A reliable assessment of 8-oxo-2-deoxyguanosine levels in nuclear and mitochondrial DNA using the sodium iodide method to isolate DNA.
A major controversy in the area of DNA biochemistry concerns the actual in vivo levels of oxidative damage in DNA. We show here that 8-oxo-2-deoxyguanosine (oxo8dG) generation during DNA isolation is eliminated using the sodium iodide (NaI) isolation method and that the level of oxo8dG in nuclear DNA (nDNA) is almost one-hundredth of the level obtained using the classical phenol method. We found using NaI that the ratio of oxo8dG/10(5 )deoxyguanosine (dG) in nDNA isolated from mouse tissues ranged from 0.032 +/- 0.002 for liver to 0.015 +/- 0.003 for brain. We observed a significant increase (10-fold) in oxo8dG in nDNA isolated from liver tissue after 2 Gy of gamma-irradiation when NaI was used to isolate DNA. The turnover of oxo8dG in nDNA was rapid, e.g. disappearance of oxo8dG in the mouse liver in vivo after gamma-irradiation had a half-life of 11 min. The levels of oxo8dG in mitochondrial DNA isolated from liver, heart and brain were 6-, 16- and 23-fold higher than nDNA from these tissues. Thus, our results showed that the steady-state levels of oxo8dG in mouse tissues range from 180 to 360 lesions in the nuclear genome and from one to two lesions in 100 mitochondrial genomes.
Hamilton
M L
ML
Department of Physiology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229-3900, USA.
Guo
Z
Z
Fuller
C D
CD
Van Remmen
H
H
Ward
W F
WF
Austad
S N
SN
Troyer
D A
DA
Thompson
I
I
Richardson
A
A
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism; D002467:Cell Nucleus / Q000235:genetics* / Q000528:radiation effects; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D004272:DNA, Mitochondrial / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000032:analysis*; D005720:Gamma Rays; D006207:Half-Life; D006801:Humans; D008099:Liver / Q000166:cytology / Q000378:metabolism / Q000528:radiation effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D016877:Oxidants / Q000378:metabolism; D018384:Oxidative Stress / Q000528:radiation effects; D010240:Parakeets; D019800:Phenol / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D012680:Sensitivity and Specificity; D012974:Sodium Iodide / Q000378:metabolism*
D004272:DNA, Mitochondrial; D016877:Oxidants; D019800:Phenol; D000080242:8-Hydroxy-2'-Deoxyguanosine; D004247:DNA; D012974:Sodium Iodide; D003849:Deoxyguanosine