In Vitro Assay for Plasmid Length DNA Strand Exchange by Human DMC1.

Autor: Goodson SD; Department of Genetics and Biochemistry, Clemson University, Clemson, SC, USA.; Eukaryotic Pathogens Innovation Center, Clemson University, Clemson, SC, USA., Hawes RB; Department of Genetics and Biochemistry, Clemson University, Clemson, SC, USA., Waldvogel SM; Department of Genetics and Biochemistry, Clemson University, Clemson, SC, USA., Sehorn MG; Department of Genetics and Biochemistry, Clemson University, Clemson, SC, USA. msehorn@clemson.edu.; Eukaryotic Pathogens Innovation Center, Clemson University, Clemson, SC, USA. msehorn@clemson.edu.; Clemson University School of Health Research, Clemson, SC, USA. msehorn@clemson.edu.; Center for Optical Materials Science and Engineering Technologies, Clemson University, Clemson, SC, USA. msehorn@clemson.edu.
Jazyk: angličtina
Zdroj: Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2019; Vol. 1999, pp. 265-270.
DOI: 10.1007/978-1-4939-9500-4_17
Abstrakt: Meiosis is a specialized cell division that generates gametes. Meiotic recombination is essential not only to generate diversity in offspring, but also to hold homologous chromosomes together through chiasma allowing proper chromosome segregation. This process requires the meiosis-specific recombinase, DMC1. DMC1 facilitates the search for homology between the homologous chromosomes and is followed by DNA strand invasion and strand exchange to produce a linkage between the two homologous chromosomes. The development of biochemical in vitro assays and the purification of the requisite proteins factors has led to a better understanding of the molecular mechanisms of meiotic homologous recombination. In this chapter, a detailed in vitro assay to examine DNA strand exchange over 5000 bases of DNA catalyzed by human DMC1 is described. This method has proved to be valuable for examining the catalytic potential of hDMC1 and delineating the functional interaction with other HR factors.
Databáze: MEDLINE
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