Abstract
For many years, microtubule research has depended on tubulin purified from cow and pig brains, which may not be ideal for experiments using proteins or extracts from non-brain tissues and cold-blooded organisms. Here, we describe a method to purify functional tubulin from the eggs of the frog, Xenopus laevis. This tubulin has many benefits for the study of microtubules and microtubule based structures assembled in vitro at room temperature. Frog tubulin lacks many of the highly stabilizing posttranslational modifications present in pig brain-derived tubulin, and polymerizes efficiently at room temperature. In addition, fluorescently labeled frog egg tubulin incorporates into meiotic spindles assembled in egg extract more efficiently than brain tubulin, and is thus superior as a probe for Xenopus egg extract experiments. Frog egg tubulin will provide excellent opportunities to identify active nucleation complexes and revisit microtubule polymerization dynamics in vitro.
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Acknowledgements
Our work was funded primarily by NIH grant GM23928. I would like to thank Dr. Mary Badon, MD/MBA, for her guidance and contributions to the figures.
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Groen, A.C., Mitchison, T.J. (2016). Purification and Fluorescent Labeling of Tubulin from Xenopus laevis Egg Extracts. In: Chang, P., Ohi, R. (eds) The Mitotic Spindle. Methods in Molecular Biology, vol 1413. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3542-0_3
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DOI: https://doi.org/10.1007/978-1-4939-3542-0_3
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