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

通过核酸酶辅助 SELEX 快速分离小分子靶标的高亲和力 DNA 适体

• 批准号: 2135005
• 负责人: Yi Xiao
• 金额: $ 35.45万
• 依托单位: North Carolina State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2135005&HistoricalAwards=false
• 关键词: Rapid; isolation; affinity; DNA; aptamers

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.

在化学划分的化学测量和成像计划的支持下，伊肖教授及其研究团队在佛罗里达国际大学（FIU）的化学和生物化学系（FIU）的化学研究团队中，试图开发一种可广泛使用的基于DNA的生物疗法的可广泛使用的方法，该方法被称为适当的Aptamers，该方法被称为适合识别小分子的靶标。适体已显示出有望作为使用点的小分子生物传感探测器的关键传感成分，这些检测器通过适体与目标结合识别靶分子，从而使其在科学，临床和法医应用中具有价值。但是，适体生成的当前方法是劳动密集型，耗时的，并且通常产生的适体无法清楚地判断出相似形状和大小的分子之间的差异。所提出的方法产生具有优异识别特性的高质量适体，并显着加速适体隔离过程。此外，该技术可能会取代适合生成的现有方法。与这些研究的工作同时，Xiao教授与西班牙裔服务和历史悠久的黑人学院和大学合作，以及FIU的学生培训计划，计划一项多层外展工作，重点是代表性不足的少数民族（URM）学生。这些活动将为学生提供参与科学研究的机会，并鼓励他们从事科学，技术，工程和数学（STEM）领域的职业。为此，小博士为科学研讨会和演讲，组织夏季科学训练营，并提供暑期实习。 Xiao教授和她的研究团队利用皮瓣核酸内切酶1（FEN1）的高结构特异性和效率，试图开发一种创新且可推广的核酸酶辅助适体隔离技术，称为Na-selex，以快速隔离高无限，小物质，小分子结构的Atpamers。为了建立这项技术，模型SELEX系统首先用于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体，可以证明Na-Selex的可行性和普遍性。最后，通过制造基于电化学适体的传感器来评估所得的适体的传感能力，以在生物样品中检测分析物检测。这种新方法有可能有效地生成对任何小分子目标的高亲和力，可定制的适体需求，并极大地促进感应元素的发展，以检测用于食品安全，环境监测，治疗性治疗和医疗诊断等各种应用的小分子。此外，FIU的一项教育外展计划增加了研究生，本科和高中URM学生的曝光率，并促进了从事STEM的URMS的包容性环境。该奖项反映了NSF的法定任务，并认为通过使用该基金会的知识分子的智力和更广泛的影响来评估Criteria Criteria Criteria的评估，并被认为是值得的。

项目成果

Using Exonucleases for Aptamer Characterization, Engineering, and Sensing

• DOI: 10.1021/acs.accounts.3c00113
• 发表时间: 2023-06-14
• 期刊: ACCOUNTS OF CHEMICAL RESEARCH
• 影响因子: 18.3
• 作者: Alkhamis，Obtin;Canoura，Juan;Xiao，Yi
• 通讯作者: Xiao，Yi