Characterization of the structure and function of a <I>Bacteroides thetaiotaomicron</I> 16S rRNA promoter

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&#146;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> &#151;7 and &#151;33 consensus sequences instead of the well known <I>E. coli</I> &#151;10 and &#151;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>

Name thesis:degree_name

Master of Science

Level thesis:degree_level

masters

Discipline thesis:degree_discipline

Biology

Department dc:contributor.department

Biology

Grantor dc:publisher

Virginia Tech

Year dc:date.issued

2003

Author dc:creator

• Thorson, Mary Leah

Chair dc:contributor.committeechair

• Stevens, Ann M.

Committee members dc:contributor.committeemember

• Chen, Jiann-Shin
• Melville, Stephen B.

• rRNA operon
• promoter
• gene reporter
• gene expression
• Bacteroides

Statement dc:rights

• In Copyright

Licence dc:rights.uri

• http://rightsstatements.org/vocab/InC/1.0/

Dc Identifier Other

etd-06092003-130248

OAI identifier oai:identifier

oai:vtechworks.lib.vt.edu:10919/42962