Identifying significant variations in genomes can be cumbersome, as the variations span a multitude of base pairs and can make genome assembly difficult. However, large DNA molecules that span the variation aid in assembly. Due to the DNA molecule's large size, routine molecular biology techniques can break DNA.
View Article and Find Full Text PDFTo understand structural variation for personal genomics, an extensive ensemble of large DNA molecules will be required to span large structural variations. Nanocoding, a whole-genome analysis platform, can analyze large DNA molecules for the construction of physical restriction maps of entire genomes. However, handling of large DNA is difficult and a system is needed to concentrate large DNA molecules, while keeping the molecules intact.
View Article and Find Full Text PDFVisualization of DNA for fluorescence microscopy utilizes a variety of dyes such as cyanine dyes. These dyes are utilized due to their high affinity and sensitivity for DNA. In order to determine if the DNA molecules are full length after the completion of the experiment, a method is required to determine if the stained molecules are full length by digesting DNA with restriction enzymes.
View Article and Find Full Text PDFVery large DNA molecules enable comprehensive analysis of complex genomes, such as human, cancer, and plants because they span across sequence repeats and complex somatic events. When physically manipulated, or analyzed as single molecules, long polyelectrolytes are problematic because of mechanical considerations that include shear-mediated breakage, dealing with the massive size of these coils, or the length of stretched DNAs using common experimental techniques and fluidic devices. Accordingly, we harness analyte "issues" as exploitable advantages by our invention and characterization of the "molecular gate," which controls and synchronizes formation of stretched DNA molecules as DNA dumbbells within nanoslit geometries.
View Article and Find Full Text PDFNanocoding, a genome analysis platform, relies on very low ionic strength conditions to elongate DNA molecules up to 1.06 (fully stretched DNA = 1). Understanding how electroosmotic and electrophoretic forces vary, as ionic strength decreases, will enable better Nanocoding devices, or other genome analysis platforms, to be developed.
View Article and Find Full Text PDFNucleosides Nucleotides Nucleic Acids
June 2017
Optical mapping, a single DNA molecule genome analysis platform that can determine methylation profiles, uses fluorescently labeled DNA molecules that are elongated on the surface and digested with a restriction enzyme to produce a barcode of that molecule. Understanding how the cyanine fluorochromes affect enzyme activity can lead to other fluorochromes used in the optical mapping system. The effects of restriction digestion on fluorochrome labeled DNA (Ethidium Bromide, DAPI, H33258, EthD-1, TOTO-1) have been analyzed previously.
View Article and Find Full Text PDFThe analysis of very large DNA molecules intrinsically supports long-range, phased sequence information, but requires new approaches for their effective presentation as part of any genome analysis platform. Using a multi-pronged approach that marshaled molecular confinement, ionic environment, and DNA elastic properties-but tressed by molecular simulations-we have developed an efficient and scalable approach for presentation of large DNA molecules within nanoscale slits. Our approach relies on the formation of DNA dumbbells, where large segments of the molecules remain outside the nanoslits used to confine them.
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