Supplemental Materials
This article contains the following supporting material:
- Supplemental Materials
- Movie01 - Movie 1: Time-lapse movie of a K-fragment generated at anaphase onset. In the beginning at metaphase, the three autosomal bivalents are positioned at the spindle equator. At anaphase onset (00:45), cohesion between homologues is lost. Subsequent to the laser flash (between 01:00 and 01:15), the K-fragment moves poleward, and its severed arms move backward to make contact with its partner homologue. The K-fragment has a poleward velocity twice that of controls. Some collateral damage is evident in the arms of adjacent chromosomes, an unavoidable complication when using the laser beam configured as a line, as it was in this case. Bar, 5�m; time in minutes:seconds.
- Movie02 - Movie 2: Control experiment. Time-lapse alternating frame FSM/DIC movie after a spot ablation that drilled a hole in a chromosome arm but did not detach the arm from its kinetochore. With FSM following injection of Rh-tubulin, kinetochore fibers become strikingly fluorescent. Before the laser flash, locations of plus end attachment to bivalents are clearly evident. After the onset of anaphase (00:00), photobleaching is evident as the laser beam in spot flash mode was used to drill a hole in one of the trailing arms. Recovery of fluorescence at the plus ends of kinetochore microtubules is evident as plus end addition of Rh-tubulin monomers � due to the polymerization state of early anaphase kinetochores � continues after the operation. Bar, 5�m; time in minutes:seconds.
- Movie03 - Movie 3: Time-lapse alternating frame FSM/DIC movie after generating a K-fragment at anaphase onset. FSM illustrates photobleaching at and around the flash site. Recovery of fluorescence at the plus ends of control K-fibers has already begun in the first fluorescence frame (00:00) after the laser flash. Fluorescence recovery after photobleaching (FRAP) is evidet at the kinetochore regions of controls; whereas such fluorescence is not evident at the K-fragment�s kinetochore. When fluorescence is eventually recovered at the K-fragment both the fragment and its associated fluorescence move poleward at similar velocities. Bar, 5�m; time in minutes:seconds.
- Movie04 - Movie 4: Time-lapse alternating frame FSM/DIC movie after generating a K-fragment at anaphase onset. FSM images after detachment of the K-fragment illustrate the lack of fluorescence at the plus end of the fragment�s K-fiber in comparison to fluorescence emerging from the kinetochores of controls moving to the same pole. Bar, 5�m; time in minutes:seconds.