DNA double-strand break measurement in mammalian cells by pulsed-field gel electrophoresis: an approach using restriction enzymes and gene probing

DNA samples prepared from human SP3 cells, which had been exposed to various doses of X-ray, were treated with NotI restriction endonuclease before being run in a contour-clamped homogeneous electrophoresis system. The restriction enzyme cuts the DNA at defined positions delivering DNA sizes which c...

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Bibliographic Details
Published inInternational journal of radiation biology Vol. 65; no. 6; p. 623
Main Authors Löbrich, M, Ikpeme, S, Kiefer, J
Format Journal Article
LanguageEnglish
Published England 1994
Subjects
DNA - radiation effects
DNA Damage
DNA Probes
DNA, Complementary - genetics
Dose-Response Relationship, Radiation
Electrophoresis, Gel, Pulsed-Field
Humans
Lactoferrin - genetics
Restriction Mapping
X-Rays
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Summary:DNA samples prepared from human SP3 cells, which had been exposed to various doses of X-ray, were treated with NotI restriction endonuclease before being run in a contour-clamped homogeneous electrophoresis system. The restriction enzyme cuts the DNA at defined positions delivering DNA sizes which can be resolved by pulsed-field gel electrophoresis (PFGE). In order to investigate only one of the DNA fragments, a human lactoferrin cDNA, pHL-41, was hybridized to the DNA separated by PFGE. As a result, only the DNA fragment which contains the hybridized gene was detected resulting in a one-band pattern. The decrease of this band was found to be exponential with increasing radiation dose. From the slope, a double-strand break induction rate of (6.3 +/- 0.7) x 10(-3)/Mbp/Gy was deduced for 80 kV X-rays.
ISSN:0955-3002
DOI:10.1080/09553009414550731