Probing concentration-dependent behavior of DNA-binding proteins on a single-molecule level illustrated by Rad51

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Frykholm, Karolin, 1977 (author): Gothenburg University,Göteborgs universitet,Institutionen för fysik (GU),Department of Physics (GU),University of Gothenburg

Freitag, Camilla (author): Gothenburg University,Göteborgs universitet,Institutionen för fysik (GU),Department of Physics (GU),University of Gothenburg

Persson, Fredrik, 1979 (author): Uppsala universitet,Gothenburg University,Göteborgs universitet,Institutionen för fysik (GU),Department of Physics (GU),University of Gothenburg,Beräknings- och systembiologi

Tegenfeldt, Jonas O. (author): Lund University,Lunds universitet,Gothenburg University,Göteborgs universitet,Institutionen för fysik (GU),Department of Physics (GU),Fasta tillståndets fysik,Fysiska institutionen,Institutioner vid LTH,Lunds Tekniska Högskola,Solid State Physics,Department of Physics,Departments at LTH,Faculty of Engineering, LTH,University of Gothenburg

Graneli, Annette, 1973 (author): Gothenburg University,Göteborgs universitet,Institutionen för fysik (GU),Department of Physics (GU),University of Gothenburg

(creator_code:org_t)

Elsevier BV, 2013
2013
English.
In: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 443:2, s. 261-268

Related links:: http://dx.doi.org/10...; show more...; https://gup.ub.gu.se...; https://doi.org/10.1...; https://lup.lub.lu.s...; https://research.cha...; https://urn.kb.se/re...; show less...

Abstract Subject headings

Low throughput is an inherent problem associated with most single-molecule biophysical techniques. We have developed a versatile tool for high-throughput analysis of DNA and DNA-binding molecules by combining microfluidic and dense DNA arrays. We use an easy-to-process microfluidic flow channel system in which dense DNA arrays are prepared for simultaneous imaging of large amounts of DNA molecules with single-molecule resolution. The Y-shaped microfluidic design, where the two inlet channels can be controlled separately and precisely, enables the creation of a concentration gradient across the microfluidic channel as well as rapid and repeated addition and removal of substances from the measurement region. A DNA array stained with the fluorescent DNA-binding dye YOYO-1 in a gradient manner illustrates the method and serves as a proof of concept. We have applied the method to studies of the repair protein Rad51 and could directly probe the concentration-dependent DNA-binding behavior of human Rad51 (HsRad51). In the low-concentration regime used (100 nM HsRad51 and below), we detected binding to double-stranded DNA (dsDNA) without positive cooperativity. (C) 2013 Elsevier Inc. All rights reserved.

NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
NATURVETENSKAP -- Fysik -- Den kondenserade materiens fysik (hsv//swe)
NATURAL SCIENCES -- Physical Sciences -- Condensed Matter Physics (hsv//eng)

By the author/editor: Frykholm, Karoli ...; Freitag, Camilla; Persson, Fredrik ...; Tegenfeldt, Jona ...; Graneli, Annette ...

About the subject

NATURAL SCIENCES: NATURAL SCIENCES; and Biological Scien ...; and Biochemistry and ...

NATURAL SCIENCES: NATURAL SCIENCES; and Physical Science ...; and Condensed Matter ...

By the university: University of Gothenburg; Lund University; Chalmers University of Technology; Uppsala University

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