This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Pitulle, C. Articles by Fox, G. E. Search for Related Content PubMed PubMed Citation Articles by Pitulle, C. Articles by Fox, G. E. Agricola Articles by Pitulle, C. Articles by Fox, G. E.

 Previous Article  |  Next Article 

Appl. Environ. Microbiol., Oct 1995, 3661-3666, Vol 61, No. 10
Copyright © 1995, American Society for Microbiology

A novel approach for monitoring genetically engineered microorganisms by using artificial, stable RNAs

C Pitulle, KO Hedenstierna and GE Fox
Department of Biochemical and Biophysical Sciences, University of Houston, Texas 77204-5934, USA.

Further improvements in technology for efficient monitoring of genetically engineered microorganisms (GEMs) in the environment are needed. Technology for monitoring rRNA is well established but has not generally been applicable to GEMs because of the lack of unique rRNA target sequences. In the work described herein, it is demonstrated that a deletion mutant of a plasmid-borne Vibrio proteolyticus 5S rRNA gene continues to accumulate to high levels in Escherichia coli although it is no longer incorporated into 70S ribosomes. This deletion construct was subsequently modified by mutagenesis to create a unique recognition site for the restriction endonuclease BstEII, into which new sequences could be readily inserted. Finally, a novel 17-nucleotide identifier sequence from Pennisetum purpureum was embedded into the construct to create an RNA identification cassette. The artificial identifier RNA, expressed from this cassette in vivo, accumulated in E. coli to levels comparable to those of wild-type 5S rRNA without being seriously detrimental to cell survival in laboratory experiments and without entering the ribosomes. These results demonstrate that artificial, stable RNAs containing sequence segments remarkably different from those present in any known rRNA can be designed and that neither the deleted sequence segment nor ribosome incorporation is essential for accumulation of an RNA product.

This article has been cited by other articles:

• Tucker, D. L., Karouia, F., Wang, J., Luo, Y., Li, T.-B., Willson, R. C., Fofanov, Y., Fox, G. E. (2005). Effect of an Artificial RNA Marker on Gene Expression in Escherichia coli. Appl. Environ. Microbiol. 71: 4156-4159 [Abstract] [Full Text]