Single‐Round DNA Aptamer Selection by Combined Use of Capillary Electrophoresis and Next Generation Sequencing: An Aptaomics Approach for Identifying Unique Functional Protein‐Binding DNA Aptamers. Issue 39 (10th June 2021)

• Record Type: Journal Article
• Authors: Saito, Shingo; Sakamoto, Toshiki; Tanaka, Naoki; Watanabe, Ryo; Kamimura, Takuya; Ota, Kazuki; Riley, Kathryn R.; Yoshimoto, Keitaro; Tasaki‐Handa, Yuiko; Shibukawa, Masami
• Is Part Of: Chemistry. Volume 27:Issue 39(2021)
• Page Start: 10058
• Page End: 10067
• Publication Date: 2021-06-10
• Subjects: capillary electrophoresis -- data mining -- DNA aptamer selection -- next generation sequencing -- selection rule; Chemistry -- Periodicals; 540
• DOI: 10.1002/chem.202100177
• Languages: English
• ISSNs: 0947-6539

Abstract

Abstract: In DNA aptamer selection, existing methods do not discriminate aptamer sequences based on their binding affinity and function and the reproducibility of the selection is often poor, even for the selection of well‐known aptamers like those that bind the commonly used model protein thrombin. In the present study, a novel single‐round selection method (SR‐CE selection) was developed by combining capillary electrophoresis (CE) with next generation sequencing. Using SR‐CE selection, a successful semi‐quantitative and semi‐comprehensive aptamer selection for thrombin was demonstrated with high reproducibility for the first time. Selection rules based on dissociation equilibria and kinetics were devised to obtain families of analogous sequences. Selected sequences of the same family were shown to bind thrombin with high affinity. Furthermore, data acquired from SR‐CE selection was mined by creating sub‐libraries that were categorized by the functionality of the aptamers (e. g., pre‐organized aptamers versus structure‐induced aptamers). Using this approach, a novel fluorescent molecular recognition sensor for thrombin with nanomolar detection limits was discovered. Thus, in this proof‐of‐concept report, we have demonstrated the potential of a "DNA Aptaomics" approach to systematically design functional aptamers as well as to obtain high affinity aptamers. Abstract : DNA Aptaomics is proposed for finding functional DNA aptamers by a semi‐comprehensive and quantitative single‐round selection using capillary electrophoresis and next generation sequencing according to a selection rule based on analogous sequences. By mining sequence data obtained from sub‐libraries with different DNA conformations, structure‐induced (FRET type) aptamers for thrombin were reasonably selected for the first time.