Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine

We report procedures to allow incorporation and detection of 5-ethynyl-2'-deoxyuridine (EdU) in fission yeast, a thymidine analogue which has some technical advantages over use of bromode-oxyuridine. Low concentrations of EdU (1μM) are sufficient to allow detection of incorporation in cells exp...

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Hlavní autoři: Hua, H, Kearsey, S
Médium: Journal article
Jazyk:English
Vydáno: Oxford University Press 2011
Témata:
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author Hua, H
Kearsey, S
author_facet Hua, H
Kearsey, S
author_sort Hua, H
collection OXFORD
description We report procedures to allow incorporation and detection of 5-ethynyl-2'-deoxyuridine (EdU) in fission yeast, a thymidine analogue which has some technical advantages over use of bromode-oxyuridine. Low concentrations of EdU (1μM) are sufficient to allow detection of incorporation in cells expressing thymidine kinase and human equilibrative nucleoside transporter 1 (hENT1). However EdU is toxic and activates the rad3-dependent checkpoint, resulting in cell cycle arrest, potentially limiting its applications for procedures which require labelling over more than one cell cycle. Limited DNA synthesis, when elongation is largely blocked by hydroxyurea, can be readily detected by EdU incorporation using fluorescence microscopy. Thus EdU should be useful for detecting early stages of S phase, or DNA synthesis associated with DNA repair and recombination.
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spelling oxford-uuid:3be6e4cf-dbdb-4938-838a-147d280c4d592022-03-26T14:10:15ZMonitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridineJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:3be6e4cf-dbdb-4938-838a-147d280c4d59Zoological sciencesEnglishOxford University Research Archive - ValetOxford University Press2011Hua, HKearsey, SWe report procedures to allow incorporation and detection of 5-ethynyl-2'-deoxyuridine (EdU) in fission yeast, a thymidine analogue which has some technical advantages over use of bromode-oxyuridine. Low concentrations of EdU (1μM) are sufficient to allow detection of incorporation in cells expressing thymidine kinase and human equilibrative nucleoside transporter 1 (hENT1). However EdU is toxic and activates the rad3-dependent checkpoint, resulting in cell cycle arrest, potentially limiting its applications for procedures which require labelling over more than one cell cycle. Limited DNA synthesis, when elongation is largely blocked by hydroxyurea, can be readily detected by EdU incorporation using fluorescence microscopy. Thus EdU should be useful for detecting early stages of S phase, or DNA synthesis associated with DNA repair and recombination.
spellingShingle Zoological sciences
Hua, H
Kearsey, S
Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title_full Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title_fullStr Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title_full_unstemmed Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title_short Monitoring DNA replication in fission yeast by incorporation of 5-ethynyl-2'-deoxyuridine
title_sort monitoring dna replication in fission yeast by incorporation of 5 ethynyl 2 deoxyuridine
topic Zoological sciences
work_keys_str_mv AT huah monitoringdnareplicationinfissionyeastbyincorporationof5ethynyl2deoxyuridine
AT kearseys monitoringdnareplicationinfissionyeastbyincorporationof5ethynyl2deoxyuridine