基于瓜环主客体识别技术的新型电化学DNA传感器的构建及其应用于急性早幼粒细胞白血病的早期诊断

The electrochemically sensing technology to detect DNA hybridization in homogeneous solution based on host-guest recognition has an important value in the field of DNA biosensors. At present, the DNA biosensors reported and we constructed based on host-guest recognition and cyclodextrins as host compound, have higher efficiency of hybridization and higher sensitivity of DNA detection. But there are also some disadvantages, such as the long time of process of host-guest recognition, the unsatisfactory stability, and so on. The research include preparation of aligned carbon nanotubes array (ACNTs) electrodes, immobilization of cucurbit uril and derivatives to ACNTs electrode using a variety of methods, and then preparation of new ACNTs electrochemically DNA biosensors via host-guest recognition based on cucurbit uril as host compound, to realize chemically DNA sensing to detect DNA hybridization in homogeneous solution. It expected to be higher efficiency, stability, sensitivity, and specificity of our prepared biosensors. Based on this, new electrochemical biosensors based nanoparticles and aligned carbon nanotubes double nanometer materials, and cucurbit uril as the host compound via host-guest recognition technology are constructed.With the signal amplifying function of two kinds of nanometer materials, the sensitivity of biological sensors will further improved. The new electrochemical DNA biosensors prepared will be used in the early diagnosis of acute promyelocytic leukemia.The research of the project will provide the experimental basis for the application and development of cucurbit uril in the field of electrochemically DNA biosensors.

基于主客体识别技术实现溶液均相DNA杂交的传感技术在电化学DNA传感器领域具有重要的应用价值。目前报道的和我们构建的环糊精为主体的主客体识别DNA传感器，可提高DNA杂交效率和检测灵敏度。但还存在主客体识别时间长，稳定性不理想等问题。课题将制备直立碳纳米管阵列（ACNTs）电极；采用多种方法固定瓜环及衍生物至ACNTs电极；应用瓜环为主体的主客体识别技术构建新型直立碳纳米管阵列电化学DNA生物传感器，以实现溶液均相杂交的DNA检测。预期得到更高效、更稳定、更灵敏、特异性更高的DNA生物传感器。在此基础上，进一步构建纳米颗粒和直立碳纳米管阵列双纳米材料与瓜环为主体的主客体识别技术相结合的电化学DNA生物传感器。利用两种纳米材料对信号的放大作用，进一步提高检测灵敏度。构建的电化学DNA传感器可用于急性早幼粒细胞白血病的早期检测。项目为瓜环在电化学DNA生物传感器领域的应用和发展提供实验依据。

本项目获国家自然科学基金（No.31300819）资助后，通过项目组成员的共同努力，已经完成了直立碳纳米管的制备，并利用直立碳纳米管制备了一系列的DNA生物传感器；同时以瓜环作为主体化合物，利用主客体识别技术及碳纳米管和纳米颗粒双纳米材料构建了一系列的DNA生物传感器，可以实现溶液中均相杂交的DNA检测。这些传感器具有稳定性好，特异性好及灵敏度高等特点，并将这些传感器应用于急性早幼粒细胞白血病的PML/RARα 融合基因及乳腺癌基因等疾病基因的检测。项目已取得了一系列创新性研究成果。研究内容主要包括：（1）在多壁碳纳米管的作为电极的修饰材料，构建了基于多壁碳纳米管支持的分子印迹聚合物和DNA的鸟嘌呤位点制备的可多位点识别富含鸟嘌呤的DNA的新型生物传感器。（2）实验室里成功制备了直立碳纳米管，对直立碳纳米管进行功能化修饰后，构建了基于直立碳纳米管和生物素-亲和素结合的溶液中均相杂交的DNA传感器；以及利用制备的直立碳纳米管表面溅射的大面积的金颗粒，构建了基于直立碳纳米管大面积金粒子生物传感器及急性早幼粒细胞白血病 PML/RARα 融合基因的检测。（3）以七元瓜环作为主体分子，以dabcyl作为客体分子，构建了基于碳纳米管和纳米颗粒双纳米材料及瓜环为主体的主客体识别 DNA 生物传感器，可以实现DNA溶液中的均相杂交。以及七元瓜环作为主体分子，二茂铁作为客体分子，构建了一种基于均相溶液中DNA杂交及七元瓜环和二茂铁纳米微球之间的主客体识别放大信号的一种可循环利用的用于乳腺癌诊断的电化学传感器。此外，我们还开展了基于金纳米复合粒子和主客体识别技术的新型荧光传感器的研究，并用于环境水中亚硝酸根的检测。课题实施已达到预期目标，已发表论文4篇 (包括SCI收录3篇)；申请了发明专利2篇；协助培养了2名硕士研究生。