Molecular Design of DNA Conjugate and its Applications to Biosensing

我们根据自己独特的原理开发了新型的生物传感技术，并提出了简便、有效和高灵敏度的分析方法。其原理在于通过使用携带DNA的纳米颗粒的非交联聚集来进行单碱基识别，所述纳米颗粒通过接枝有寡脱氧核糖核苷酸的温敏聚合物的自组装形成。本研究基于这一原理，设计了新型的DNA结合材料，包括DNA响应性水凝胶、DNA合成聚合物嵌段型结合物和DNA连接的纳米金等，作为基因诊断的分析工具。证明了DNA固定的聚苯乙烯乳胶和DNA连接的金纳米粒子的独特聚集，表明该原理在一般情况下成立. DNA纳米粒子的电泳迁移率发生了序列依赖性的剧烈变化.以单链DNA为交联剂的新型杂化水凝胶能够根据样品DNA序列的变化而收缩或膨胀，从而实现对单碱基差异的精确识别.建立了以DNA-合成聚合物嵌段型偶联物为亲和配基的亲和毛细管电泳分离单链样品DNA及其点突变体的方法.在微芯片上进行表面等离子体共振成像，用于高灵敏度检测以非交联构型沉积在表面上的DNA连接的金纳米颗粒。基于微流控通道中的亲和电泳技术，实现了基因突变的多重分析。在该研究基金的帮助下，我们成功地通过设计各种DNA缀合材料来构建生物传感系统。

We developed novel biosensing technologies on the basis of our unique principle, and presented facile, effective and high-sensitive analytical methods. The principle consists in single-base discrimination by using non-cross-linking aggregation of DNA-carrying nanoparticles, which are formed through the self-assembly of thermoresponsive polymers grafted with oligodeoxyribonucleotides. In this study, we devised new DNA-conjugated materials, including DNA-responsive hydrogels, DNA-synthetic polymer block-type conjugates, and DNA-linked gold nanoparticles, as analytical tools for gene diagnostics based on this principle.Achievements are summarized as follows:1. The unique aggregation of DNA-immobilized polystyrene Latex and DNA-linked gold nanoparticles was demonstrated, indicating that the principle would hold in general.2. Sequence-dependent, drastic change was found in electrophoretic mobility of DNA-carrying nanoparticles.3. Novel hybrid hydrogels containing rationally designed single-stranded DNA as the cross-linker were developed to shrink or swell in response to sequence of sample DNA, allowing for precise discrimination of single base difference.4. Affinity capillary electrophoresis using DNA-synthetic polymer block-type conjugates as affinity ligands was developed to separate single-stranded sample DNA and its point mutant.5. Surface plasmon resonance imaging was conducted on a microchip for high-sensitive detection of DNA-linked gold nanoparticles deposited onto the surface in a non-cross-linking configuration.6. Multiplex gene mutation analysis was realized on the basis of affinity electrophoresis in micro fluidic channels. With the aid of this research fund, we succeeded to construct biosensing systems by designing various DNA-conjugated materials.