Abstract
Pulsed-field gel electrophoresis (PFGE) is a technique that resolves chromosome-sized DNA molecules in an agarose gel. As well as DNA mapping and karyotyping applications, PFGE techniques are well adapted to follow DNA rearrangements over time in a quantitative manner. Because of the very large sizes of the DNA molecules analyzed, DNA preparation, electrophoresis, and Southern blotting processes present unique challenges in PFGE experiments. In this chapter, we describe a robust PFGE protocol covering the preparation of intact Saccharomyces cerevisiae chromosomal DNA, specific running conditions for the resolution of small, medium- and large-sized chromosomes and their by-products, and basic Southern blotting and hybridization instructions for the analysis of these molecules.
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Acknowledgements
The setup of the PFGE protocol was initially started in the laboratory of Professor David Tollervey.
A.E.H. is a research fellow in the laboratory of Professor David Tollervey who is funded by the Welcome Trust. J.H. is funded by Wellcome Trust grant 088335.
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Hage, A.E., Houseley, J. (2013). Resolution of Budding Yeast Chromosomes Using Pulsed-Field Gel Electrophoresis. In: Makovets, S. (eds) DNA Electrophoresis. Methods in Molecular Biology, vol 1054. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-565-1_13
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DOI: https://doi.org/10.1007/978-1-62703-565-1_13
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Publisher Name: Humana Press, Totowa, NJ
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