SGER: Membrane-Elution and Differential Display Analysis of Cyclic Gene Expression

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The experimental work in the project will measure gene expression during the division cycle of eukaryotic, mammalian cells of both human and mouse origin. The newly developed membrane elution (or baby machine) method for producing undisturbed, synchronous cultures of eukaryotic cells will be used to analyze the precise pattern and timing of expression of different genes during the mammalian division cycle. This method produces unperturbed synchronized cells that exhibit three generations of synchronized divisions with the proper DNA contents and cell sizes observed throughout the successive cell cycles. The hypothesis to be tested is that there are few cell-cycle-regulated genes in eukaryotic cells, and in particular, there are no G1-phase specific patterns of gene expression. In the membrane-elution method, exponentially growing, unperturbed cells are bound to a membrane. The membrane is washed with warm medium so cells grow and divide on the membrane. Only newborn cells are released from the membrane by division. A short collection period yields a synchronized culture. The broader impact of this work is that the entire field of cell biology, arguably depends on a correct understanding of the nature of the cell cycle. If the hypothesis proves correct, it could have a major impact on how we understand the cell cycle.