DNA电化学生物传感器在转基因植物特定序列基因检测中的应用

摘要

在玻碳电极(GCE)上采用循环伏安法电聚合硫堇(PTh)得到PTh/GCE修饰电极,并利用聚硫堇层共价结合和静电作用吸附金纳米粒子(AuNP's)制得AuNP's/PTh/GCE修饰电极.然后通过将ss-DNA/AuNP's/PTh修饰电极置于cDNA杂交液中,于42℃杂交制得ds-DNA/AuNP's/PTh修饰玻碳电极,实现了脱氧核糖核酸(DNA)探针在AuNP's/PTh修饰的玻碳电极上的固定,制得DNA电化学生物传感器.在[Fe(CN)6]3-/4-溶液中采用微分脉冲伏安法(DPV)及交流阻抗谱技术(EIS)对DNA的固定和杂交进行了表征.试验结果表明:在1.0×10-10～1.0×10-6mol·L-1的浓度范围内,该传感器可对转基因植物外源基因草丁膦乙酰转移酶基因(PAT基因)片段进行检测,检出限(3s)为3.2×10-11mol·L-1.%The glassy carbon electrode (GCE) was modified with polythionine (PTh) to form PTh/GCE by electropolymerization of thionine with cyclic voltammetry, and nanoparticles of gold were adhered to the PTh/GCE through coordination and electrostatic action of the PTh layer to form the modified electrode of AuNP's/PTh/GCE.Finally, DNA probe was immobilized onto the AuNP' s/PTh/GCE through the interaction between ss-DNA and nano-gold to form ss-DNA/AuNP's/PTh/GCE, which was further hydridized in cDNA solution at 42 ℃ to form dsDNA/AuNP's/PTh/GCE.The immobilization and hydridization of DNA were characterized by differential pulse voltammetry and AC impedance spectroscopy in a solution of [Fe(CN)6]3-/4-.It was found that linear relationship between values of detection signals (△Ret) and concentration of the 20-base PAT genetic sequence was obtained in the range of 1.0 × 10-10- 1.0 × 10-6 mol · L-1.Detection limit (3s) of this biosensor was found to be 3.2 ×10-11mol · L-1.