The bacteroides group is a subdivision in the <I>Cytophaga-Flavobacterium-Bacteroides</I> phylum. This group is as phylogenetically distinct from other Gram-negative enterics, including <I>Escherichia coli</I>, as they are from Gram-positive organisms. Furthermore, there is no cross expression between genes of <I>E. coli</I> and <I>Bacteroides</I> species. It is thought that this difference in gene expression lies in part at the level of transcription initiation and is due to the sequences within the promoter region itself. A putative consensus sequence for <I>Bacteroides</I> promoters has been published by C. Jeff Smith’s research group based on alignments of the sequences upstream of certain regulated genes. However, this consensus has not been found within all putative <I>Bacteroides</I> promoters. In this study, the promoter structure and function of a strong housekeeping <I>B. thetaiotaomicron</I> 16S rRNA promoter was examined and compared to an <I>E. coli</I> 16S rRNA promoter. Our hypothesis is that there are significant differences between the promoters of these two organisms. Analysis of <I>B. thetaiotaomicron</I> sequence upstream of the 16S rRNA gene has revealed the same overall structure known for <I>E. coli</I> 16S rRNA promoters in that there are two putative promoters separated by approximately 150 bp. However, the <I>B. thetaiotaomicron</I> 16S rRNA promoter contains the proposed <I>Bacteroides</I> —7 and —33 consensus sequences instead of the well known <I>E. coli</I> —10 and —35 consensus sequences. The biological activity of the<I> B. thetaiotaomicron</I> 16S rRNA full-length promoter was confirmed using a <I>Bacteroides lux</I> reporter system. A newly designed <I>Bacteroides lux</I> reporter was used to analyze specific regions of the <I>B. thetaiotaomicron</I> 16S rRNA promoter. In addition, by pairing the <I>B. thetaiotaomicron</I> 16S rRNA promoter with an <I>E. coli</I> ribosomal binding site, and vice-versa, the improved <I>lux</I> reporter was used to further confirm that the difference in gene expression between the two species lies at the level of transcription in <I>E. coli</I>. In <I>Bacteroides</I>, however, transcription and translation may work together to create a barrier to efficient gene expression of foreign genes. </P> / Master of Science
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/42962 |
| Date | 13 June 2003 |
| Creators | Thorson, Mary Leah |
| Contributors | Biology, Stevens, Ann M., Chen, Jiann-Shin, Melville, Stephen B. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
| Relation | Thesis2.PDF |
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