Figure 3. Guiding the refinement of self-assembly protocols by quality feedback.

(a) Laser-scanned false-coloured photographs of agarose gels on which a screen of self-assembly reaction mixtures for isothermal folding of a 42-helix-bundle DNA origami object was electrophoresed. Reaction mixtures were incubated for 2 h at the indicated temperatures (after a brief scaffold-denaturing heatshock to 65 °C). Top and bottom images give object and defect label fluorescence intensities, respectively. P indicates the gel pocket and F, the target object band. (b) Relative defect label brightness as a function of incubation temperature. (c,d) As in a,b, but for a time-resolved analysis of a 42-helix-bundle self-assembly reaction mixture incubated at constant 52 °C. The electrophoretic mobility of the target object band displays a slight ondulation that stems from the apparatus used. R1: stepwise ‘annealing’ from 60 to 44 C° with a cooling rate of 1 °C per hour. R2: 30 min at 52 °C; 30 min at 45 °C; and 30 min at 25 °C. Note that the relative brightness values in c,d should not be compared directly because the data sets were obtained from two different gels. Blue squares give data obtained for a design variant of the 42-helix bundle that shows best constant-temperature folding at 48 °C. See also Supplementary Fig. 35.