Single molecule DNA intercalation in continuous homogenous elongational flow
| Autor: | Sean J. Gerrin, Gene Malkin, Griffis Joshua, Robert H. Meltzer, Ekaterina Protozanova, Shilpi P. Vyas, Mikhail M. Safranovitch |
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| Rok vydání: | 2014 |
| Předmět: |
Benzoxazoles
Microfluidics Intercalation (chemistry) Biomedical Engineering Bioengineering Nanotechnology General Chemistry DNA Microfluidic Analytical Techniques Biochemistry Fluorescence Elasticity chemistry.chemical_compound DNA Intercalation chemistry Optical mapping Biophysics Molecule A-DNA Fluorescent Dyes |
| Zdroj: | Lab on a chip. 14(19) |
| ISSN: | 1473-0189 |
| Popis: | Sequence-nonspecific staining of DNA with intercalating fluorophores is required for fluorescence-based length estimation of elongated DNA in optical mapping techniques. However, the observed length of a DNA molecule is affected by the relative concentrations of DNA and dye. In some applications, predetermination of DNA concentration may not be possible. Here we present a microfluidic approach in which individual DNA molecules are entrained by converging laminar sheath flows containing the intercalating dye PO-PRO-1. This provides uniform staining regardless of DNA concentration, and uniform elastic stretching of DNA in continuous elongational flow. On-chip intercalation provides a unique process for concentration-independent staining of long DNA fragments for the optical mapping method Genome Sequence Scanning (GSS), and normalizes intramolecular elasticity across a broad range of molecule lengths. These advances permit accurate mapping of observed molecules to sequence derived templates, thus improving detection of complex bacterial mixtures using GSS. |
| Databáze: | OpenAIRE |
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