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Structural Analysis of the TOL pDK1 xylGFJQK Region and Partial Characterization of the xylF and xylG Gene Products

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TOL plasmids encode enzymes responsible for utilization of toluene and related aromatic compounds by Pseudomonas putida, ultimately converting them to central metabolic intermediates. The nucleotide sequence for the 5.6 kb xylGFJQK region of the pDK1 TOL meta operon was determined. DNA sequence analysis revealed the presence of five open reading frames corresponding to xylG (1458 bp), xylF (846 bp), xylJ (783 bp), xylQ (936 bp) and xylK (1047 bp), encoding predicted protein products of 51.6, 31.3, 27.8, 32.8, and 36.6 kDa in size, respectively. The average G+C content of the xylLTEGFJQK region was 65.7%, somewhat higher than the 58.9% seen in the immediately upstream xylXYZ region and substantially more than the 50% G+C content reported for the upper TOL operon of this plasmid. Homology comparisons were made with genes and proteins of related catabolic plasmids. The dmpCDEFG and pWWO xylGFJQK regions exhibit consistently high levels of nucleotide and amino acid homology to pDK1 xylGFJQK throughout the entire region. In contrast, although the nucleotide sequence homology of the Acinetobacter atdCDE region to xylGFJ is high, the homology of atdFG to xylQK is markedly less. Such radical changes in homology between corresponding regions of different operons, combined with variable base and codon …
Date: December 1999
Creator: Poulter, Melinda D.
System: The UNT Digital Library
Cassette Systems for Creating Intergeneric Hybrid ATCases (open access)

Cassette Systems for Creating Intergeneric Hybrid ATCases

Cassette systems for creating intergeneric hybrid ATCases were constructed. An MluI restriction enzyme site was introduced at the carbamoylphosphate binding site within the pyrB genes of both Pseudomonas putida and Escherichia coli. Two hybrids, E. coli pyrB polar domain fused with P. putida pyrB equatorial domain and P. putida pyrB polar domain fused with E. coli pyrB equatorial domain, are possible. The intergeneric E. coli-P. putida hybrid pyrB gene was constructed and found to encode an active ATCase which complemented an E. coli Pyr- strain. These hybrids are useful for kinetic and expression studies of ATCase in E. coli.
Date: December 1999
Creator: Simpson, Luci N.
System: The UNT Digital Library

Comparative Mitochondrial DNA Sequence Diversity in Isolated and Open Populations of Southern Flying Squirrels

Access: Use of this item is restricted to the UNT Community
Three populations of Southern flying squirrels were studied in the Ouachita Mountains of Arkansas to assess the impact of population subdivision-due to island formation--on the population genetics of Glaucomys volans. One island, one mainland, and one open population were investigated. A 367 nucleotide hypervariable region of mitochondrial DNA was sequenced in individuals from each population. Individuals and populations were compared to assess relatedness. Higher sequence diversity was detected in the open and island populations. One island individual shared characters with both the island and mainland populations. Results support the hypothesis that the mainland population may have reduced gene flow. Also, the island population may have been originally founded by at least two maternal lineages.
Date: August 1999
Creator: Cook, Melaney Birdsong
System: The UNT Digital Library
Mutagenized HLA DNA Constructs: Tools for Validating Molecular HLA Typing Methodologies (open access)

Mutagenized HLA DNA Constructs: Tools for Validating Molecular HLA Typing Methodologies

This study describes the development and validation of mutagenized cloned DNA constructs, which correspond to the polymorphic regions of the class II region of the HLA complex. The constructs were used to verify the allelic specificity of primers and probes in polymerase chain reaction (PCR)-based HLA typing assays such as Sequence Specific Primers (SSP) and Sequence Specific Oligonucleotide Probes (SSOP). The constructs consisted of the entire polymorphic region of exon 2 of class II HLA allele sequences that included primer annealing sites or probe hybridization sites. An HLA allele sequence was inserted into a plasmid, cloned, then mutagenized to match a specific HLA allele, and finally, the correct clone was verified by bidirectional sequencing of the insert. Thus, the construct created a cloned reference DNA sample for any specific allele, and can be used to validate the accuracy of various molecular methodologies.
Date: May 1999
Creator: Schulte, Kathleen Q.
System: The UNT Digital Library