Microbiology Commons^™

The Expression And Regulation Of The Hsdk Genes Of Escherichia Coli K-12, Ainu Parkash

Loma Linda University Electronic Theses, Dissertations & Projects

The DNA restriction and modification (R-M) genes in bacteria play a defensive role in the organism, offering protection to the host from invaders such as bacteriophages. The expression of these genes in E. coli K-12 had not previously been studied due to the complexity of the EcoK R-M enzymes. The use of lacZ gene fusion techniques allowed studies of the expression of the hsdK genes for the first time. Operon fusions of Mu(/ac) to the hsdK genes show that the two hsdK promoters, p _res and p _mod, are weak (50 Miller units) and that …

Altered Expression Of Adenovirus 12 Dna-Binding Protein But Not Dna Polymerase During Abortive Infection Of Hamster Cells, Lynne A. Lucher, Benjawan Khuntirat, Jiansheng Zhao, Peter C. Angeletti

Nebraska Center for Virology: Faculty Publications

Replication of human adenovirus type 12 DNA is blocked in abortively infected baby hamster kidney cells. The activity and accumulation of adenovirus 12 DNA polymerase is equivalent in infected hamster and human cell extracts. However, the accumulation of adenovirus type 12 DNA-binding protein is approximately 120-fold lower in extracts from infected hamster cells when compared to infected permissive human cells. This difference in accumulation is not because of replication of viral DNA during productive infection, since this difference is observed in the presence of hydroxyurea. The DNA-binding protein from infected hamster cells retains the ability to bind denatured DNA-cellulose. An …

Ecdysterone Regulatory Elements Function As Both Transcriptional Activators And Repressors., Leonard Dobens, Karen Rudolph, Edward M. Berger

Dartmouth Scholarship

A synthetic, 23-bp ecdysterone regulatory element (EcRE), derived from the upstream region of the Drosophila melanogaster hsp27 gene, was inserted adjacent to the herpes simplex virus thymidine kinase promoter fused to a bacterial gene for chloramphenicol acetyltransferase (CAT). Hybrid constructs were transfected into Drosophila S3 cells and assayed for ecdysterone-inducible CAT expression. In the absence of ecdysterone a tandem pair of EcREs repressed the high constitutive level of CAT activity found after transfection with the parent reporter plasmid alone. After hormone addition very high levels of CAT activity were observed. Insertion of the EcRE pair 3' of the CAT gene …

Characterization Of Hsd::Mudx128 Operon Fusion Mutant Of Escherichia Coli K-12, Marjorie Ann T. Reyno

Loma Linda University Electronic Theses, Dissertations & Projects

The study of the regulation of gene expression in complex genes can be facilitated by the use of operon fusions which place a well characterized lacZ gene under the control of a promoter of interest. An hsd::MudX128 operon fusion mutant of E. coli K-12 isolated by Prakash (1986) was observed to have a high β-galactosidase activity (1000 units), ten to twenty times higher than other hsd::MudX mutants. This β-galactosidase activity was suppressed to a very low level (25 units) by the introduction of an F’ plasmid, F'101, which carries the 98 min to 2 min region of the …

Genetic Transformation In Salmonella Typhimurium, Siao Ping Tasi

Loma Linda University Electronic Theses, Dissertations & Projects

Plasmid transformation is an essential step in many gene cloning experiments. In Salmonella tynhimurium, mutants of rough strain have shown improved transformation frequency previously. In this work a stable restriction-minus (r^-) and modification-plus (m⁺) strain and its derivatives were constructed. Effect of restriction barrier on transformation of plasmid pBR322 was tested. Several factors which may affect transformation frequency have been systematically examined to determine the optimum conditions for transformation in S. tvphimurium. Efforts were made to simplify the procedure.

Using pBR322 plasmid as an experimental model, the r^- strain demonstrated 10⁵ to …

Partial Characterization Of Lpss From Rhizobium Fredii Usda 205 Before And After Nod Gene Induction, Bradley Lawrence Reuhs

Masters Theses

The lipopolysaccharides (LPS) from Rhizobium fredii USDA 205 and its nod- mutant, HC 205, which has been cured of the 192-MDa symbiotic plasmid, were isolated by the phenol-water extraction method. The LPS from the wild-type strain which was grown in association with host-plant root extract or a flavone analog to a constituent of the root extract, which is required for the induction of at least a portion of the nodulation genes, was also isolated. The conditions required for the induction of the nod genes were determined by work with a nod- mutant of R. fredii USDA 201 which …

A Simple Solid-Phase Electrophoretic Procedure For The Separation Of Plasmid Dna, Linda Ann Simurra

Biological Sciences Theses & Dissertations

A method was developed for extraction of plasmid DNA from bacterial cells embedded in agarose blocks. Cell - containing blocks were treated with various lysing reagents and inserted into the wells of an agarose gel. Upon electrophoresis the plasmid DNA migrated out of the embedding block and into the gel leaving intact chromosomal DNA in the well. This method was tested with various organisms and found to be effective for plasmid isolation. In comparison to "traditional" procedures, this new method is less tedious since chemical separation of plasmids is not required prior to electrophoresis. Also, a higher yield of plasmid …

Restriction Analysis Of Bacteriophage, Ali Reza Mostaghimi

Loma Linda University Electronic Theses, Dissertations & Projects

A restriction map of DNA from bacteriophage P3-C₁ for the endonucleases BamH I, Bgl II, EcoR I, Hind III, Sac I and Sca I is described in this study. The phage DNA was determined to be a linear double stranded molecule with a unit genomic length of 31 kb. After electrophoresis in 0.5% agarose gels, the undigested DNA appeared as three molecules of 31, 62 and 93 kb in length. The Hind III digestion pattern of the individual 31 kb molecule was identical to the 62 kb and 93 kb Hind III pattern. With all enzymes except Hind III, …

Restriction Mapping And Expression Of Recombinant Plasmids Containing The Arsenic Resistance Genes Of The Plasmid R45, Terry M. Coons

Dissertations and Theses

The trivalent (arsenite) and pentavalent (arsenate) forms of arsenic are introduced into the environment through the use of arsenic in herbicides, pesticides, fertilizers, and the smelting of arsenic-bearing ores. Bacteria resistant to arsenic are readily isolated from surface waters, sewage, and clinical infections. Although some bacterial resistance is provided by inducible phosphate transport systems that discriminate against arsenate, marked resistance is carried on bacterial plasmids.

A 6.9 kilobase fragment previously derived from one such plasmid, R45, and containing the genes for inducible resistance to arsenite and arsenate was ligated into the cloning vectors puce and pUC9 in opposite orientations and …

A Comparison Of The Acidic Extracellular Polysaccharides From Rhizobium Trifolii Mutants Affected In Trifolium Repens Root Hair Curling, Brian Andrew Hanley

Masters Theses

The compositions and sugar linkages of the acidic extracellular polysaccharide (EPS) from several Rhizobium trifolii mutants were compared. These mutants were derived from a wild type Rhizobium trifolii, ANU843, and are unable to cause normal root hair curling, Hac-, or nodulate clover, Nod-. Three of the mutants contain a Tn5 insertion in a 5.3 kb BglII fragment of the symbiotic plasmid (pSym). A fourth mutant is missing the pSym, which includes much of the 5.3 kb BglII fragment, both of these deletions result in Hac-Nod- phenotypes. When the 5.3 kb BglII fragment is cloned (pRT150) and reinserted into the …

Seasonal Responses Of Filtration Rates To Temperature, Oxygen Availability, And Particle Concentration Of The Freshwater Clam, Musculium Partumeium (Say), Albert J. Burky, Richard B. Benjamin, Denis G. Conover, Jennifer Detrick

Biology Faculty Publications

Seasonal filtration rates of a pond population of the freshwater pisidiid clam, Musculium par­tumeium (Say), were assessed by measuring the clearance of 2.02 µ.m latex beads from suspension and expressed in terms of FR = a(AFDW}b where FR = filtration rate (ml H20 · hr- 1 ) , AFDW = mg ash-free dry weight of a whole clam and "a" and "b" are constants. The a-values ( = FR of a 1-mg AFDW clam} vary seasonally with the highest rates corresponding to periods of maximum growth and reproduction in the spring and fall. Changes in "b" reflect the influence of …

Cole1 Copy Number Mutants., Londa Schmidt, Joseph Inselburg

Dartmouth Scholarship

A deletion mutant of the colicin E1-derived plasmid, pDMS6642, exhibited an approximately fourfold increase in copy number. We subsequently isolated hydroxylamine-induced mutants of that plasmid that had a further increase in copy number. Analysis of them suggests that the increased copy number of pDMS6642 is associated with transcriptional readthrough from a Tn3 transposon into the region of ColE1 containing information that influences plasmid replication. The hydroxylamine mutation in one copy number mutant appeared to increase the plasmid copy number by stimulating readthrough transcription from the Tn3 transposon into the ColE1 replication control region, whereas the other hydroxylamine mutation acts by …

Evidence For The Horizontal Acquisition Of Murine Akr Virogenes By Recent Horizontal Infection Of The Germ Line, Stephen J. O'Brien, Janet L. Moore, Malcolm A. Martin, James E. Womack

Biology Faculty Articles

Several recent reports have established the biological and molecular genetic similarity between the endogenous AKV virus of strain AKR, and an N-ecotropic endogenous virus found in the genome of feral Japanese mice, Mus musculus molossinus. The similarities are so striking as to suggest a common origin of these viruses, which are present in some, but not all, inbred mouse strains. The virogenes of AKR mice may have been acquired by either: (a) common descent of AKR (and other AKV⁺ strains) from a common ancestor of AKR and molossinus animals, or (b) horizontal germ line infection of the AKR …

Analysis Of Multiple Isoenzyme Expression Among Twenty-Two Species Of Mycoplasma And Acholeplasma, Stephen J. O'Brien, Janice M. Simonson, Marion W. Grabowski, Michael F. Barile

Biology Faculty Articles

Crude extracts of triple-cloned, purified cultures of 22 species of Mycoplasma and Acholeplasma were examined for expression of 21 isozyme systems routinely used to type mammalian cells. Nine previously described enzymes (purine nucleoside phosphorylase, adenylate kinase, dipeptidase, esterase, glyceraldehyde-3-phosphate dehydrogenase, glucose phosphate isomerase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and superoxide dismutase) and three enzymes not previously reported in mycoplasma (triose phosphate isomerase, inorganic pyrophosphatase, and acid phosphatase) were detected in some or all of the species examined. These findings provide new information on the enzymatic expressions of these organisms. Three of the isozyme systems (superoxide dismutase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase) …

Interpopulation Variation In Calcareous And Proteinaceous Shell Components In The Stream Limpet, Ferrissia Rivularis, Carl M. Way, Daniel J. Hornbach, Albert J. Burky

Biology Faculty Publications

Natural populations of the North American stream limpet, F. rivularis, were studied in upstate New York [USA], in a set of localities whose waters have a 15-fold range of dissolved Ca (4.6-67.6 mg/liter) and also range from oligotrophy to eutrophy. Shell component analyses (CaCO3, total organic C and total N) are reported as component mass-fractions (mg/g or .mu.g/g dry weight) and as values for a standard limpet shell of 35 mm aperture length (AL). More than 2-fold differences occur between populations in all 3 components, with relatively little variation occurring within each population. Expressed per standard limpet, CaCO3 values for …

Seasonal Metabolism Of The Sphaeriid Clam, Musculium Partumeium (Say), From A Permanent And A Temporary Pond, Carl M. Way, Daniel J. Hornbach, Albert J. Burky

Biology Faculty Publications

Seasonal metabolic rates were examined for the sphaeriid clam, Musculium partumeium (Say) from a permanent and a temporary pond. For the fall-born generation from the permanent pond and for the single generation from the temporary pond, metabolic rates peak during periods of greatest growth and reproduction. Metabolic rates were consistently higher for the permanent pond population. These rates for Musculium partumeium extend the available information on the family Sphaeriidae to the genus Musculium.

Growth Of Pisidium Casertanum (Poli) In West Central Ohio, Albert J. Burky, Daniel J. Hornbach, Carl M. Way

Biology Faculty Publications

Clarke (1973) states that Pisidium casertanum (Poli) is perhaps the most common species of Pisidium and is truly cosmopolitan, occurring throughout Eurasia, Africa, Australia, and South, Central and North America. Populations of these clams are reported in habitats ranging from ephemeral ponds to benthic zones of deep lakes. Our study describes Manuscript received 29 May 1979 and in revised form 25 July 1980 (#79-34). 2 Present address: Department of Biology, University of Virginia, Charlottesville, VA 22901. 3 Present address: Department of Zoology, Miami University, Oxford, OH 45056. 41 aspects of the life-cycle of P. casertanum in a spring-fed stream with …

Conservation And Variation Of Nucleotide Sequences In Escherichia Coli Strains Isolated From Nature, Lawrence G. Harshman, Monica Riley

Lawrence G. Harshman Publications

A group of Escherichia coli isolates from nature were compared with one another and with laboratory strains of E. coli with respect to size distribution of chromosomal restriction endonuclease fragments and differences in nucleotide sequences in selected small portions of the genomes. The estimated frequency of base substitutions in nucleotide sequences in and near the trp operons of 26 of the 28 E. coli strains examined ranged from 0.008 to 0.066. Nucleotide sequences in or near λ prophage homologs were significantly more variable than the sequences in or near trp, tnaA, and thyA genes. Thus, the λ-homologous regions …

Studies Of The Lysogenic Response As A Function Of Multiplicity Of Infection Of Lambdoid Bacteriophages, Mark Floyd Fedyk

Masters Theses

The lambdoid bacteriophages λimm^λ, λimm⁴³⁴ and λcI are ideal subjects for the study of gene product interactions at the molecular level due to their closely related genetic makeup. Previous work in this laboratory has indicated the possibility that phage λimm is sensitive to phage λimm⁴³⁴cI gene product and that phage λimm⁴³⁴ is sensitive to phage λimm^λcI gene product. The purpose of this research was to determine if the frequency of lysogeny for phage λimm^λ is affected by the multiplicity of coinfection with the λimm⁴³⁴ phage. By comparing the graph of …

Bvr-1, A Restriction Locus Of A Type C Rna Virus In The Feline Cellular Genome: Pleiotropic Restriction Of Endogenous Balb Virus In Cat X Mouse Somatic Cell Hybrids, Stephen J. O'Brien, Janice M. Simonson

Biology Faculty Articles

Bvr-1 is a dominant X-linked feline gene which restricts the replication of B-tropic murineleukemia virus (B-MuLV) in somatic cell hybrids between murine BALB/c-RAG cells and FL-74 feline cells. Since the hybrids were originally derived by the hypoxanthine aminopterin thymidine selection scheme, counter selection experiments on 6-thioguanine result in preferential survival of hybrid cells which have spontaneously lost the feline X-chromosome on which is located the structural gene for hypoxanthine guanine phosphoribosyl transferase (IMP: pyrophosphate phosphoribosyl transferase, E.C. 2.4.2.8) and Bvr-1. Back selected Bvr-1^- cells express high parental levels of B-MuLV. Bvr-1^- effectively restricts the IdU-mediated induction of …

Buoyancy Changes As Related To Respiratory Behavior In An Amphibious Snail, Pomacea Urceus (Müller), From Venezuela, Kathleen A. Burky, Albert J. Burky

Biology Faculty Publications

The ampullariid Pomacea urceus uses its ctenidium and lung in respiration. The snails reach the surface with their siphon and ventilate their lung by withdrawing their head-foot in a pumping action. The lung gas also serves to change the buoyancy of the snails. After a ventilation a snail does not necessarily return to the same buoyancy level. They may achieve overall specific gravities of <1 to > 1 after a ventilation. They may remain submerged (s.g. > 1) or float (s.g.

Biochemical Properties Of Salmonella Potsdam Bacteriophage P4/2, Clyde L. Roggenkamp

Loma Linda University Electronic Theses, Dissertations & Projects

Over 90% recovery of phage P4/2 was realized with differential centrifugation concentration and purification, whereas only about 16% was recovered from the aqueous two-phase polymer - Genetron - ECTEOLA-cellulose column system. Purity of preparations was greatest with the density gradient centrifugation (A₂₆₀ = 2.66 and A₂₈₀ = 1.80 per 10¹² plaque-forming units). With the chemical system of concentration and purification followed by an ECTEOLA anion exchange run, the A₂₆₀ was 6.6 and the A₂₈₀ was 4.3.

Three tests were employed to determine the configuration of P4/2 DNA: (1) a formalin denaturation test, (2) a thermal …

Replicating Single-Cycle Adenovirus Vectors Generate Amplified Influenza Vaccine Responses, Catherine M. Crosby, William E. Matchett, Stephanie S. Anguiano-Zarate, Christopher A. Parks, Eric A. Weaver, Larry R. Pease, Richard J. Webby, Michael A. Barry

Nebraska Center for Virology: Faculty Publications

Head-to-head comparisons of conventional influenza vaccines with adenovirus (Ad) gene-based vaccines demonstrated that these viral vectors can mediate more potent protection against influenza virus infection in animal models. In most cases, Ad vaccines are engineered to be replication-defective (RD-Ad) vectors. In contrast, replication-competent Ad (RC-Ad) vaccines are markedly more potent but risk causing adenovirus diseases in vaccine recipients and health care workers. To harness antigen gene replication but avoid production of infectious virions, we developed “single-cycle” adenovirus (SC-Ad) vectors. Previous work demonstrated that SC-Ads amplify transgene expression 100-fold and produce markedly stronger and more persistent immune responses than RD-Ad vectors …