导　　师： 李毅群

学科专业： 070302

授予学位： 硕士

作　　者： ;

机构地区： 暨南大学

摘　　要： DNA传感器是一类重要的DNA检测手段，近年来由于发展迅速受到了广泛的关注。DNA传感器以DNA为敏感元件，DNA固定在用作换能器的电极上，并通过电极将DNA与DNA、RNA、药物、自由基等相互作用所产生的生物学信号转换成光、电、声波等物理学信号。DNA传感器的应用范围十分广泛，可用于疾病检测、环境监测、药物筛选、法医鉴定等多个领域。 艾滋病HIV-1的传统诊断方法由于存在步骤复杂，需用放射性同位素以及分析时间过长等缺陷，不适于常规环境。本论文以糊电极为基底电极，研制成电化学传感器，建立了一种灵敏度高，方法简单，能快速定量检测与HIV-1有关的特定序列目的基因的方法，为建立核酸探针电化学传感器进行基因诊断提供了依据。 利用一定电压下将探针固定在活化的碳糊电极表面的技术，研制成DNA电化学传感器，用该传感器对互补链DNA进行杂交检测。在分析的过程中对探针的固定和杂交条件进行了探索。实验得出2min为最佳固定时间。同时杂交时间对电化学信号有较大影响。随着杂交时间的增加，电极上指示剂的还原峰电流差迅速增加，20min后增加趋缓。杂交液中NaCl的浓度和杂交电压对电化学信号有较大影响。NaCl的浓度在0.1～0.5mol//L范围内电化学信号迅速增加；0.5～0.75mol//L范围内，电化学信号增加缓慢。用该传感器，分别在0.1、0.25、0.75、1.0和0.5v电压下杂交。研究结果表明：0.5v为最佳杂交电压。同时比较了0.5v电压下和无电压时的杂交效率，无电压时杂交2小时，其杂交量几乎相当于0.5v电压下杂交20min。 该传感器能对与HIV-1有关的DNA片段进行识别和鉴定。当互补链DNA浓度在1.25×10~/(-7/)-5.00×10~/(-7/)mol//L范围内，其浓度与电化学信号大小呈一次线性关系。其检测下限40ng//ml。 DNA sensor is one of the important detective means and noted widely in recent years for rapid development.DNA sensor takes DNA as sensitive element by immobilizing DNA on the transduser.lt can convert biological signals into physical signals such as light,electrical,acoustic etc. by DNA on the surface of electrode interaction with DNA,RNA,drug and radicals.DNA sensor is widely used in many fields such as illness detection,enviornment monitoring,drug screening, forensic identifying. Traditional diagnostic method on HIV-1 is not suitable for routin environment because of radioisotopes,complex steps and long assay times.We manufactured a kind of high-sensitive DNA electrochemical sensor based on carbon paste electrode which can quantify DNA sequences related to HIV-1 simply in short time.This forms the bases for gene diagnosis on nucleic acid probe electrochemical biosensor. The DNA electrochemical sensor is manufactured by immobilizing ssDNA on the activated surface of CPE in fixed potential.The complementary DNA is detected by hybridization on the this sensor.The conditions on probe immobilizing and hybridizing are studied during analysis.We concluded 2min is the optimal immobilizing time.Hybridization time has great effect on the response.With the hybridization time increasing, difference of reductive peak current of indicator on the electrode increases sharply.Then it increases slowly in 20 minutes.The concentration of NaCl in hybridization solution and hyridizayion potential influence electrochemical response greatly.The concentration of NaCl being between 0.1 and 0.5mol//L,the response increase quickly. It increases slowly when NaCl's concentration is between 0.5 and 0.75mol//L.If this sensor hybridizes in 0.1,0.25,0.75,1.0 and 0.5v respectively,the study show 0.5v is optimal hybridization voltage.Hybridization efficiency without voltage comparing with that in 0.5v,the hybri dization quantities for two hours is almost the same as that for 20 minutes in 0.5v. This sen

关 键 词： 电化学传感器 探针 杂交 基因诊断

领　　域： [自动化与计算机技术] [自动化与计算机技术]