基于荧光标记发夹式DNA探针的汞离子定量检测技术的研究

发布时间：2018-01-14 05:34

本文关键词：基于荧光标记发夹式DNA探针的汞离子定量检测技术的研究　出处：《太原理工大学》2017年硕士论文　论文类型：学位论文

【摘要】：环境污染已成为一个全球性问题,而重金属污染在环境污染中备受关注,汞污染作为较严重的一种重金属污染,对生态环境具有严重的危害。由于汞难被降解,能够在土壤、水、空气中广泛富集,最后经过食物链和大气等进入人体,严重破坏蛋白和酶的活性,造成器官中毒。为了更好的预防汞污染带来的食品安全、疾病危机等问题,开发一种方便快捷、灵敏高效,并且能够实现现场检测,用于临床诊断的检测手段变得越来越重要。在汞离子的检测方法中使用最广泛的属于常规的大型仪器检测方法,如原子吸收光谱法、ICP-MS等方法,这些方法在检测灵敏度和精度上都非常高,但是操作比较复杂,而且耗时也相对较长。而面对日益严峻复杂的汞污染挑战,为了实现对不同环境现场的汞污染检测,研究人员也在不断开发一些新型的Hg~(2+)检测方法。如试纸条法、化学传感器法,纳米材料传感器法以及DNA探针法等,试纸条法是比较简便的一种方法,但只能做定性检测,无法实现定量分析,而其他检测方法基本都是利用化学分子或一些官能团能够结合Hg~(2+),引起分子结构或者分子的发光特性的改变,最后通过观察吸收谱或光谱来实现对Hg~(2+)的定量检测,相对传统大型仪器方法,这些方法在操作和成本上具有很大改善,但在灵敏度和耗时上仍需改进。结合DNA链特性和荧光共振能量转移技术,在本课题中我们设计了一种发夹式的富含T碱基的DNA链,无需繁琐的温度处理,实现在常温下一步法检测Hg~(2+)。在DNA链两端分别修饰荧光染料Cy3、Cy5,通过观察Cy3荧光强度的变化,将其变化与Hg~(2+)浓度进行线性拟合,我们发现随着Hg~(2+)浓度的增加,Cy3的荧光强度在不断增大,它们之间具有线性相关性,并且线性相关系数为0.9836,而且我们的方法最低检测限能够达到1.3 nM,低于国家饮用水中Hg~(2+)的标准检测限5 nM。随后对方法的选择性进行了研究,加入不同的干扰离子,发现我们的方法对Hg~(2+)具有好的选择性,能够特异性识别Hg~(2+),实现了对Hg~(2+)的选择性的定量检测。
[Abstract]:Environmental pollution has become a global problem, and heavy metal pollution in the environmental pollution has received much attention. As a serious heavy metal pollution, mercury pollution has serious harm to the ecological environment, because mercury is difficult to be degraded. Can be widely enriched in soil, water, air, and finally through the food chain and atmosphere into the human body, seriously damage the activity of proteins and enzymes, resulting in organ poisoning. In order to better prevent mercury pollution brought food safety. Disease crisis and other problems, the development of a convenient, rapid, sensitive and efficient, and can achieve on-site detection. The detection methods for clinical diagnosis are becoming more and more important. The most widely used methods of mercury ion detection are the conventional large-scale instrument methods, such as atomic absorption spectrometry (AAS) and ICP-MS. These methods are very sensitive and accurate, but the operation is complex and time-consuming, and face the increasingly serious challenges of mercury pollution. In order to detect mercury pollution in different environmental sites, researchers have been developing some new Hg~(2 detection methods, such as test strip method, chemical sensor method. Nanomaterial sensor method and DNA probe method, the test strip method is a relatively simple method, but can only do qualitative testing, can not achieve quantitative analysis. Other detection methods are based on the use of chemical molecules or some functional groups can bind to Hg~(2, resulting in changes in molecular structure or molecular luminescence properties. Finally, the quantitative detection of Hg~(2 is realized by observing the absorption spectrum or spectrum. Compared with the traditional large-scale instrument method, these methods have a great improvement in operation and cost. However, we still need to improve the sensitivity and time consuming. Combining with the characteristics of DNA chain and fluorescence resonance energy transfer technology, we designed a hairpin DNA chain rich in T-base. The Hg~(2 can be detected by one step method at room temperature without complicated temperature treatment. The fluorescent dye Cy3C Cy5 is modified at the two ends of DNA chain, and the change of fluorescence intensity of Cy3 is observed. By linear fitting with the concentration of Hg~(2, we found that the fluorescence intensity of Cy3 increased with the increase of the concentration of Hg~(2, and there was a linear correlation between them. The linear correlation coefficient is 0.9836, and the minimum detection limit of our method can reach 1.3 nm. The detection limit of 5 nm was lower than the standard detection limit of Hg~(2 in drinking water. Then the selectivity of the method was studied and different interference ions were added. It is found that our method has good selectivity for Hg~(2, and can specifically identify Hg~(2, thus realizing the selective quantitative detection of Hg~(2.
【学位授予单位】：太原理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.3

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