ELECTROCHEMICAL DNA-BASED SENSORS

基于电化学 DNA 的传感器

基本信息

批准号：
6087204
负责人：
JACQUELINE K BARTON
金额：
$ 25.78万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-03-01 至 2004-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6087204
关键词：
DNA binding protein analytical method bioengineering /biomedical engineering biosensor device biotherapeutic agent electrochemistry electrodes electron transport gene mutation microarray technology microelectrodes molecular stacking nucleic acid sequence p53 gene /protein phosphoester ligase protein sequence

项目摘要

The proposed program has as its goal the design and application of DNA- based electrochemical sensors. Electrodes modified with DNA oligonucleotide duplexes and a non-covalently bound intercalator as redox probe will be constructed as sensors for single base mismatches. The approach is not based fundamentally on differential hybridization ir a molecular recognition event. Instead these DNA-based sensors rely upon DNA-mediated charge transport and its sensitivity to perturbations in DNA stacking. Preliminary results have established the individual elements necessary to construct a sensor for mutational analysis: 9i) different single base-mismatches can be detected electrochemically and the sequence composition of the film can be varied substantially; (ii0 single-stranded DNAs can be assayed with repeated cycling of hybridization and denaturation on the electrode; significantly, a high level of sensitivity and discrimination for single base mismatches are achieved by coupling of the DNA-mediated charge transported to an electrocatalytic cycle. It is proposed first that the scope and sensitivity associated with this design can be systematically examined, where oligonucleotide length, sequence, and intercalator probe are varied; additionally variations in linker length and the construction of mixed surfaces will be carried out. These studies will enable not only the development of mismatch probes but also of new electrochemical probes for DNA-binding proteins and small molecules. These DNA films will be utilized to develop electrochemical assays for DNA-binding proteins, for example base flipping enzymes, based upon their perturbations to DNA stacking. The DNA-modified electrodes will also be utilized to probe the redox chemistry of DNA repair proteins, such as photolyase and Mut Y, and chemotherapeutics, such as the enediynes and anthracycline antibiotics. Finally, DNA chips will be fabricated for the analysis first of mutational hot spots in the p53 gene and thereafter for the analysis of the entire p53 gene sequence. The proposed program therefore intends to exploit DNA-mediated charge transport for the development of a completely new family of DNA-based sensors.

该计划的目标是基于DNA的电化学传感器的设计和应用。用DNA寡核苷酸双链体和非共价结合的介导子作为氧化还原探针修饰的电极将被构造为单个碱基不匹配的传感器。该方法从根本上基于差异化杂交IR分子识别事件。相反，这些基于DNA的传感器依赖于DNA介导的电荷转运及其对DNA堆叠中扰动的敏感性。初步结果已经确定了构造一个传感器进行突变分析所需的个体元素：9i）可以从电化学上检测到不同的单个基本不匹配，并且膜的序列组成可以大大变化；（II0单链DNA可以通过反复的杂交和变性的循环进行测定；在显着，通过耦合将DNA介导的电荷耦合到电催化循环的DNA介导的电荷来实现单个碱基不匹配的高灵敏度和歧视。首先，可以系统地检查与此设计相关的范围和敏感性，其中寡核苷酸的长度，序列和嵌入量探针在接头长度上有所不同；只有不匹配探针的发展，但也将使用新的结合蛋白和小分子的电化学探针来开发这些DNA膜。 DNA修饰的电极也将用于探测DNA修复蛋白的氧化还原化学，例如光溶解酶和mut Y，以及化学治疗剂，例如eneDiynes和antallacyclace抗生素。最后，将制造DNA芯片，用于在p53基因中的突变热点进行分析，然后以分析整个p53基因序列。因此，该计划旨在利用DNA介导的电荷运输来开发全新的基于DNA的传感器。