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A low molecular weight artificial RNA of unique size with multiple probe target regionsArtificial RNAs (aRNAs) containing novel sequence segments embedded in a deletion mutant of Vibrio proteolyticus 5S rRNA have previously been shown to be expressed from a plasmid borne growth rate regulated promoter in E. coli. These aRNAs accumulate to high levels and their detection is a promising tool for studies in molecular microbial ecology and in environmental monitoring. Herein a new construct is described which illustrates the versatility of detection that is possible with aRNAs. This 3xPen aRNA construct carries a 72 nucleotide insert with three copies of a unique 17 base probe target sequence. This aRNA is 160 nucleotides in length and again accumulates to high levels in the E. coli cytoplasm without incorporating into ribosomes. The 3xPen aRNA illustrates two improvements in detection. First, by appropriate selection of insert size, we obtained an aRNA which provides a unique and hence, easily quantifiable peak, on a high resolution gel profile of low molecular weight RNAs. Second, the existence of multiple probe targets results in a nearly commensurate increase in signal when detection is by hybridization. These aRNAs are naturally amplified and carry sequence segments that are not found in known rRNA sequences. It thus may be possible to detect them directly. An experimental step involving RT-PCR or PCR amplification of the gene could therefore be avoided.
Document ID
20040089518
Acquisition Source
Headquarters
Document Type
Reprint (Version printed in journal)
Authors
Pitulle, C.
(University of Houston Texas, United States)
Dsouza, L.
Fox, G. E.
Date Acquired
August 21, 2013
Publication Date
January 1, 1997
Publication Information
Publication: Systematic and applied microbiology
Volume: 20
Issue: 1
ISSN: 0723-2020
Subject Category
Exobiology
Funding Number(s)
CONTRACT_GRANT: R821205-010
CONTRACT_GRANT: 3652-978
CONTRACT_GRANT: NAGW-2108
Distribution Limits
Public
Copyright
Other
Keywords
Non-NASA Center
NASA Discipline Exobiology

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