title: Electrophoretic mobility of supercoiled, catenated and knotted DNA molecules.
abstract: We systematically varied conditions of two-dimensional (2D) agarose gel electrophoresis to optimize separation of DNA topoisomers that differ either by the extent of knotting, the extent of catenation or the extent of supercoiling. To this aim we compared electrophoretic behavior of three different families of DNA topoisomers: (i) supercoiled DNA molecules, where supercoiling covered the range extending from covalently closed relaxed up to naturally supercoiled DNA molecules; (ii) postreplicative catenanes with catenation number increasing from 1 to ∼15, where both catenated rings were nicked; (iii) knotted but nicked DNA molecules with a naturally arising spectrum of knots. For better comparison, we studied topoisomer families where each member had the same total molecular mass. For knotted and supercoiled molecules, we analyzed dimeric plasmids whereas catenanes were composed of monomeric forms of the same plasmid. We observed that catenated, knotted and supercoiled families of topoisomers showed different reactions to changes of agarose concentration and voltage during electrophoresis. These differences permitted us to optimize conditions for their separation and shed light on physical characteristics of these different types of DNA topoisomers during electrophoresis.
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: Jorge
authors.initials: J
authors.lastname: Cebrián
authors.affiliation: Scientific and Applied Computing Laboratory, Polytechnic School, National University of Asunción, P.O. Box 2111, SL. San Lorenzo, Paraguay.
authors.forename: Maridian J
authors.initials: MJ
authors.lastname: Kadomatsu-Hermosa
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: Alicia
authors.initials: A
authors.lastname: Castán
authors.affiliation: Scientific and Applied Computing Laboratory, Polytechnic School, National University of Asunción, P.O. Box 2111, SL. San Lorenzo, Paraguay.
authors.forename: Víctor
authors.initials: V
authors.lastname: Martínez
authors.affiliation: Scientific and Applied Computing Laboratory, Polytechnic School, National University of Asunción, P.O. Box 2111, SL. San Lorenzo, Paraguay.
authors.forename: Cristina
authors.initials: C
authors.lastname: Parra
authors.affiliation: Scientific and Applied Computing Laboratory, Polytechnic School, National University of Asunción, P.O. Box 2111, SL. San Lorenzo, Paraguay.
authors.forename: María José
authors.initials: MJ
authors.lastname: Fernández-Nestosa
authors.affiliation: Scientific and Applied Computing Laboratory, Polytechnic School, National University of Asunción, P.O. Box 2111, SL. San Lorenzo, Paraguay.
authors.forename: Christian
authors.initials: C
authors.lastname: Schaerer
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: María-Luisa
authors.initials: ML
authors.lastname: Martínez-Robles
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Hernández
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: Dora B
authors.initials: DB
authors.lastname: Krimer
authors.affiliation: Center for Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, CH-1015 Lausanne, Switzerland schvartzman@cib.csic.es.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Stasiak
authors.affiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas (CSIC), Ramiro de Maeztu 9, 28040 Madrid, Spain schvartzman@cib.csic.es.
authors.forename: Jorge B
authors.initials: JB
authors.lastname: Schvartzman
mesh_terms: D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification; D043864:DNA, Catenated / Q000737:chemistry* / Q000302:isolation & purification; D004278:DNA, Superhelical / Q000737:chemistry* / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D009690:Nucleic Acid Conformation
chemical_list: D043864:DNA, Catenated; D004278:DNA, Superhelical; D004247:DNA