Accuracy of AFM measurements of the contour length of DNA fragments adsorbed on mica in air and in aqueous buffer

The measurement by atomic force microscope of the contour length of DNA fragments adsorbed on mica has been made as accurate as possible by revisiting the different steps of image acquisition and processing. In air, the DNA helical rise was estimated at 2.97 +/- 0.15 Angstrom per base pair (bp) (mean +/- standard deviation) by imaging a 648-bp DNA fragment and 2.95 +/- 0.14 Angstrom per bp for a 1115-bp fragment. This confirms earlier observations suggesting that drying DNA fragments on mica in the presence of nickel induces limited conformational changes. At this point the exact nature of these conformational changes remains unknown. Simple hypotheses are the transconformation of stretches of the DNA molecules to the A-form of the double helix or alteration of the helix structure at the points of contact between DNA and mica. By contrast, in aqueous buffer, the measured helical rise was 3.14 +/- 0.15 Angstrom per bp for the 648bp fragment and 3.17 +/- 0.13 Angstrom per bp for the 1115-bp fragment. Thus, measured helical rises do not depend on the fragment length and are significantly shorter than the 3.38 Angstrom per bp measured by crystallography, but close to the 3.18 Angstrom per bp found in NMR studies. These findings are discussed with respect to discrepancies in earlier results published in the literature. (C) 2002 Elsevier Science B.V. All rights reserved.