Fig. 4.

Investigating the dynamics of SCRaMbLE. a The endpoint 520 nm fluorescence values of SCRaMbLEd synV-pBAB016 colonies picked randomly after 2, 4, 8 or 24 h of β-estradiol induction and then grown in 96-well plate format for 24 h. For each induction length, ≥81 randomly selected induced colonies were characterised and 6 colonies were also characterised from an uninduced synV-pBAB016 control plate. Fluorescence values were normalised for culture OD₆₀₀ and are given as a proportion of the mean un-SCRaMbLEd control fluorescence value for that induction length. Blue dots denote un-SCRaMbLEd controls (cont), grey dots denote SCRaMbLEd cells (SCR) that have a fluorescence value equal to or lower than the highest un-SCRaMbLEd control, green dots denote SCRaMbLEd cells that have a higher fluorescence values than any of the un-SCRaMbLEd controls and purple dots denote the relative VB2-pBAB016 fluorescence value from Fig. 1e. Percentage values above the plotted data are the proportion of SCRaMbLEd cells that have a higher fluorescence value than any of the un-SCRaMbLEd controls and values below the plotted data give the proportion of SCRaMbLEd cells with a lower fluorescence value than those controls. Black horizontal lines denote the mean average relative fluorescence of that group of cells, excluding the VB2-pBAB016 value in the case of the controls. b The equivalent information but with SCRaMbLEd colonies visually screened under blue light to selectively pick colonies with high fluorescence. For each induction length, ≥19 selected colonies were characterised along with 3 un-SCRaMbLEd controls. c For each induction length, the proportion of cell death and the proportion of improved cultures, i.e., those with higher fluorescence output than any of the equivalent un-SCraMbLEd controls, with either selective or random picking of colonies. Lines are second-degree polynomial curves of best fit, with R² values stated. d OD₆₀₀ achieved after 48 h growth in synthetic xylose medium of XD4-pJCH006, synV-pJCH006 and the 13 fastest growing re-SCRaMbLEd XD4-pJCH006 colonies. Values plotted are mean averages of 3 biological replicates and error bars represent 1 standard deviation from the mean