Abstract
Southwestern blotting is a technique used to study DNA-protein interactions. This method detects specific DNA-binding proteins by incubating radiolabeled DNA with a gel blot, washing, and visualizing through autoradiography. A blot resulting from 1-dimensional SDS-PAGE reveals the molecular weight of the binding proteins. To increase separation and determine isoelectric point a 2-dimensional gel can be blotted. Additional dimensions of electrophoresis, such as a gel shift (EMSA), can precede isoelectric focusing and SDS-PAGE to further improve separation. Combined with other techniques, such as mass spectrometry, the DNA-binding protein can be identified.
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Acknowledgement
This work was supported by NIH grant R01GM043609. LIN receives partial stipend support from NIH/NIGMS MBRS-RISE GM060655.
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Jia, Y., Nagore, L., Jarrett, H. (2015). Southwestern Blotting Assay. In: Leblanc, B., Rodrigue, S. (eds) DNA-Protein Interactions. Methods in Molecular Biology, vol 1334. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2877-4_5
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DOI: https://doi.org/10.1007/978-1-4939-2877-4_5
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2876-7
Online ISBN: 978-1-4939-2877-4
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