Young-Soo Seo 1 Vladimir A. Samuilov 1 Jonathan Sokolov 1 Miriam Rafailovich 1 Bernard Tinland 2 Jaeseung Kim 3 Benjamin Chu 3 1 Department of Materials Science and Engineering, SUNYat Stony Brook, NY, USA 2 Institut Charles Sadron, CNRS, Strasbourg, France 3 Department of Chemistry, SUNYat Stony Brook, NY, USA DNA separation at a liquid-solid interface We demonstrate that it is possible to separate a broad band of DNA on a solid sub- strate without topological obstacles. The mobility was found to scale with molecular size (N) as N –0.25 , while the resolution scaled as N 0.75 indicating that diffusivity on this substrate was minimal. By varying the buffer concentration we were able to show that the mobility for a given chain length scaled with the persistent length (p) as p 1/2 . This could be shown to be related to the Gaussian conformation of the chains adsorbed on the surface. A two-dimensional corrugated surface of nonporous silica beads was produced using a self-assembling process at the air/water interface. Even though the surface corrugations were comparable to persistence length we show that they do not affect the mobility, indicating that surface friction rather than topological constraints are the predominant mechanism of separation on a surface. Keywords: DNA separation / Liquid-solid interface/ Self-assembled silica beads / Silicon sub- strate EL 5070 1 Introduction The separation of DNA molecules in a wide molecular weight range is required in many applications of molecu- lar biology. DNA fragments cannot be separated under an applied electric field in simple solutions since the electro- phoretic mobility of a DNA molecule is related to its charge-to-friction ratio, a quantity that is independent of chain length. Separation of DNA therefore requires the use of some type of sieving matrix [1] such as those produced by topological constrictions, such as junction points in a gel, entanglements in a polymer solution, or porous arrays of particles [1, 2]. The most common meth- ods used for separating long double-stranded (ds)DNA molecules are pulsed-field slab-gel electrophoresis (PFGE) or pulsed-field capillary gel electrophoresis (PFCGE) [3, 4]. Since the gels used in these techniques can be very complex and susceptible to degradation with use, new methods for DNA separation that rely on topological contstraints produced by inorganic corru- gated matrices have been proposed. For example, Tin- land [5] has shown that separation can be achieved using a highly symmetric lattice with well-defined porosity which is produced by “crystallization” of silica beads within a cell. Ordered two-dimensional arrays of posts such as those produced lithographically on Si wafers by Austin and co-workers [6] and microfabricated channel arrays produced by Craighead and co-workers [7], have demonstrated that dsDNA can also be separated on sur- faces by introducing well defined topological constraints. In the first case, the topological constraints provide obstacles to electrodiffusion by “hooking” molecules in a manner similar to three-dimensional electrophoresis [8]. In the second case, mobility is hindered by “entropic trapping” where the DNA strands of different lengths are separated by their escape times from the channel [9]. In all these instances where rigid, inorganic topographical sieving matrices are introduced, the effect on DNA chain mobility due to interactions with the surface must also be considered. In particular when the degree of confinement is large, such as in the microfabricated arrays [9] where the constriction is less than the radius of gyration of the DNA chains, the relative importance of surface interac- tions versus entropic pinning must be evaluated in order to quantitatively interpret the results. We have recently shown that it was possible to obtain separation on a flat Si wafer without any topographical constraints [10]. When DNA molecules are adsorbed on a “flat” surface, the balance between the loss of entropy due to the localization of the DNA at the surface and the energetic gain on adsorption of the molecule results in the classic picture of DNA segments being present as either loops (that extend into the buffer solution) or trains (that are contiguous segments adsorbed on the surface) [11]. For a fixed amplitude of the attraction potential, longer DNA molecules have more adsorbed trains (contiguous segments) which implies that they experience more fric- tion than the short ones for motion in the plane of the sub- strate. This introduces a length-dependent mobility and, as a result, separation. Correspondence: Professor Vladimir Samuilov, Department of Materials Science and Engineering, SUNY at Stony Brook, NY 11794, USA E-mail: vladimir.samuilov@sunysb.edu Fax: 1631-632-5764 Abbreviations: FWHM, full width at half maximum; PMT , photo- multiplier tube; TBE, Tris-boric acid-EDTA buffer 2618 Electrophoresis 2002, 23, 2618–2625  2002 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 0173-0835/02/1608–2618 $17.501.50/0