Mitotic spindle assembly on chromatin patterns made with deep UV photochemistry.
We provide a detailed method to generate arrays of mitotic spindles in vitro. Spindles are formed in extract prepared from unfertilized Xenopus laevis eggs, which contain all the molecular ingredients of mitotic spindles. The method is based on using deep UV photochemistry to attach chromatin-coated beads on a glass surface according to a pattern of interest. The immobilized beads act as artificial chromosomes, and induce the formation of mitotic spindles in their immediate vicinity. To perform the experiment, a chamber is assembled over the chromatin pattern, Xenopus egg extract is flowed in and after incubation the spindles are imaged with a confocal microscope.
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
Katarzyna
K
Tarnawska
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
Céline
C
Pugieux
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
François
F
Nédélec
D000818:Animals; D002457:Cell Extracts; D002843:Chromatin / Q000378:metabolism*; D005260:Female; D010063:Ovum / Q000378:metabolism / Q000528:radiation effects; D010777:Photochemistry / Q000379:methods*; D008941:Spindle Apparatus / Q000378:metabolism* / Q000528:radiation effects*; D014466:Ultraviolet Rays; D014982:Xenopus laevis
D002457:Cell Extracts; D002843:Chromatin
Deep UV photochemistry; Micro-patterning; Microtubules; Mitotic spindle; Xenopus egg extract