Koppen G, Angelis K J
Flemish Institute of Technological Research, Environmental Toxicology, Mol, Belgium.
Environ Mol Mutagen. 1998;32(3):281-5.
The comet assay was used to measure DNA damage and repair in nuclei released from 1 cm root ends of Vicia faba after X-ray irradiation. Irradiation induced a linear increase of DNA content in comet tail with doses under various denaturation and electrophoretic conditions. The pH of the electrophoresis solution played the most important role in the detection of DNA damage. After irradiation with 30 Gy of X-rays, most of the DNA damage was removed during the first 20 min, even in the presence of DNA repair inhibitors. This first, rapid phase of DNA repair was not affected by incubation on ice, but was partially blocked by 3-aminobenzamide. When DNA was exposed to alkali (0.3 M NaOH) and electrophoresed at neutral pH, all DNA damage was removed in 2 hr, even in the presence of 3-aminobenzamide. Complete repair was inhibited by incubation on ice (30% of DNA remaining in tail) and partially by aphidicolin (13% DNA remaining in tail). Under alkaline (0.3 M NaOH) pretreatment and electrophoresis, more than 20% of detected DNA damage remained unrepaired after 2 hr of postirradiation incubation with and without 3-aminobenzamide at room temperature. Aphidicolin and incubation on ice inhibited the removal of DNA damage to 33% and 39% DNA, respectively. Moreover, aphidicolin treatment attenuated the first phase of damage removal.
彗星试验用于测量经X射线照射后蚕豆1厘米根尖释放细胞核中的DNA损伤与修复情况。在不同变性和电泳条件下，照射剂量与彗星尾中DNA含量呈线性增加。电泳溶液的pH值在DNA损伤检测中起最重要作用。经30 Gy X射线照射后，即使存在DNA修复抑制剂，大部分DNA损伤在最初20分钟内也会被去除。DNA修复的这一快速初始阶段不受冰上孵育的影响，但会被3-氨基苯甲酰胺部分阻断。当DNA暴露于碱（0.3 M NaOH）并在中性pH下电泳时，即使存在3-氨基苯甲酰胺，所有DNA损伤在2小时内也会被去除。冰上孵育（尾中剩余30%的DNA）和阿非科林（尾中剩余13%的DNA）会分别部分抑制完全修复。在碱性（0.3 M NaOH）预处理和电泳条件下，在室温下照射后孵育2小时，无论有无3-氨基苯甲酰胺，超过20%检测到的DNA损伤仍未修复。阿非科林和冰上孵育分别将DNA损伤的去除率抑制到33%和39%。此外，阿非科林处理减弱了损伤去除的第一阶段。
