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United States Department of Agriculture

Agricultural Research Service

Title: Use of the Bacteroides Ovatus Xsa Gene As a Transcriptional Reporter Gene for Various Gram-Negative Anaerobes and Escherichia Coli

Authors
item Whitehead, Terence
item Cotta, Michael
item Xing, W - UNIV OF NEB LINCOLN NEB
item Morrison, M - UNIV OF NEB LINCOLN NEB

Submitted to: Microbial Ecology in Health and Disease
Publication Type: Abstract Only
Publication Acceptance Date: November 9, 1997
Publication Date: N/A

Technical Abstract: Members of the genera Bacteroides, Prevotella, and Porphyromonas are capable of causing diseases in humans. Genetic studies of these organisms are important for determining factors involved in disease development. A reporter gene for transcriptional fusions would be useful for studies of gene regulation in these organisms. A xylan -inducible operon from Bacteroides ovatus V975 contains a novel bifunctional xylosidase/arabinosidase gene (xsa) which may serve this purpose. The xsa gene was isolated by polymerase chain reaction and inserted into the Escherichia coli plasmid pBluescript. The xsa gene was determined to be under transcriptional regulation in E. coli by the lac promoter. The xsa gene was subcloned into E. coli/Bacteroides shuttle vectors and introduced into different Bacteroides species and Porphyromonas gingivalis by conjugation. The xsa gene was expressed in Bacteroides species when placed downstream of a P. ruminicola xylanase gene promoter region. The advantages of the xsa reporter system include the low background or total lack of arabinosidase and xylosidase activities in most Bacteroides species, P. gingivalis, and E. coli, and the ease of performing enzymatic assays. The xsa reporter gene is being tested to study the regulation of the glutamate dehydrogenase genes (gdhA and gdhB) of B. thetaiotaomicron.

Last Modified: 7/28/2014
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