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PLoS One. 2014 Feb 13;9(2):e88629. doi: 10.1371/journal.pone.0088629. eCollection 2014.

Cell-cycle analyses using thymidine analogues in fission yeast.

Author information

1
Department of Cell Biology, Institute for Cancer Research, Oslo University Hospital, Oslo, Montebello, Norway.

Abstract

Thymidine analogues are powerful tools when studying DNA synthesis including DNA replication, repair and recombination. However, these analogues have been reported to have severe effects on cell-cycle progression and growth, the very processes being investigated in most of these studies. Here, we have analyzed the effects of 5-ethynyl-2'-deoxyuridine (EdU) and 5-Chloro-2'-deoxyuridine (CldU) using fission yeast cells and optimized the labelling procedure. We find that both analogues affect the cell cycle, but that the effects can be mitigated by using the appropriate analogue, short pulses of labelling and low concentrations. In addition, we report sequential labelling of two consecutive S phases using EdU and 5-bromo-2'-deoxyuridine (BrdU). Furthermore, we show that detection of replicative DNA synthesis is much more sensitive than DNA-measurements by flow cytometry.

PMID:
24551125
PMCID:
PMC3923809
DOI:
10.1371/journal.pone.0088629
[Indexed for MEDLINE]
Free PMC Article

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