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Optimization of RNA Aptamer SELEX Methods: Improved Aptamer Transcript 3'-End Homogeneity, PAGE Purification Yield,
Chandan Narayan1, Suresh Veeramani1,2, William H Thiel1
1Department of Internal Medicine and University of Iowa, Iowa City, Iowa, USA.
Nucleic Acid Therapeutics
|November 10, 2021
Summary
Optimizing key steps in RNA aptamer selection (SELEX) enhances efficiency. Improvements in RNA transcription, purification, and recovery yield better aptamers for diverse targets.
Area of Science:
- Molecular biology
- Biotechnology
- Biochemistry
Background:
- Systematic Evolution of Ligands by Exponential Enrichment (SELEX) is a cornerstone for discovering RNA aptamers.
- While SELEX has evolved, fundamental steps like DNA template preparation and in vitro transcription remain largely unchanged.
- These foundational steps present opportunities for optimization to improve overall SELEX efficiency.
Purpose of the Study:
- To enhance the homogeneity of in vitro transcribed aptamer RNA.
- To increase the efficiency of purifying aptamer RNA using polyacrylamide gel electrophoresis (PAGE).
- To improve the quality of target-bound aptamer RNA recovered during the SELEX process.
Main Methods:
- Focus on optimizing in vitro transcription for improved RNA homogeneity.
- Enhance polyacrylamide gel electrophoresis (PAGE) for more efficient RNA purification.
- Develop improved methods for recovering target-bound aptamer RNA.
Main Results:
- Achieved greater homogeneity in in vitro transcribed aptamer RNA.
- Increased the efficiency of aptamer RNA purification via PAGE.
- Enhanced the recovery quality of target-bound aptamer RNA.
Conclusions:
- Optimizations in RNA transcription, purification, and recovery significantly improve the SELEX process.
- These enhancements are applicable to both protein-based and cell-based RNA aptamer selections.
- The refined SELEX methodology contributes to more efficient aptamer discovery.
