DNA aptamers for common buffer molecules: possibility of buffer interference in SELEX

• Published in: Organic & biomolecular chemistry 2024-10, Vol.22 (41), p.8337-8343
• Main Authors: Ding, Yuzhe, Zhang, Ziyu, Kaiyum, Yunus A, Heng, Yicheng, Johnson, Philip E, Liu, Juewen
• Format: Article
• Language: English
• Subjects: Aptamers; Aptamers, Nucleotide - chemistry; Binding; Buffers; Calorimetry; Fluorescence spectroscopy; HEPES - chemistry; Magnetic resonance spectroscopy; NMR spectroscopy; Nucleotide sequence; SELEX Aptamer Technique; Spectrometry, Fluorescence; Spectroscopy; Titration; Titration calorimetry; Tromethamine - chemistry
• ISSN: 1477-0520; 1477-0539; 1477-0539
• DOI: 10.1039/d4ob00622d

During a typical aptamer selection experiment, buffer molecules are used at the 10 to 50 mM range, whereas target molecules could be used at much lower concentrations even in low μM levels. Therefore, doubts existed regarding the potential enrichment of buffer binding aptamers, particularly for failed selections that cannot validate binding of enriched sequences. In this study, we used two common buffer molecules, Tris and HEPES, as target molecules. While we successfully isolated aptamers for Tris buffer, our attempts to generate aptamers for HEPES buffer failed. Thioflavin T (ThT) fluorescence spectroscopy showed the dissociation constant ( K d ) of the Tris buffer aptamer to be 2.9 mM, while isothermal titration calorimetry showed a K d of 43 μM. NMR spectroscopy also confirmed aptamer binding. Finally, we discussed the implications of this buffer selection work and recommended the use of certain buffers. Using capture-SELEX, aptamers for Tris were isolated but the selection for HEPES failed, and the possibility of obtaining buffer binding aptamers during aptamer selection experiments is discussed.