镉对小麦叶片DNA伤害的彗星实验研究

The comet assay detects Cd-induced DNA damages in wheat leaves

为了建立植物基因毒性的研究方法 ,以小麦为对象 ,进行了植物彗星实验方法的研究 .以 0 1mg·L- 1 的Cd2 + 溶液处理小麦叶片后 ,采用机械方法分离细胞核 ,制备玻片后变性 0 ,5 ,15 ,3 0min ,在 10 0 ,2 0 0 ,3 0 0mA下电泳 5 ,15 ,3 0min .实验结果表明 ,采用机械分离的方法可以获得大量的、能够进行彗星实验的细胞核 ,增加DNA的变性时间、电泳电流大小和电泳时间可以增加DNA片段在电场中的迁移 ,提高实验的灵敏度 ,但变性时间和电泳时间过长增加了对照处理DNA片段的迁移 .因此 ,本实验提出了小麦叶片彗星实验的最佳条件是DNA变性 15min ,3 0 0mA下电泳 15min ,并在该条件下研究了重金属镉对小麦的基因损伤 .结果表明 ,重金属镉能引起小麦基因的损伤 。

The comet assay for the detection of DNA damages in wheat leaves had been established, and its application in assessing cadmium (Cd) toxicity was studied. In the experiment, as soon as the treatment with Cd, the nuclei of wheat leaves were separated mechanically and subjected to comet assay. The effects of denaturation in different time and electrophoresis in various time and current were explored. With the time of denaturation and electrophoresis increased, the sensitivity of this method increased simultaneously. But DNA migration also increased in control. Therefore, the appropriate operation conditions had been chosen. The DNA damages (mainly DNA breakages) in nuclei isolated from wheat leaves were detected by the alkaline comet assay. Furthermore, a dose-dependent increase in DNA damages was found with increasing the concentration of Cd in the solution.