Rapid isolation of high-affinity DNA aptamers for small-molecule targets via nuclease-assisted SELEX

在化学系化学测量与成像项目的支持下，佛罗里达国际大学（FIU）化学与生物化学系的Xiao Yi教授和她的研究团队寻求开发一种广泛可用的方法，用于快速生成基于dna的生物受体，称为适体，它可以选择性地识别小分子靶标。适配体作为点用小分子生物传感探测器的关键传感组件，通过适配体与目标分子的结合来识别目标分子，这使得它们在科学、临床和法医应用中具有潜在的价值。然而，目前的适体生成方法是劳动密集型的，耗时的，并且通常产生的适体不能清楚地区分相似形状和大小的分子之间的差异。该方法产生了具有优异识别性能的高质量适配体，并显著加快了适配体的分离过程。此外，该技术可能取代现有的适体生成方法。在开展这些研究工作的同时，肖教授还与面向拉美裔学生和传统黑人学生的学院和大学以及国际金融大学的学生培训项目合作，计划开展多层次的外联工作，重点关注代表性不足的少数族裔学生。这些活动将为学生提供参与科学研究的机会，并鼓励他们在科学、技术、工程和数学（STEM）领域从事职业。为了实现这一目标，肖博士举办科学研讨会和演讲，组织暑期科学营，并提供暑期实习。利用皮瓣内切酶1 （FEN1）的高结构特异性和高效率，肖教授和她的研究小组寻求开发一种创新的、可推广的核酸酶辅助核酸适体分离技术，称为NA-SELEX，用于快速分离高亲和力、小分子结合的核酸适体。为了建立该技术，首先使用模型SELEX系统系统地研究和优化基于fen1的链分离。然后，通过分离具有临床和法医相关性的不同小分子靶标的高亲和力适配体，证明了NA-SELEX的可行性和普遍性。最后，通过制造用于生物样品中分析物检测的电化学适配体传感器来评估所得适配体的传感能力。这种新方法有可能根据任何小分子靶标的需求有效地产生高亲和力、可定制的适配体，并极大地促进了用于检测小分子的传感元件的发展，这些小分子检测用于各种应用，如食品安全、环境监测、治疗治疗和医疗诊断。此外，FIU的教育推广计划增加了研究生，本科生和高中URM学生对STEM领域的接触，并为从事STEM的URM提供了一个包容的环境。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

With support from the Chemical Measurement and Imaging Program in the Division of Chemistry, Professor Yi Xiao and her research team in the Department of Chemistry and Biochemistry at Florida International University (FIU) seek to develop a widely usable method for the rapid generation of DNA-based bioreceptors, termed aptamers, which selectively recognize small molecules targets. Aptamers have shown promise as key sensing components for point-of-use, small-molecule biosensing detectors that recognize target molecules by aptamer binding to the targets, making them potentially valuable for scientific, clinical, and forensic applications. However, current methods for aptamer generation are labor intensive, time consuming, and typically yield aptamers that are unable to clearly tell the difference between molecules of similar shape and size. The proposed approach produces high-quality aptamers with superior recognition properties and significantly accelerates the aptamer isolation process. Moreover, this technique might replace existing methods for aptamer generation. In parallel with these research efforts, Professor Xiao, in partnership with Hispanic-serving and historically black colleges and universities, as well as student training programs at FIU, plans a multi-level outreach effort focusing on underrepresented minority (URM) students. These activities will provide students with opportunities to participate in scientific research and encourage them to pursue careers in science, technology, engineering, and mathematics (STEM) fields. In pursuit of this, Dr. Xiao gives science seminars and presentations, organizes summer science camps, and provides summer internships. Leveraging the high structural specificity and efficiency of flap endonuclease 1 (FEN1), Professor Xiao and her research team seek to develop an innovative and generalizable nuclease-assisted aptamer isolation technique, termed NA-SELEX, for the rapid isolation of high-affinity, small-molecule-binding aptamers. To establish this technology, a model SELEX system is first used to systematically study and optimize FEN1-based strand separation. The feasibility and generalizability of NA-SELEX is then demonstrated by isolating high-affinity aptamers for different small-molecule targets with clinical and forensic relevance. Finally, the sensing capabilities of the resulting aptamers is assessed by fabricating electrochemical aptamer-based sensors for analyte detection in biological samples. This new approach has the potential to efficiently generate high-affinity, customizable aptamers on demand for any small-molecule target and greatly facilitate the development of sensing elements for the detection of small molecules for various applications such as food safety, environmental monitoring, therapeutic treatment, and medical diagnostics. In addition, an educational outreach program at FIU increases the exposure of graduate, undergraduate, and high school URM students to STEM fields and promotes an inclusive environment for URMs engaged in STEM.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.