Superhelix dimensions of a 1868 base pair plasmid determined by scanning force microscopy in air and in aqueous solution.
We have used scanning force microscopy (SFM) to study the conformation of a 1868 base pair plasmid (p1868) in its open circular form and at a superhelical density of sigma= -0.034. The samples were deposited on a mica surface in the presence of MgCl2. DNA images were obtained both in air and in aqueous solutions, and the dimensions of the DNA superhelix were analysed. Evaluation of the whole plasmid yielded average superhelix dimensions of 27 +/- 9 nm (outer superhelix diameter D), 107 +/- 51 nm (superhelix pitch P), and 54 +/-8 degrees (superhelix pitch angle alpha). We also analysed compact superhelical regions within the plasmid separately, and determined values of D = 9.2 +/- 3.3 nm, P = 42 +/- 13 nm and alpha= 63 +/- 20 degrees for samples scanned in air or rehydrated in water. These results indicate relatively large conformation changes between superhelical and more open regions of the plasmid. In addition to the analysis of the DNA superhelix dimensions, we have followed the deposition process of open circular p1868 to mica in real time. These experiments show that it is possible to image DNA samples by SFM without prior drying, and that the surface bound DNA molecules retain some ability to change their position on the surface.
Rippe
K
K
Deutsches Krebsforschungszentrum, Abteilung Biophysik der Makromoleküle, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany. Karsten.Rippe@dkfz-heidelberg.de
Mücke
N
N
Langowski
J
J
D000388:Air; D001482:Base Composition; D004274:DNA, Recombinant / Q000737:chemistry*; D018625:Microscopy, Atomic Force; D010957:Plasmids / Q000737:chemistry*; D012996:Solutions; D014867:Water
D004274:DNA, Recombinant; D012996:Solutions; D014867:Water