Plasmid DNA Topology Assayed by Two-Dimensional Agarose Gel Electrophoresis
Two-dimensional (2D) agarose gel electrophoresis is nowadays one of the best methods available to analyze DNA molecules with different masses and shapes. The possibility to use nicking enzymes and intercalating agents to change the twist of DNA during only one or in both runs, improves the capacity of 2D gels to discern molecules that apparently may look alike. Here we present protocols where 2D gels are used to understand the structure of DNA molecules and its dynamics in living cells. This knowledge is essential to comprehend how DNA topology affects and is affected by all the essential functions that DNA is involved in: replication, transcription, repair and recombination.
Key wordsDNA topology DNA replication Supercoiling Catenanes Knots Topoisomerases
We acknowledge current and past members of the laboratory for their continuous suggestions and support. We would like to strengthen that this work could not be accomplished without the continuous support and constructive criticism of Andrzej Stasiak. This work was sustained by grant BFU2011-22489 to J.B.S. from the Spanish Ministerio de Economía y Competitividad.
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