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Author Tyler, Tina Lynn
Title Detection and characterization of urease genes in groundwater bacterial communities
book jacket
Descript 117 p
Note Source: Dissertation Abstracts International, Volume: 65-07, Section: B, page: 3292
Adviser: Peter Sheridan
Thesis (Ph.D.)--Idaho State University, 2004
Molecular methods for detecting and characterizing ureolytic bacteria were developed and tested for use in conjunction with a proposed groundwater remediation strategy for the immobilization of radionuclides by calcite precipitation. A field test of the strategy to promote in situ calcite precipitation was performed in which molasses and urea were injected into groundwater to stimulate bacterial growth and urease activity. Bacterial growth and urease expression were stimulated, resulting in enzymatic urea hydrolysis, the products of which promoted calcite precipitation. To investigate the ureolytic bacterial populations that were stimulated during this field test, oligonucleotide primers for the ureC gene were designed with an emphasis on detecting a diverse range of ureolytic bacterial species in groundwater. Primers were tested and optimized in PCR using known strains, bacterial cultures isolated from the test aquifer, and community DNA extracts from groundwater samples. Several novel, previously undescribed ureC sequences were obtained from bacterial isolates commonly found in groundwater. These sequences, along with others from the GenBank database were used to evaluate the ureC gene as a phylogenetic marker. A comparative phylogenetic analysis of ureC and 16S rRNA genes demonstrated that the two gene phylogenies are incongruent, but tentative species identifications based on ureC sequences could be made in some cases. A denaturing gradient gel electrophoresis (DGGE) assay using ureC-targeted PCR primers was performed using genomic DNA extracts collected during the field test. This analysis was performed to examine the bacterial community response to molasses and urea treatments and to identify urea-hydrolyzing species. DGGE analysis revealed that molasses treatments increased the apparent diversity of bacterial species containing ureC gene sequences, while a subsequent urea treatment selected for a few key urea hydrolyzers that were recruited from within the greater community of ureC-containing organisms. ureC sequence data suggested that the key urea hydrolyzers belonged to the alpha-, beta-, and gamma-Proteobacterial sub-division of the Bacterial Domain. The ureC-targeted molecular methods described herein have been used to detect and monitor ureolytic bacteria and will serve as the basis for developing future methods to enumerate gene copy number and quantify urease expression rates during applications of the remediation strategy
School code: 0320
Host Item Dissertation Abstracts International 65-07B
Subject Biology, Microbiology
Alt Author Idaho State University
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