Atomic force microscopy of supercoiled DNA structure on mica

Abstract

Atomic force microscopy (AFM) has been used to investigate supercoiled DNA structure. In AFM imaging, the deformation of DNA structure in the adsorption process onto the mica surface is a serious problem. To investigate the structure of supercoiled DNA in solution by AFM, the method by which DNA is fixed on the substrate without any deformation of the structure should be developed. In this paper, it is made clear that the structure of supercoiled DNA on mica is dependent on the method of treatment of the mica substrate. We prepared four kinds of mica surfaces; which were (1) no treatment (freshly cleaved) mica, treated with (2) spermine, (3) spermidine and (4) 3-aminopropyltriethoxysilane. The values of roughness of the mica treated with spermine and spermidine were 0.18 and 0.19 nm, respectively, while that of the mica treated with 3-aminopropyltriethoxy silane was 0.38 nm. Supercoiled DNA molecules (pUC19) were fixed in the shape of open circles on the freshly cleaved mica surface, whereas supercoiled DNA molecules on the mica treated with spermine or spermidine interwound tightly. We could, moreover, observe the clockwise handedness of the supercoiled DNA with AFM using the mica treated with spermine as substrate. The apparent length of DNA in the supercoil was shorter than that in open circular DNA. It was made clear by analysis of the differences of these measured lengths that the superhelix winding angle and the value of writhe of pUC19 were 61° and 10.2, respectively.