• Korean Journal of Microbiology
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• v.13 no.3
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• pp.109-115
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• 1975

DNA's were extracted from Saccharomyces cerevisiae and Mycobacterium phlei and the damaging cell walls of these microoragnisms were examined under an electron microscope in the extraction process in which a number of physico-chemical tratments of cells was involved. While the DNA was easily extracted from S. cerevisiae using conventional meylelded very little DNA, of M. phlei was extremely difficult to isolate and yielded very little DNA, applying various methods of isolation published earlier. When the cell walls of S. cerevisiae were examined with the electron microscope, they were not yet damaged even after the cells were treated with sodium lauryl sulfate(SLS) and ethylene diamine tetracetic acid(EDTA), but they were completely destroyed by the treatment of sodium perchlorate followed by the addition of chloroform and a vigorous agitation. Oozing cytoplasm through the broken cell walls was also observed. In the extraction of DNA from M.phlei, the pronase was not effective at the aerobic environment of the sample. When phenol was applied at the last step of DNA isolation, an extreme damage mass yielding little DNA into the solution. Unlike the cells of S.cerevisiae.M.phlei cells showed a tendency of aggregation, thus the destruction of cell walls by sodium hydroxide was seen only on the walls of peripheral cells in the aggregated mass, leaving the walls of the inner cells undamaged.

• Journal of Microbiology
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• v.34 no.1
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• pp.30-36
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• 1996

We have exmained the re-establishment of HIMRE mediated silencing function on the transcriptional activity of yeast heast shock gene HSP82. To test whether the onset of SIR repression can occur in growing cells in the rpesence of a potent inhibitor of DNA replication, HMRa/HSP82 strains with SIR4- and SIR4S$^{+}$ genetic backgrounds were arrested in S phase by incubation of a culture in 200 mM hydroxyurea for 120 min. It was clear that following a 20 minute heat shock, silencing of the HMRa/HSP82 allele in cells pretreated with hydroxyurea does occur in a SIR4-dependen fashion, even though the kinetics of repression appears to be substantially delayed. We also have tested whether re- establishement of silencing at the HMR/hsp82 locus can occur in G1-arrested cells. Cell cycle arrest at G1 phase was achieved by treatment of early log a cell cultures with .alpha.-factor mating pheromone, which induces G1 arrest. The result suggests that passage through S phase (and therefore DNA replication) is nor required for re-establishing silencer-mediated repression at the HMNRa/HSP82 locus. Finally, to test whether de nono protein synthesis is required for re-establishment of silencer-mediated repression, cells were pretreated with cycloheximide (500 /.mu.g/ml) 120 min. It was apparent that inhibiting protein synthesis delays, but does not prevent, re-establishment of silencer-mediated repression. Altogether, these results indicate that re-establishment of silencer-mediated repression is not dependent on the DNA replication and has no requirement for protein synthesis.s.