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Development of Aptamers for RNase Inactivation in Xtract-Free™ Sample Collection and Transport Medium
Luke T Daum1, John D Rodriguez1, James P Chambers2
1LuJo BioScience Laboratory, San Antonio, TX 78209, USA.
Diagnostics (Basel, Switzerland)
|June 27, 2024
Summary
New DNA aptamers were developed to neutralize ribonuclease (RNase) enzymes, preserving RNA integrity in self-collection kits. This innovation enhances genetic material stability for home-based and point-of-care diagnostic testing.
Area of Science:
- Biotechnology
- Molecular Biology
- Genomics
Background:
- RNA is highly susceptible to degradation by ribonuclease (RNase) enzymes.
- Current sample collection methods often require cold chain or immediate processing, limiting accessibility.
- There is a need for stable, user-friendly sample collection mediums for genetic analysis.
Purpose of the Study:
- To develop novel DNA aptamers targeting RNase A.
- To integrate these aptamers into an extraction-free collection medium for RNA preservation.
- To evaluate the efficacy of aptamer-mediated RNase inactivation.
Main Methods:
- Systematic Evolution of Ligands by Exponential Enrichment (SELEX) was employed to generate anti-RNase DNA aptamers.
- Aptamers were selected using paramagnetic beads and stringent conditions, including a novel collection medium (Xtract-Free™) and elevated temperature.
- Next-generation sequencing and bioinformatic analysis identified five candidate aptamers.
- RNA integrity was assessed using fluorometric and real-time RT-PCR assays to quantify RNase activity.
Main Results:
- Five DNA aptamers with anti-RNase activity were successfully generated and identified.
- Inclusion of aptamers reduced RNase activity by 8,800- to 11,200-fold, significantly protecting viral RNA from digestion.
- The developed aptamers maintained efficacy under conditions simulating home-collection scenarios.
Conclusions:
- Anti-RNase DNA aptamers can effectively inactivate RNase enzymes, preserving RNA integrity.
- These aptamers are suitable for integration into extraction-free sample collection mediums and diagnostic kits.
- This technology holds significant potential for improving the quality and accessibility of RNA-based genomic analyses, particularly for point-of-care and mail-back testing.
