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Unexpected detection of DNA by nucleic acid sequence-based amplification technique
David Rodríguez-Làzaro1, Joy Lloyd, John Ikonomopoulos
1Institute of Food and Agricultural Technology (INTEA), University of Girona, Campus Montilivi s/n, E-17071 Girona, Spain.
Molecular and Cellular Probes
|July 24, 2004
Summary
Nucleic acid sequence-based amplification (NASBA) can detect DNA in some bacteria, not just RNA. Researchers found that enzymatic treatments are crucial to confirm if NASBA targets DNA or RNA for accurate microbial detection.
Area of Science:
- Microbiology
- Molecular Biology
- Biotechnology
Background:
- Nucleic acid sequence-based amplification (NASBA) is a method for detecting RNA.
- Accurate identification of microbial targets relies on confirming the nucleic acid type.
Purpose of the Study:
- To investigate the nucleic acid specificity of NASBA in different bacterial species.
- To determine the necessity of enzymatic treatments for validating NASBA results.
Main Methods:
- Extraction of nucleic acids from Salmonella enterica serotype Typhimurium and Mycobacterium avium subsp. paratuberculosis.
- Application of enzymatic treatments: DNase, RNase, S1 nuclease, and combined RNase/S1 nuclease.
- Detection of nucleic acids using NASBA.
Main Results:
- For S. enterica Typhimurium, RNase and RNase/S1 nuclease treatments eliminated the NASBA signal, confirming RNA detection.
- For M. avium subsp. paratuberculosis, DNase treatment abolished the NASBA signal, while RNase, S1, and RNase/S1 treatments did not affect it.
- This indicates NASBA detected DNA in M. avium subsp. paratuberculosis.
Conclusions:
- NASBA's ability to detect DNA varies between bacterial species.
- Enzymatic pre-treatment is essential to verify the nucleic acid target (DNA or RNA) for reliable NASBA-based microbial detection.