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DNA aptamers for common buffer molecules: possibility of buffer interference in SELEX.
Yuzhe Ding1, Ziyu Zhang1, Yunus A Kaiyum2
1Department of Chemistry, Waterloo Institute for Nanotechnology, University of Waterloo, Waterloo, Ontario, N2L 3G1, Canada. liujw@uwaterloo.ca.
Organic & Biomolecular Chemistry
|September 24, 2024
Summary
Researchers developed aptamers that bind to Tris buffer, a common laboratory chemical. However, attempts to create aptamers for HEPES buffer were unsuccessful, highlighting potential challenges in aptamer selection experiments.
Area of Science:
- Biochemistry
- Molecular Biology
- Biotechnology
Background:
- Aptamer selection experiments typically use buffer molecules at high concentrations (10-50 mM) compared to target molecules (low μM).
- This concentration difference raises concerns about the potential for aptamers to bind buffer molecules, especially in failed selection experiments.
Purpose of the Study:
- To investigate the feasibility of selecting aptamers against common buffer molecules.
- To assess the potential for buffer-binding aptamers to interfere with aptamer selection outcomes.
Main Methods:
- Aptamer selection was performed using Tris and HEPES as target molecules.
- Binding affinity of the selected Tris aptamer was characterized using Thioflavin T (ThT) fluorescence spectroscopy and isothermal titration calorimetry (ITC).
- Nuclear Magnetic Resonance (NMR) spectroscopy was employed to confirm aptamer-target binding.
Main Results:
- Aptamers specific for Tris buffer were successfully isolated.
- The Tris aptamer exhibited dissociation constants (Kd) of 2.9 mM (ThT) and 43 μM (ITC).
- Attempts to generate aptamers for HEPES buffer were unsuccessful.
Conclusions:
- It is possible to select aptamers that bind to buffer molecules like Tris.
- The study highlights the importance of considering buffer composition in aptamer selection protocols.
- Recommendations are provided regarding the use of specific buffers to mitigate potential selection biases.
