基于DNA杂交的新型液晶传感器及其应用
发布时间：2018-10-08 14:06
【摘要】：近年来,DNA检测广泛应用于临床诊断、基因表达、农业、食品安全以及环境监测等领域,因此对DNA进行快速、便捷、灵敏的检测十分必要。传统的DNA检测方法有基于荧光的生物芯片技术、表面等离子共振(Surface Plasmon Resonance,SPR)等,但是这些检测方法存在诸多不足,仍需改进。近年来,具有独特物理化学性质的新型材料——液晶已经引起研究人员的广泛关注。液晶是介于液态与固态之间的一种物质形态,具有双折射性、弹性应变等特性,基底表面微小的变化即可引起液晶分子取向的变化,而液晶分子取向的变化可以转变为光学信号,肉眼即可观察,因此在DNA检测中备受关注。本文基于DNA杂交和液晶的双折射性,构建了固态-液晶传感界面,并在其基础上对DNA杂交进行了检测;除此之外,本文还构建了水-液晶界面,探究了不同表面活性剂对液晶分子排列的影响,并在其基础上探索了单链DNA(ssDNA)对水-液晶界面的影响,为DNA杂交的检测奠定了基础。具体内容如下:(1)利用ssDNA作为取向剂在固态-液晶界面构建功能化基底。首先将三乙氧基硅基丁醛(TEA)修饰于固态基底表面,利用带有醛基的TEA与修饰有氨基的ssDNA之间的反应,将ssDNA修饰在固态基底表面。探究了玻片在TEA中的修饰时间以及ss DNA在基底表面的表面覆盖率对于功能化基底构建的影响。(2)基于固态-液晶界面检测DNA杂交的探索。采用上述方法构建一种新型的利用ssDNA诱导液晶4-氰基-4’-戊基联苯(5CB)垂直排列的功能化基底,通过优化实验条件,ssDNA会诱导5CB分子垂直排列,偏光显微镜下观察到均一的黑色,当加入互补ssDNA之后,会发生杂交反应,扰乱5CB分子的排列,偏光显微镜下观察到彩色纹理,从而达到检测DNA的效果。在优化实验条件(玻片在TEA溶液中的修饰时间为1h,ssDNA的最佳浓度为20 nM)下,该检测方法的检测限为0.1 nM,并且具有良好的特异性和重复性。(3)在水-液晶界面不同表面活性剂对5CB分子取向的影响。根据表面活性剂与5CB分子之间的相互作用,探讨了在水-液晶界面不同表面活性剂(十八烷基三甲基溴化铵,十二烷基三甲基溴化铵,正辛基三甲基溴化铵,十二烷基硫酸钠)对5CB分子取向的影响,探究了每种表面活性剂诱导5CB分子垂直排列的最佳浓度与响应时间,链长不同的同类表面活性剂对5CB分子取向的影响,以及链长相同的不同类表面活性剂对5CB分子取向的影响。除此之外,我们还探究了在水-液晶界面ssDNA对5CB分子取向的影响,为后续DNA杂交的检测提供了方法与条件。
[Abstract]:In recent years, DNA detection has been widely used in clinical diagnosis, gene expression, agriculture, food safety and environmental monitoring, so it is necessary to detect DNA quickly, conveniently and sensitively. Traditional DNA detection methods are based on fluorescence biochip technology, surface plasmon resonance (Surface Plasmon Resonance,SPR) and so on, but these detection methods have many shortcomings and still need to be improved. In recent years, liquid crystal, a new material with unique physical and chemical properties, has attracted wide attention. Liquid crystal is a kind of material form between liquid and solid. It has properties of birefringence, elastic strain and so on. The change of the orientation of liquid crystal molecules can be transformed into optical signals, which can be observed by the naked eye, so it has attracted much attention in DNA detection. Based on the birefringence of DNA hybridization and liquid crystal, the solid-liquid crystal sensing interface was constructed, and the DNA hybridization was detected on the basis of it. In addition, the water-liquid crystal interface was constructed in this paper. The effects of different surfactants on the arrangement of liquid crystal molecules were investigated, and the effects of single strand DNA (ssDNA) on the water-liquid crystal interface were explored, which laid a foundation for the detection of DNA hybridization. The main contents are as follows: (1) ssDNA was used as the orientation agent to construct the functional substrate at the solid-liquid crystal interface. Firstly, triethoxysilyl butyraldehyde (TEA) was modified on the solid substrate surface. SsDNA was modified on the solid substrate surface by the reaction between TEA with aldehyde group and ssDNA modified with amino group. The effects of glass modification time in TEA and ss DNA surface coverage on the construction of functional substrate were investigated. (2) Detection of DNA hybridization based on solid-liquid crystal interface. The above method was used to construct a novel functional substrate for the vertical alignment of liquid crystal 4-cyano-4keto-pentyl biphenyl (5CB) induced by ssDNA. The vertical arrangement of 5CB molecules was induced by optimizing the experimental conditions, and the uniform black was observed under polarizing microscope. When the complementary ssDNA is added, the hybridization reaction will occur, and the arrangement of 5CB molecules will be disturbed. The color texture can be observed under the polarizing microscope, thus the effect of DNA detection can be achieved. Under the optimized experimental conditions (the time of modification of glass slides in TEA solution is 1 h), the optimum concentration of DNA is 20 nM. The detection limit of this method is 0.1 nM, and has good specificity and repeatability. (3) the effect of different surfactants on the orientation of 5CB molecules at the water-liquid crystal interface. Based on the interaction between surfactants and 5CB molecules, different surfactants (octadecyl trimethylammonium bromide, dodecyl trimethyl ammonium bromide, octyl trimethylammonium bromide) at the water-liquid crystal interface were studied. The effect of sodium dodecyl sulfate on the orientation of 5CB molecules was investigated. The optimum concentration and response time of each surfactant to induce the vertical alignment of 5CB molecules, and the effects of the same surfactants with different chain length on the orientation of 5CB molecules were investigated. And the effects of different surfactants with the same chain length on the orientation of 5CB molecules. In addition, we also investigated the effect of ssDNA on the orientation of 5CB molecules at the water-liquid crystal interface, which provided the methods and conditions for the subsequent detection of DNA hybridization.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TP212;Q523
本文编号：2257041
[Abstract]:In recent years, DNA detection has been widely used in clinical diagnosis, gene expression, agriculture, food safety and environmental monitoring, so it is necessary to detect DNA quickly, conveniently and sensitively. Traditional DNA detection methods are based on fluorescence biochip technology, surface plasmon resonance (Surface Plasmon Resonance,SPR) and so on, but these detection methods have many shortcomings and still need to be improved. In recent years, liquid crystal, a new material with unique physical and chemical properties, has attracted wide attention. Liquid crystal is a kind of material form between liquid and solid. It has properties of birefringence, elastic strain and so on. The change of the orientation of liquid crystal molecules can be transformed into optical signals, which can be observed by the naked eye, so it has attracted much attention in DNA detection. Based on the birefringence of DNA hybridization and liquid crystal, the solid-liquid crystal sensing interface was constructed, and the DNA hybridization was detected on the basis of it. In addition, the water-liquid crystal interface was constructed in this paper. The effects of different surfactants on the arrangement of liquid crystal molecules were investigated, and the effects of single strand DNA (ssDNA) on the water-liquid crystal interface were explored, which laid a foundation for the detection of DNA hybridization. The main contents are as follows: (1) ssDNA was used as the orientation agent to construct the functional substrate at the solid-liquid crystal interface. Firstly, triethoxysilyl butyraldehyde (TEA) was modified on the solid substrate surface. SsDNA was modified on the solid substrate surface by the reaction between TEA with aldehyde group and ssDNA modified with amino group. The effects of glass modification time in TEA and ss DNA surface coverage on the construction of functional substrate were investigated. (2) Detection of DNA hybridization based on solid-liquid crystal interface. The above method was used to construct a novel functional substrate for the vertical alignment of liquid crystal 4-cyano-4keto-pentyl biphenyl (5CB) induced by ssDNA. The vertical arrangement of 5CB molecules was induced by optimizing the experimental conditions, and the uniform black was observed under polarizing microscope. When the complementary ssDNA is added, the hybridization reaction will occur, and the arrangement of 5CB molecules will be disturbed. The color texture can be observed under the polarizing microscope, thus the effect of DNA detection can be achieved. Under the optimized experimental conditions (the time of modification of glass slides in TEA solution is 1 h), the optimum concentration of DNA is 20 nM. The detection limit of this method is 0.1 nM, and has good specificity and repeatability. (3) the effect of different surfactants on the orientation of 5CB molecules at the water-liquid crystal interface. Based on the interaction between surfactants and 5CB molecules, different surfactants (octadecyl trimethylammonium bromide, dodecyl trimethyl ammonium bromide, octyl trimethylammonium bromide) at the water-liquid crystal interface were studied. The effect of sodium dodecyl sulfate on the orientation of 5CB molecules was investigated. The optimum concentration and response time of each surfactant to induce the vertical alignment of 5CB molecules, and the effects of the same surfactants with different chain length on the orientation of 5CB molecules were investigated. And the effects of different surfactants with the same chain length on the orientation of 5CB molecules. In addition, we also investigated the effect of ssDNA on the orientation of 5CB molecules at the water-liquid crystal interface, which provided the methods and conditions for the subsequent detection of DNA hybridization.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TP212;Q523
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