改良彗星实验和RAPD技术检测DNA损伤的实验研究

Experimental study of improvements on comet assay and RAPD for detecting DNA damage

为了有效检测细胞DNA的断裂损伤效应和碱基突变,对彗星实验和RAPD技术进行改进和优化,建立了一套快速、稳定的实验流程。结果显示,改进后的彗星实验方法操作简便、耗时短、实验结果稳定,较好地解决了脱胶、细胞核分离操作繁琐、实验结果重复性低等问题;采用优化后的PCR反应体系和扩增条件,RAPD扩增条带清晰,分辨率高,实验结果稳定,重复性高。将改进和优化后的彗星实验与RAPD技术结合,可快速检测出细胞DNA断裂和碱基突变的损伤效应。

This article studies the improvement and optimization of two methods on the technique process of the comet assay and the RAPD reactive conditions,and the establishment of the quick and stable experiment process for detecting effectively the effects of DNA damage in cells. The results show that the agarose gel debonding,very complex results of the nucleus separation and low repeatability of experimental results are better solved by the improved comet assay and the method is simple and rapid for analysing the DNA damage degree. By optimizing the different RAPD reaction conditions, the stripes of amplification are better distinctive, the resolution is higher and there are good repeatability. The results indicate that the rupture damage and the base mutation of DNA could be detected quickly and efficiently by the combination the comet assay with the RAPD technique.