Flow cytometry is a technique used to determine cell cycle stage by measuring DN
ID: 3166543 • Letter: F
Question
Flow cytometry is a technique used to determine cell cycle stage by measuring DNA content. Shown to the right are three flow cytometry profiles. Choose the experimental situation that is most likely to lead to each profile and provide a brief explanation for how the cell distribution in the profile supports your choice. (2 points each, 6 points total) IN 2N IN 2N 1N 2N A) an asynchronous population of wildtype cells f a temperature-sensitive yeast mutant that is defective in securin degradation at the non- permissive temperature C) wildtype cells that were synchronized in G1 and then released into media that contains hydroxyurea, which is known to limit free nucleotides D) You have identified a novel yeast mutant, googlenope, in a screen for genes defective in the activation of the G2 DNA damage checkpoint pathway. As an initial test to determine whether googlenope might be part of the checkp DNA damaging agent. Cells are synchronized in G a DNA damaging then analyzed by flow cytometry. When this is performed with wildtype cells, the proportion of the cells with 2N DNA content accumulates but no cells esult do you expect to observe if googlenope is in fact part of the DNA damage checkpoint? Briefly explain. (3 oint pathway, you perform a cell synchronization experiment to observe how the cells respond to a 1 and then allowed to proceed into S phase, at which point chemical is added to the cells. Samples are collected at 20-minute intervals for 2 hours and with 1N DNA content appear, not even in the later timepoints. What points)Explanation / Answer
(A) In an asynchronous population of cells , some will exists in G0/1(1N) or G2/M phase (2N) therefore 2nd profile is of this situation.
?(B) 1st profile is of this situation as securin degradation will effect cell division at metaphase, therefore cell at G2 will accumulate normally but due to restriction of m phase new duaghter cell wll not form thereby reducing G1 stage population.
?(C) hydroxyurea effect cell cycle by inhibiting dna replication which is by exhuasting or depleting dNTPs before s phase but this does not happens at 100% . The dNTPs pool inspite of hydroxyurea presence does not completely exhuast therefore there is a mixed population of G1 andG2 cells. hence the 3rd profile is this stiuation.
?(D)if googlenope is defective in G2 DNA damage checkpoint pathway then for these mutant the result will not the same there will be cells in G1 phase as DNA damage was not detected due to this defect.
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