基于富勒烯衍生物修饰玻碳电极的DNA电化学传感器

Electrochemical DNA Biosensor Based on the Immobilization of Probe DNA on a Fulleropyrrolidine Derivative Modified Glassy Carbon Electrode

以富勒烯C60,L-苏氨酸及对苯二甲醛为原料,在氮气保护下反应得到含醛基官能团的2-(4-醛基苯基)-5-(1-羟乙基)富勒烯吡咯烷衍生物(C60-CHO).将该材料修饰于玻碳电极表面,并利用醛基与氨基之间温和、高效的缩合反应,将5'-氨基修饰的寡聚核苷酸共价固定到了C60-CHO修饰的玻碳电极表面,构建了一种新型的电化学DNA传感器.以[Fe(CN)6]3?/4?为电活性探针,采用电化学阻抗法对转基因植物CaMV35S启动子基因特征片段进行检测.实验结果表明,杂交前后的电子传递电阻差值(ΔRet)与目标序列浓度对数(lg CS2)在1.0×10-13～1.0×10-9 mol/L浓度范围内呈良好的线性关系,线性回归方程为ΔRet/(103?)=3.471 lg(CS2/mol/L)+50.425(r=0.9977),检测限为1.5×10?14 mol/L.杂交特异性实验进一步表明该传感器对完全互补、碱基错配和非互补序列具有良好的识别能力.

An aldehyde-containing fulleropyrrolidine derivative,2-（4-formylphenyl）-5-（1-hydroxyethyl）-fulleropyrrolidine（C60-CHO）,was synthesized by the reaction of C60 with L-threonine and terephthalalde-hyde in refluxing toluene under a nitrogen atmosphere.To fabricate a novel DNA biosensor,the C60-CHO was cast on the surface of a glassy carbon electrode and grafted with 5＇-amino modified probe DNA through a facile amine-aldehyde condensation reaction.When the oligonucleotides related to CaMV35S promoter fragment were determined with the developed biosensor via electrochemical impedance spectroscopy using [Fe（CN）6]3？/4？ as indicating probe,the difference values of the electron transfer resistance（？Ret） before and after hybridization showed good linear relationship with logarithm of the target-sequence concentrations in the range from 1.0×10？13 to 1.0×10？9 mol/L with the regression equation of ？Ret/（103 Ω）=3.471 lg（CS2/mol/L）＋50.425.The detection limit of 1.5×10？14 mol/L was also obtained.In addition,the hybridiza-tion specificity experiments showed that the sensing system can accurately discriminate the complementary sequence from the mismatched and the noncomplementary sequences.