|
|
|
GFP
tagged proteins |
|
Cell Cycle progression in yeast can be monitored by observing mitotic spindle morphology using a GFP-tagged tubulin molecule. The spindle forms toward the end of S-phase, but must not elongate (anaphase) to separate the daughter nuclei until all of the genome has been replicated. The S-phase Checkpoint controls spindle dynamics during DNA replication and is essential for genome stability.
|
  |
 |
The S-phase Checkpoint also prevents genome instability by maintaining cohesion between newly replicated DNA molecules of sister chromatids. Without cohesion, the spindle is not able to ensure that sisters segregate away from one another during anaphase. As a result, daughter nuclei become aneuploid. We monitor cohesion at a specified chromosome locus by integrating Tetracyclin operator sequences at that locus. A Tet repressor protein fused to GFP lights up the locus as a discrete fluorescent spot in the yeast nucleus. |