Efficient labeling of fission yeast Schizosaccharomyces pombe with thymidine and BUdR.
In this paper we report the construction of a Schizosaccharomyces pombe strain that facilitates analysis of replicating DNA. The strain co-expresses the Herpes simplex virus thymidine kinase gene (hsv-tk) and a human equilibrative nucleoside transporter (hENT1). The double integrant efficiently incorporates 3H-thymidine into nuclear DNA as monitored by scintillation counting. These strains also incorporate the thymidine analog Bromodeoxy uridine (BUdR) into newly replicated DNA, which can be detected by immunofluorescence and flow cytometry. This strain provides a valuable tool for direct study of DNA replication in S.pombe.
Molecular and Cell Biology Laboratory, The Salk Institute, 10010 North Torrey Pines Road, La Jolla, CA 92037, USA.
Jeffrey A
JA
Hodson
Julie M
JM
Bailis
Susan L
SL
Forsburg
D001973:Bromodeoxyuridine / Q000378:metabolism*; D004261:DNA Replication; D033721:Equilibrative Nucleoside Transporter 1 / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D016193:G1 Phase; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D009584:Nitrogen / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016196:S Phase; D012568:Schizosaccharomyces / Q000166:cytology / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics; D013936:Thymidine / Q000378:metabolism*; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D019076:Transgenes / Q000235:genetics
D033721:Equilibrative Nucleoside Transporter 1; D012333:RNA, Messenger; C438649:SLC29A1 protein, human; D013937:Thymidine Kinase; D001973:Bromodeoxyuridine; D009584:Nitrogen; D013936:Thymidine