The use of biphasic linear ramped pulsed field gel electrophoresis to quantify DNA damage based on fragment size distribution.
OBJECTIVE
The development of biphasic linear pulse ramping gel electrophoresis has permitted resolution of DNA fragments from 200 Kbp to 6 Mbp in a single gel. We used this technique to measure radiation-induced DNA damage based on fragment size.


METHODS
Human colon cancer cells (HT29 and LS174T) and Chinese hamster ovary cells were embedded in agarose, deproteinized, irradiated with 5-80 Gy, and assessed for DNA double strand breakage using pulsed field gel electrophoresis. The frequency of DNA double strand breakage determined using a previously published method was compared to the breakage frequency calculated using the fragment size distribution.


RESULTS
Both methods produced similar estimates for breakage frequency of approximately 5 x 10(-9) breaks Gy-1 bp-1.


CONCLUSIONS
These findings suggest that biphasic linear pulse ramping gel electrophoresis can yield a quantitative estimate of DNA fragment distribution resulting from irradiation. The ability to quantify the distribution of DNA fragment sizes produced by irradiation should yield important information concerning the mechanisms of both DNA double strand break induction and repair.
Lawrence
T S
TS
University of Michigan Medical Center, Department of Radiation Oncology, Ann Arbor 48109.
Normolle
D P
DP
Davis
M A
MA
Maybaum
J
J
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA / Q000032:analysis*; D004249:DNA Damage; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D014407:Tumor Cells, Cultured
D004247:DNA