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Isolation and Purification of Kinesin from Drosophila Embryos
JoVE Journal
Biology
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JoVE Journal Biology
Isolation and Purification of Kinesin from Drosophila Embryos
DOI:

09:49 min

April 27, 2012

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Chapters

  • 00:05Title
  • 02:28Fly Cup Preparation and Embryo Collection
  • 04:00Embryo Homogenization and Clarification
  • 04:48Microtubule Polymerization, Kinesin Binding and Differential Sedimentation of Microtubules and Kinesin
  • 06:49Representative Kinesin Purification Results
  • 08:29Conclusion

Summary

Automatic Translation

This is a protocol to isolate active full length Kinesin from Drosophila embryos for single-molecule biophysical studies. We show how to collect embryos, make the embryo lysate, and then polymerize microtubules (MTs). Kinesin is purified by immobilizing it on the MTs, spinning down the Kinesin-MT complexes, and then releasing the kinesin from the MTs via ATP addition.

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