (left) and dissociation (right). Starting number were 100 molecules. The scale bar is 5 µm. c, d Histograms of fluorescent “on” times at different time-lapse conditions of c mEos2-TBP and d mEos2-Sox19b in nuclei of oblong embryos (mean ± s.d.). Time-lapse intervals are written on top of the data points. Lines: global fit of a bi-exponential decay model (Equation 1 in Methods). Data includes 4780 molecules from 15 embryos for mEos2-TBP and 3330 molecules from 4 embryos for mEos2-Sox19b. For clarity, only the first six points of histograms are shown. e, f Residence times of e mEos2-TBP and f mEos2-Sox19b. Errors are the s.d. from the fits in c and d
