DETECTION OF SINGLE-STRAND BREAKS AND FORMAMIDOPYRIMIDINE-DNA GLYCOSYLASE-SENSITIVE SITES IN DNA OF CULTURED HUMAN FIBROBLASTS

被引：41

作者：

CZENE, S

HARMSRINGDAHL, M

机构：

[1] CANC RES INST,DEPT MOLEC VIROL,BRATISLAVA 81232,SLOVAKIA

[2] SWEDISH RADIAT PROTECT INST,S-17116 STOCKHOLM,SWEDEN

来源：

MUTATION RESEARCH-DNA REPAIR | 1995年 / 336卷 / 03期

关键词：

IONIZING RADIATION; SINGLE-STRAND BREAKS; FPG-SENSITIVE DNA DAMAGE; NICK TRANSLATION ASSAY; HUMAN FIBROBLASTS;

D O I：

10.1016/0921-8777(94)00058-E

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Under oxidative stress 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxodG), a damaged base with mutagenic potential, and single-strand breaks (SSB) are formed in DNA. Both lesions are frequently used as a parameter for oxidative damage of DNA. Here we report on results from the evaluation of a modified nick translation assay, where 8-oxodG and SSB formation in cellular DNA of cultured human fibroblasts were simultaneously detected. The assay is based on a method previously described by others, with several modifications in reaction conditions and type of substrate. We used formamidopyrimidine-DNA glycosylase (FPG) in our assay in order to measure the formation of FPG-sensitive lesions (which include 8-oxodG) in DNA of human fibroblasts in response to ionising radiation. The quantification of the DNA damage was based on calibration experiments with plasmid DNA pUC19. Dose-response curves of SSB and FPG-sensitive lesion formation in human fibroblasts VH-10 were established. A very low background level of 8-oxodG was detected in unirradiated fibroblasts (approx. 500 residues per cell).

引用

页码：235 / 242

页数：8

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