| Autor: |
Henley RY; Department of Physics, Northeastern University, Boston, Massachusetts, United States of America., Vazquez-Pagan AG; Department of Biology, Northeastern University, Boston, Massachusetts, United States of America., Johnson M; Department of Chemistry and Chemical Biology, Northeastern University, Boston, Massachusetts, United States of America., Kanavarioti A; Yenos Analytical LLC, El Dorado Hills, California, United States of America., Wanunu M; Department of Physics, Northeastern University, Boston, Massachusetts, United States of America.; Department of Chemistry and Chemical Biology, Northeastern University, Boston, Massachusetts, United States of America. |
| Abstrakt: |
Nanopores are a promising platform in next generation DNA sequencing. In this platform, an individual DNA strand is threaded into nanopore using an electric field, and enzyme-based ratcheting is used to move the strand through the detector. During this process the residual ion current through the pore is measured, which exhibits unique levels for different base combinations inside the pore. While this approach has shown great promise, accuracy is not optimal because the four bases are chemically comparable to one another, leading to small differences in current obstruction. Nucleobase-specific chemical tagging can be a viable approach to enhancing the contrast between different bases in the sequence. Herein we show that covalent modification of one or both of the pyrimidine bases by an osmium bipyridine complex leads to measureable differences in the blockade amplitudes of DNA molecules. We qualitatively determine the degree of osmylation of a DNA strand by passing it through a solid-state nanopore, and are thus able to gauge T and C base content. In addition, we show that osmium bipyridine reacts with dsDNA, leading to substantially different current blockade levels than exhibited for bare dsDNA. This work serves as a proof of principle for nanopore sequencing and mapping via base-specific DNA osmylation. |
