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Unravelling DNA Organization with Single-Molecule Force Spectroscopy Using Magnetic Tweezers.
Brouwer, Thomas B; Kaczmarczyk, Artur; Zarguit, Ilias; Pham, Chi; Dame, Remus T; van Noort, John.
Afiliación
  • Brouwer TB; Leiden Institute of Physics, Leiden University, Leiden, The Netherlands.
  • Kaczmarczyk A; Leiden Institute of Physics, Leiden University, Leiden, The Netherlands.
  • Zarguit I; Leiden Institute of Chemistry, Leiden University, Leiden, The Netherlands.
  • Pham C; Leiden Institute of Physics, Leiden University, Leiden, The Netherlands.
  • Dame RT; Leiden Institute of Chemistry, Leiden University, Leiden, The Netherlands.
  • van Noort J; Centre for Interdisciplinary Genome Research, Leiden University, Leiden, The Netherlands.
Methods Mol Biol ; 2819: 535-572, 2024.
Article en En | MEDLINE | ID: mdl-39028523
ABSTRACT
Genomes carry the genetic blueprint of all living organisms. Their organization requires strong condensation as well as carefully regulated accessibility to specific genes for proper functioning of their hosts. The study of the structure and dynamics of the proteins that organize the genome has benefited tremendously from the development of single-molecule force spectroscopy techniques that allow for real-time, nanometer accuracy measurements of the compaction of DNA and manipulation with pico-Newton scale forces. Magnetic tweezers, in particular, have the unique ability to complement such force spectroscopy with the control over the linking number of the DNA molecule, which plays an important role when DNA-organizing proteins form or release wraps, loops, and bends in DNA. Here, we describe all the necessary steps to prepare DNA substrates for magnetic tweezers experiments, assemble flow cells, tether DNA to a magnetic bead inside a flow cell, and manipulate and record the extension of such DNA tethers. Furthermore, we explain how mechanical parameters of nucleoprotein filaments can be extracted from the data.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: ADN / Imagen Individual de Molécula Idioma: En Revista: Methods Mol Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2024 Tipo del documento: Article País de afiliación: Países Bajos Pais de publicación: Estados Unidos
Texto completo
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XML
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: ADN / Imagen Individual de Molécula Idioma: En Revista: Methods Mol Biol Asunto de la revista: BIOLOGIA MOLECULAR Año: 2024 Tipo del documento: Article País de afiliación: Países Bajos Pais de publicación: Estados Unidos