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Microbial detection with low molecular weight RNAThe need to monitor microorganisms in the environment has increased interest in assays based on hybridization probes that target nucleic acids (e.g., rRNA). We report the development of liquid-phase assays for specific bacterial 5S rRNA sequences or similarly sized artificial RNAs (aRNAs) using molecular beacon technology. These beacons fluoresce only in the presence of specific target sequences, rendering as much as a 27-fold fluorescence enhancement. The assays can be used with both crude cell lysates and purified total RNA preparations. Minimal sample preparation (e.g., heating to promote leakage from cells) is sufficient to detect many Gram-negative bacteria. Using this approach it was possible to detect an aRNA-labeled Escherichia coli strain in the presence of a large background of an otherwise identical E. coli strain. Finally, by using a longer wavelength carboxytetramethylrhodamine beacon it was possible to reduce the fraction of the signal due to cellular autofluorescence to below 0.5%.
Document ID
20040088677
Acquisition Source
Legacy CDMS
Document Type
Reprint (Version printed in journal)
External Source(s)
Authors
Kourentzi, K. D.
(University of Houston 4800 Calhoun Avenue, Houston, TX 77204-4004, United States)
Fox, G. E.
Willson, R. C.
Date Acquired
August 21, 2013
Publication Date
December 1, 2001
Publication Information
Publication: Current microbiology
Volume: 43
Issue: 6
ISSN: 0343-8651
Subject Category
Life Sciences (General)
Report/Patent Number
ISSN: 0343-8651
Distribution Limits
Public
Copyright
Other
Keywords
NASA Discipline Life Sciences Technologies
Non-NASA Center
Evaluation Studies
RNA, Ribosomal, 5S/genetics
Gram-Negative Bacteria/classification/genetics/isolation & purification
RNA, Bacterial/genetics
Environmental Microbiology
RNA Probes/genetics
Support, Non-U.S. Gov't
Biological Markers
Nucleic Acid Hybridization
Support, U.S. Gov't, Non-P.H.S
Molecular Weight

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