华东师范大学学报(自然科学版) ›› 2006, Vol. 2006 ›› Issue (4): 39-48.

• 化学 • 上一篇    下一篇

运用交流阻抗技术直接监测DNA杂交

徐 颖, 杨 琳, 叶晓燕, 何品刚, 方禹之   

  1. 华东师范大学 化学系, 上海 200062
  • 收稿日期:2005-06-10 修回日期:2006-04-20 出版日期:2006-07-25 发布日期:2006-07-25
  • 通讯作者: 方禹之

Direct Detection of DNA Hybridization by Impedance Technique(Chinese)

XU Ying, YANG Lin, YE Xiao-yan, HE Pin-gang, FANG Yu-zhi   

  1. Department of Chemistry, East China Normal University, Shanghai 200062, China
  • Received:2005-06-10 Revised:2006-04-20 Online:2006-07-25 Published:2006-07-25
  • Contact: FANG Yu-zhi

摘要： 介绍了基于交流阻抗技术构建非标记型脱氧核糖核酸（DNA）杂交传感器的方法.以24个碱基长度的寡聚DNA作为实验对象，将5′端巯基化的单链寡聚DNA（SH-ssDNA）探针与巯基乙酸（RSH）同时自组装到金电极表面，形成杂交识别层，利用交流阻抗技术测量出杂交前后金电极表面电子传递电阻Ret的增量作为杂交信号.实验中对DNA探针的自组装时间、杂交温度、杂交时间和阻抗测量液等实验条件进行了观察和优化；通过选择自组装液中SH-ssDNA探针和RSH的浓度，减少DNA在金电极表面的非特异性吸附，同时保证金电极表面自组装的SH-ssDNA探针有合适的疏密度，提高了杂交效率.在各优化条件下，无需扩增杂交信号，此非标记型DNA杂交传感器的检测下限为3.0×10-14mol/L；和完全互补序列相比，一个和三个碱基错配序列分别产生55.6%和1.3%的杂交信号.

关键词: DNA杂交传感器, 交流阻抗, 非标记法, DNA杂交传感器, 交流阻抗, 非标记法

Abstract: A label-free DNA hybridization biosensor based on impedance technique was introduced. A 24-mer 5'-mercapto-group ended single-stranded DNA probe (SH-ssDNA) was self-assembled onto Au electrode surface with mercaptoacetic acid (RSH) to form a recognition layer. By Faradic impedance technique, a significant increase of the interfacial electronic transfer resistance Ret was determined when the SH-ssDNA probe/Au electrode hybridized with its complementary sequence. Further, several experimental conditions, including the SH-ssDNA probe immobilization time, hybridization temperature and time, the eletrolyte for impendance detections were optimized. Especially, the proportion of SH-ssDNA and RSH in the immobilization solution was important for a suitable density of ssDNA probe on electrode surface to improve the hybridization efficiency. Based on these optimized conditions, this label-free DNA hybridization biosensor has a low detection limit of 3.0×10-14 mol/L of complementary sequence in solution. When compared with the complementary sequence, the hybridization signal of one-base mismatched sequence and three-base mismatched sequence was 55.6% and 1.3% respectively.

Key words: impedance, label-free, DNA hybridization biosensor, impedance, label-free

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