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. 2014 Oct 24;53(47):12735–12740. doi: 10.1002/anie.201405991

Figure 2.

Figure 2

Agarose-gel-electrophoretic characterization of PEG purification using exemplarily a 42-helix bundle object. a) Image of a gel on which samples extracted after each of ten consecutive PEG purification cycles (1–10) were separated. Labels: sc, reference sample containing only scaffold strands; U, unpurified self-assembly reaction mixture; po, gel loading pocket; m, folded objects; ex, non-integrated excess staple strands. b) Cross-sectional lane profiles determined from (a). c) Recovery of folded objects (left) and residuals of excess staple strands (right) relative to unpurified reaction mixture, as determined by integrating and comparing the areas of the peaks reflecting folded objects and excess strands, respectively. The experiment was run in triplicate, each experiment gave data as in (a). Error bars in (c) indicate the standard deviation in the recovery and residuals, respectively. d) Image of a gel containing samples extracted after one and five cycles of PEG purification (1xP, 5xP), after one and five cycles of molecular-weight cut-off filtration (1xF, 5xF), after AGE extraction (1xG), and after PEG purification of a previously AGE-extracted sample (G+P). U refers to the unpurified self-assembly reaction mixture. Other labels as in (a). e) Cross-sectional lane profiles determined from (d). f) Recovery of folded objects and residual excess strands as in (c) but for samples as in (d). Values were obtained from three independent experiments, each giving data as in (d). Error bars indicate the standard deviation in the recovery and residuals, respectively. g–i) Estimation of residual PEG content in purified samples using fluorescein-labeled PEG (fPEG). g) Overlay image of two scans of the same gel, recorded separately for the ethidium bromide and fluorescein emission channels. Samples were taken before (U) and after (U+PB) addition of precipitation buffer (PB) and compared to the supernatant (SN) and the redissolved pellet (P) of the precipitation. h) Cross-sectional lane profiles from ethidium bromide channel (dark gray) and fluorescein channel (light gray). i) Estimated concentrations of folded DNA objects (dsDNA) and PEG at all steps of a PEG purification cycle. Labels: fPEG, fluorescein-labeled PEG; other labels as in (a).