Localized Nanoscopic Surface Measurements of Nickel-Modified Mica for Single-Molecule DNA Sequence Sampling

Abstract

Cleaved, cation-derivatized Muscovite mica is utilized extensively in AFM imaging due to its flatness over large areas (millimeter cleavage planes with local RMS roughness <0.3 nm), ease of preparation and ability to adsorb charged biomolecules such as DNA1–3. In particular, NiCl2 treatment has become a common method for controlling DNA adsorption on mica substrates while retaining the mica’s ultra flat surface4. While several studies have modeled the mica:metal-ion:DNA system using macroscopic colloidal theory (DLVO, etc) 4–8, Ni-mica’s physicochemical properties have not been well characterized on the nanoscale. Efforts to manipulate and engineer DNA nanostructures would benefit greatly from a better understanding of the surface chemistry of Ni:mica. Here we present in-situ, nanometer- and attogram-scale measurements, and thermodynamic simulation results that show the surface chemistry of nickel treated mica is more complex than generally appreciated by AFM practitioners, due to metal ion speciation effects present at neutral pH. We also show that under certain preparations, Ni:Mica allows in situ, nanoscopic nucleotide sequence mapping within individual surface-adsorbed DNA molecules by permitting localized, controlled desorption of the double helix by soluble DNA binding enzymes. These results should aid efforts to precisely control DNA:mica binding affinity, particularly at physiological pH ranges required by enzymatic biochemistry (pH 7.0–8.5)4–8, and facilitate development of more complex and useful biochemical manipulations of adsorbed DNA, such as single molecule sequencing.

Tuning the translational freedom of DNA for high speed AFM

“…This behavior is more prominent with molecules imaged with Ni 2+ concentrations of 5 mM or lower. Although a direct correlation with surface distributions of Ni 2+ would be purely speculative without further in depth studies, the results together with previous findings on the distribution of Ni 2+ ions [30,40] suggest that positively charged Ni 2+ domains act as anchor points for the negatively charged DNA chains, with the interstitial regions governed by weak Mg 2+ associations, allowing for the desired mobility.…”

“…To consider the role of Ni 2+ within our system, we adopted the idealized polka dot surface model discussed by others [30,32]. It is reasonable to assume that the density of surface-associated Ni 2+ ions increases with increasing Ni 2+ concentration during incubation.…”

“…It is reasonable to assume that the density of surface-associated Ni 2+ ions increases with increasing Ni 2+ concentration during incubation. It has previously been demonstrated that the distribution of Ni 2+ on the surface is highly non-uniform [40] and lacks homogeneity [30]. However, it may be possible to gain further insight into the Ni 2+ /Mg 2+ -governed surface interactions by identifying regions of specific surface association by examining the motion of the DNA chains.…”

“…These alternative methods either put significant constraints on the experimental setup or do not provide the required functionality, and therefore pre-incubation of mica with divalent cations remains the common practice. [22,30], leading to phase-separated domains of positive surface charge [31]. Where transition metal cations are utilized, a strong adhesion between DNA and mica is achieved due to directional bonding of the d-orbitals of the divalent cation.…”

Ion Exchange and DNA Molecular Dip Sticks: Studying the Nanoscale Surface Wetting of Muscovite Mica

“…We have demonstrated that in addition to ion species, ion concentration, and pH of the solution 8,18,37 , the time-dependent incubation and aging process of the sample also play important roles in determining the ion distribution on mica surfaces. When the sample was treated with Ni 2+ solution for a shorter period (30 seconds), the tip-sample interaction underwent a long time-scale dependent transition related to the accumulation of water on the mica surface and possibly as a consequence of surface compositional change and spatial distribution.…”