基于磁性微球的无标记化学发光端粒传感新技术

Magnetic Bead-based Label-free Chemiluminescence Detection of Specific DNA Sequence—Telomere

近年来无标记型DNA传感技术的研究已成为病原基因测定和基因疾病诊断等领域新的研究热点之一.基于磁性微球分离和富集的方法,建立了一种新型的无标记化学发光检测技术,并成功地应用于特定序列DNA——端粒的检测.首先采用dT20修饰的磁性微球,与连接有dA20的捕获探针DNA杂交,然后再与端粒进行第二步杂交反应.磁性分离洗涤后,利用端粒中富含的G碱基与3,4,5-三甲氧基苯乙二醛反应产生特异性化学发光,从而实现特定序列DNA——端粒的无标记检测.实验结果表明该法具有操作简便、分析快速、灵敏度高、专属性好等特点.目标DNA浓度在5×10-9～1×10-7mol/L浓度范围内具有良好的线性关系,相关系数为0.9918.

The label-free DNA sensing techniques are currently receiving considerable attention for the determination of pathogenic genes and the diagnosis of genetic disease, etc. Based on magnetic separation and preconceneration, a novel label-free chemiluminescence (CL) detection protocol was presented as exemplified by the successful determination of specific DNA sequence-telomere. This method was composed of three steps: (1) the dT(20) arm modified in the magnetic beads was hybridized with the dA(20)-modified capture DNA; (2) the first step was followed by the second hybridization of the bead-captured probe with telomere; (3) the amount of telomere was determined by the guanine CL reaction with 3,4,5-trimethoxyphenylglyoxal (TMPG). The results showed that this simple and sensitive protocol was much suitable for the determination of specific DNA sequence such as telomere with a good linear correlation in the concentration range of 5 X 10(-9)similar to 1 X 10(-7) mol/L (R-2=0.9918).