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 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.
Löbrich
M
M
Strahlenzentrum der Justus-Liebig-Universität, Giessen, Germany.
Ikpeme
S
S
Kiefer
J
J
D004247:DNA / Q000528:radiation effects*; D004249:DNA Damage; D015342:DNA Probes; D018076:DNA, Complementary / Q000235:genetics; D004307:Dose-Response Relationship, Radiation; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D007781:Lactoferrin / Q000235:genetics*; D015183:Restriction Mapping; D014965:X-Rays
D015342:DNA Probes; D018076:DNA, Complementary; D004247:DNA; D007781:Lactoferrin