Factors influencing DNA migration from individual cells subjected to gel electrophoresis.
Alkaline and neutral gel electrophoresis of individual mammalian cells allows detection of DNA single- and double-strand breaks, respectively. For both the alkaline and the neutral assays, lysis conditions influence how much DNA migrates, and factors in addition to DNA size play a role in migration. In particular, the tight packing of DNA in individual nuclei appears to reduce the ability to detect double-strand breaks in all of the genome. Tangling of DNA molecules is probably also responsible for the presence of "wings" associated with each nucleus after application of pulsed-field gel electrophoresis; these wings were aligned in the directions of the pulsed field, not along the resultant vector of the fields as was expected. The choice of fluorescent staining methods (propidium iodide, Hoechst 33342, or antibodies against bromodeoxyuridine) did not influence sensitivity for detecting DNA damage.
British Columbia Cancer Research Centre, Vancouver, Canada.
P L
PL
Olive
D
D
Wlodek
R E
RE
Durand
J P
JP
Banáth
D002460:Cell Line; D004247:DNA / Q000737:chemistry* / Q000187:drug effects / Q000528:radiation effects; D004249:DNA Damage; D004307:Dose-Response Relationship, Radiation; D016521:Electrophoresis, Gel, Pulsed-Field; D005720:Gamma Rays / Q000009:adverse effects; D008466:Mechlorethamine / Q000633:toxicity; D008856:Microscopy, Fluorescence; D012680:Sensitivity and Specificity; D013194:Staining and Labeling
D008466:Mechlorethamine; D004247:DNA