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At least 109 records · Page 6Linked to original sources
PubMed2004

Animal models for studying respiratory syncytial virus infection and its long term effects on lung function.

BACKGROUND: Human respiratory syncytial virus (hRSV) infection causes a spectrum of illnesses ranging from mild infection to life-threatening bronchiolitis and respiratory failure. Human studies on the pathogenesis of RSV infection are invaluable, but animal models permit advances with the use of experimental strategies that would be inappropriate in human studies. METHODS: We review the advantages and disadvantages of various animal models for the study of hRSV infection. RESULTS: No animal model of hRSV infection replicates the complete spectrum of disease severity seen in humans. Available models differ in their ability to incorporate genetic technology and to allow the study of immunity, vaccine efficacy and treatment interventions. Although hRSV establishes disease in primates, this advantage is outweighed by the impracticalities and cost of using such models. The study of bovine RSV infection in calves is appealing because of parallels with human disease. Among rodent models, BALB/c mice have helped delineate the mechanisms underlying vaccine-enhanced RSV disease, and cotton rats have been used for preclinical testing. The single major disadvantage of studying hRSV in rodent models is the limited extent to which this host-restricted human pneumovirus replicates in mouse lung tissue. The rodent-specific Pneumovirus pathogen, pneumonia virus of mice, causes an infection that mirrors severe bronchiolitis and pneumonia in infants infected with RSV, including robust virus replication with profound inflammation. CONCLUSION: The recent development of the pneumonia virus of mice model has permitted exploration of the mechanisms of severe Pneumovirus disease in vivo with the use of sophisticated genetic tools and genetically manipulated mouse strains.

Animals↗
PubMed2003

Electrotransformation of Clostridium paraputrificum M-21 with some plasmids.

Clostridium paraputrificum M-21 was transformed with several shuttle plasmids constructed for Clostridium acetobutylicum-Escherichia coli and Clostridium perfringens-E. coli by electroporation. The Clostridium stercorarium xylanase gene xyn10B was successfully expressed in C. paraputrificum M-21 and the expressed protein did not suffer from proteolysis by host protease(s). This system will provide us with a genetic tool for genetic and metabolic engineering of this bacterium.

Journal Article↗
PubMed1989

Genetic and mutational tools for investigating the genetics and molecular biology of trichothecene production in Gibberella pulicaris (Fusarium sambucinum).

Gibberella pulicaris (Fusarium sambucinum) is a promising organism for studying the genetics and regulation of trichothecene biosynthesis; conditions for obtaining fertile crosses have been defined (Desjardins & Beremand, 1987) and crosses between natural variants have provided some information about the number, location, arrangement, and role of genes which determine trichothecene production (Desjardins & Beremand, 1987; Beremand & Desjardins, 1988). The development of some additional experimental tools and methodologies required for the further genetic analysis of trichothecene production in G. pulicaris are described in the present study. A highly fertile, isogenic line was constructed for G. pulicaris strain R-6380. The ability to readily generate mutants in this strain was also demonstrated. Both biochemical and morphological mutants were obtained following UV-mutagenesis. The inheritance of some of these mutations through meiosis indicated that they will be useful genetic markers for crosses and mapping studies. Since strain R-6380 is also transformable (Salch & Beremand, 1988), it is an excellent choice for transmission and molecular genetic studies involving trichothecene production.

Crosses, Genetic↗
PubMed1987

What's new in mutagenicity and carcinogenicity--status of short-term assay systems as tools in genetic toxicology and carcinogenesis.

The status of short term assay systems as tools in genetic toxicology has shifted from the goal to predict carcinogenic activity of chemicals toward their employment for understanding and elucidating the mechanisms of biological activity. This altered mode of application arises from research development in two main areas. One resulted in the observation that the calculated predictive values of individual assay systems (alone or in combination with supplementary tests) to detect carcinogens as genotoxic and non-carcinogens as non-genotoxic are lower than originally expected. The other is the increasing recognition that by employing relevant in vitro procedures, various aspects of a compound's activity can be studied which may otherwise not be clarified with available in vivo methods. This report focuses on the reasons why mutagenesis assays, or short term genotoxicity assays, in general, are not unambiguously employed for determining the carcinogenic potential of unknown compounds. One reason is that a safe prediction is not possible, since non-genotoxic carcinogens will not induce alterations of the DNA. Another reason is that even genotoxic carcinogens may not accurately respond as positive in a given test system due to the limitations of the specific assay. These are mainly seen in the incomplete metabolic conversion of the test compound, irrelevancy the measured effect may actually have for carcinogenesis, and the lack of regarding pharmacokinetic influences by a host animal, when testing in vitro. A rationale testing strategy compiled of assay systems which individually are included to overcome the restrictions is described.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1996

Genetic diagnosis from formalin-fixed fetal tissue using FISH: a new tool for genetic counseling in subsequent pregnancies.

We evaluated the feasibility of retrospective genetic testing for numerical chromosomal aberrations by applying the FISH technique to formalin-fixed fetal tissue. Fetal tissue from 10 old cases with known aneuploidy and from 13 cases with known fetal malformations, were tested with specific DNA probes for pericentromeric repeat regions of chromosomes 13/21, 18, X and Y. FISH diagnosis concurred with karyotype in all nine cases with sufficient cells. Numerical aberration was diagnosed in six out of 13 cases with fetal malformations.

Aneuploidy↗
PubMed1983

The family history and genetic counseling. Tools for preventing and managing inherited disorders.

Genetic disorders are more common than is generally appreciated and are the cause of significant morbidity and mortality. A number of these diseases are treatable, and in some cases, their clinical manifestations are preventable by early diagnosis and therapeutic intervention. A brief family history, obtained in a careful and thoughtful manner, can provide useful information to the physician for the diagnosis and management of patients with inherited diseases and, most importantly, can provide a unique opportunity to practice preventive medicine. Patients with known or suspected genetic disorders, including birth defects and mental retardation, deserve appropriate consultation and the opportunity for genetic counseling. This counseling is essential to permit couples at risk to make appropriate, informed reproductive decisions. Specialized diagnostic tests, such as chromosome analysis, are available, and many disorders can be diagnosed prenatally by amniocentesis.

Adult↗
PubMed1999

Hyperactive transposase mutants of the Himar1 mariner transposon.

Mariner-family transposable elements are active in a wide variety of organisms and are becoming increasingly important genetic tools in species lacking sophisticated genetics. The Himar1 element, isolated from the horn fly, Haematobia irritans, is active in Escherichia coli when expressed appropriately. We used this fact to devise a genetic screen for hyperactive mutants of Himar1 transposase that enhance overall transposition from approximately 4- to 50-fold as measured in an E. coli assay. Purified mutant transposases retain their hyperactivity, although to a lesser degree, in an in vitro transposition assay. Mutants like those described herein should enable sophisticated analysis of the biochemistry of mariner transposition and should improve the use of these elements as genetic tools, both in vivo and in vitro.

Animals↗
PubMed2005

Development of a method for markerless genetic exchange in Enterococcus faecalis and its use in construction of a srtA mutant.

Enterococcus faecalis is a gram-positive commensal bacterium of the gastrointestinal tract and an important opportunistic pathogen. Despite the increasing clinical significance of the enterococci, genetic analysis of these organisms has thus far been limited in scope due to the lack of advanced genetic tools. To broaden the repertoire of genetic tools available for manipulation of E.faecalis, we investigated the use of phosphoribosyl transferases as elements of a counterselection strategy. We report here the development of a counterselectable markerless genetic exchange system based on the upp-encoded uracil phosphoribosyl transferase of E. faecalis. Whereas wild-type E. faecalis is sensitive to growth inhibition by the toxic base analog 5-fluorouracil (5-FU), a mutant bearing an in-frame deletion of upp is resistant to 5-FU. When a cloned version of upp was ectopically introduced into the deletion mutant, sensitivity to 5-FU growth inhibition was restored, thereby providing the basis for a two-step integration and excision strategy for the transfer of mutant alleles to the enterococcal chromosome by recombination. This method was validated by the construction of a DeltasrtA mutant of E. faecalis and by the exchange of the surface protein Asc10, encoded on the pheromone-responsive conjugative plasmid pCF10, with a previously isolated mutant allele. Analysis of the DeltasrtA mutant indicated that SrtA anchors Asc10 to the enterococcal cell wall, facilitating the pheromone-induced aggregation of E. faecalis cells required for high-frequency conjugative plasmid transfer in liquid matings. The system of markerless exchange reported here will facilitate detailed genetic analysis of these important pathogens.

Aminoacyltransferases↗
PubMed1987

Transposition of gram-positive transposon Tn916 in Acholeplasma laidlawii and Mycoplasma pulmonis.

Mycoplasma genetics has been limited by a lack of genetic tools such as selectable markers, methods to transfer DNA, and suitable vectors for cloning. Studies were undertaken to examine the potential of using the streptococcal transposon Tn916 as a mycoplasma genetic tool. The Escherichia coli plasmid pAM120, which contains Tn916, was transformed into Acholeplasma laidlawii and Mycoplasma pulmonis. Transposition of Tn916 into the mycoplasma chromosome apparently occurred by an excision-insertion mechanism. This example shows that newly introduced DNA from other bacteria can be successfully expressed in mycoplasma and that Tn916 should serve as a powerful genetic tool for the study of mycoplasmas.

Acholeplasma laidlawii↗
PubMed2007

Functionality of the beta/six site-specific recombination system in tobacco and Arabidopsis: a novel tool for genetic engineering of plant genomes.

The beta recombinase is a member of the prokaryotic site-specific serine recombinases (invertase/resolvase family), which in the presence of a DNA bending cofactor can catalyse DNA deletions between two directly oriented 90-bp six recombination sites. We have examined here whether the beta recombinase can be expressed in plants and whether it displays in planta its specific catalytic activity excising DNA sequences that are flanked by six sites. In plant protoplasts, the enzyme could be expressed as a GFP-beta recombinase fusion which can localise to the cell nucleus. Beta recombinase stably expressed in tobacco plants can catalyse deletion of a spacer region that is flanked by directly oriented six sites and has been placed between promoter and a GUS reporter gene (preventing GUS expression). In transient transformation experiments, beta recombinase-mediated elimination of the spacer results in transcriptional induction of the GUS gene. Similarly, beta recombinase in stably double-transformed Arabidopsis plants deletes specifically the spacer region of a reporter construct that has been incorporated into the genome. In the segregating T1 generation, plants were identified that contain exclusively the recombined reporter construct. In summary, our results demonstrate that functional / recombinase can be expressed in plants and that the enzyme is suitable to precisely eliminate undesired sequences from plant genomes. Therefore, the beta/six recombination system (and presumably related recombinases) may become an attractive tool for plant genetic engineering.

Amino Acid Sequence↗
PubMedDate not supplied

Molecular genetic identification tools for three commercially cultured oysters (Crassostrea belcheri, Crassostrea iredalei, and Saccostrea cucullata) in Thailand.

Molecular genetic keys for identification of 3 commercially cultured oysters (Crassostrea belcheri, Crassostrea iredalei, and Saccostrea cucullata) in Thailand were developed based on restriction analysis of 18S ribosomal DNA and cytochrome oxidase subunit I (COI). Digestion of the amplified 18S rDNA with Hinf I unambiguously differentiated Crassostrea oysters from Saccostrea oysters and Striostrea (Parastriostrea) mytiloides. In addition, species-specific restriction fragment length polymorphism patterns of C. belcheri, C. iredalei, and S. cucullata were consistently observed when the gel-eluted COI was digested with Mbo I and Dde I. Thirty composite haplotypes were observed across all individuals. Species-specific composite haplotypes were found in C. belcheri (AAAA and AAAB), C. iredalei (AABC and AABU), and S. cucullata (BBCD and BBCE), respectively. The most common composite haplotype of COI in C. belcheri (AAAA), C. iredalei (AABC), and S. cucullata (BBCD) was amplified, cloned, and sequenced. Detection of C. belcheri and C. iredalei based on polymerase chain reaction was further developed using more specific primers (HCO2198 and R372) followed by digestion of a 372-bp product with Mbo I.

Animals↗
PubMed1994

Construction of composite transposons for halophilic Archaea.

Transposons with selectable marker genes (e.g., antibiotic resistance) have been extremely useful tools in bacterial genetics but have not been found naturally in Archaea. We constructed synthetic transposons consisting of halobacterial ISH elements (ISH2, ISH26, or ISH28) flanking a mevinolin resistance determinant. Introduction of these constructs into Haloferax volcanii cells can produce drug-resistant transformants through homologous recombination between the plasmid hmgA gene and the chromosomal hmgA locus. This problem was overcome by using another host, Haloarcula hispanica, the hmgA gene of which shares little homology with that from Haloferax volcanii. Introduction of an ISH28-based transposon (ThD28) into Haloarcula hispanica cells produced numerous transformants. Each of these was shown to contain an ISH-flanked mevinolin resistance determinant integrated into the cellular DNA. Integration was not obviously site specific. Transposon ThD26 (based on ISH26a), was less mobile, relative to ThD28, and the ISH2-based construct (ThD22) did not transpose at all in these cells. The further development of halobacterial transposons may provide useful genetic tools allowing rapid isolation and analysis of halobacterial genes, particularly those with no selectable phenotype.

Base Sequence↗
PubMed1998

Genetic studies in Borrelia burgdorferi.

Borrelia burgdorferi, the agent of Lyme disease, has recently joined a growing number of micro-organisms for which the entire genomic sequence is known. Despite this wealth of information, little is known about the contribution of specific spirochetal components to the pathogenesis of Lyme disease or their function in the normal life cycle of the organism. This discrepancy is due in part to the lack of a well-developed genetic system in B. burgdorferi, which in turn can be attributed to its relatively recent isolation and the dissimilarity of Borrelia from other genetically tractable bacteria. We are interested in several plasmid-encoded gene products in B. burgdorferi that may play a role in sensing and adaptation to the different environments the spirochete encounters as it completes an infectious cycle between the tick vector and the mammalian host. We are developing genetic tools with which to test the roles of specific B. burgdorferi gene products in the transmission cycle in an animal model of Lyme disease. We have demonstrated targeted gene inactivation by allelic exchange, using the gyrBr gene encoding coumermycin-resistant topoisomerase as a selectable marker. Spirochetes are transformed by electroporation and coumermycin-resistant colonies are screened by PCR for allelic exchange at the targeted locus. We have successfully inactivated several genes of interest in the type strain B31. We are investigating the utility of additional antibiotic resistance genes as selectable markers in B. burgdorferi. Targeted gene inactivation is a powerful tool with which to investigate the role of particular proteins in the basic biology and virulence of a pathogenic microorganism. We have made significant advances in our ability to genetically manipulate B. burgdorferi in order to address these issues. However, the available methods are incomplete and far from routine. We are currently improving existing methods as well as developing additional genetic tools with which to augment genetic studies in B. burgdorferi.

Borrelia burgdorferi Group↗
PubMed2005

Development of an audit tool for genetic services.

Rapid growth in demand and altered professional roles have produced changes in the delivery of genetic services over the past decade, but these have not been rigorously evaluated because of the paucity of appropriate audit tools. The aim of this study was to use clients' accounts and factor analysis to develop a robust assessment and audit tool. Qualitative data abstracted from several published studies were used to generate a number of statements related to outcomes of genetic services. A total of 57 statements were incorporated into a questionnaire. The questionnaire was mailed to clients of the Wales genetic service (n = 133) who had completed their episode of care. Respondents were asked to rank each statement on a seven-point Likert type scale. Responses were subjected to factor analysis. A total of 97 anonymized responses were received (73% response rate). Six main factors were found to contribute to the outcome of the service from the client's perspective. These were labeled (i) enhanced understanding, (ii) positive psychological change, (iii) respect for autonomy, (iv) adaptation, (v) disequilibirium, and (vi) value of contact. The audit tool has now been refined for use in evaluating genetic services. Questions are included to investigate the six outcome areas shown to be relevant from the client's perspective. Data from the next phase of the study will be analyzed to validate the tool for use in both clinical audit and research contexts.

Data Collection↗
PubMed2004

Prevention of type 2 diabetes mellitus from the viewpoint of genetics.

The prevalence of type 2 diabetes mellitus has increased worldwide over the past decades. The economic and social cost associated with diabetes mellitus and its complications have also increased enormously. Since current methods of treating diabetes is inadequate, the most effective way to reduce the burden associated with type 2 diabetes would be to prevent diabetes itself. Current strategies to prevent type 2 diabetes mellitus are based on efforts to reduce insulin resistance and to preserve or increase pancreatic beta cell function in high risk individuals. Now there is substantial evidence that life style intervention or pharmacologic agents can reduce the development of diabetes in individuals with impaired glucose tolerance. However, it remains to be determined whether the result would be similar in other risk groups for type 2 diabetes. Although development of type 2 diabetes mellitus is influenced by a complex interaction between genetic and environmental factors, current prevention strategies do not reflect contribution of genetic factors. Over the past decade, many investigators tried to find diabetogenic genes by traditional gene discovery methods. It appears that multiple genes with weak effect are involved in the development of type 2 diabetes mellitus which makes searching diabetogenic genes more complicated. In addition to the completion of human genome project, new genetic tools to identify genetic variations and analyze gene to gene or gene to environmental interaction will facilitate the discovery of susceptibility genes for type 2 diabetes mellitus in near future. Once we could identify susceptibility genes and gene to environmental interaction of type 2 diabetes mellitus, we can clarify the role of both environmental and genetic factors in the development of type 2 diabetes mellitus. Moreover, we can identify people who can get more benefit from prevention and also can provide better and personalized preventive measure based on genetic information.

Diabetes Mellitus, Type 2↗
PubMed2001

Lactobacillus sakei: recent developments and future prospects.

Lactobacillus sakei is one of the most important bacterial species involved in meat preservation and meat fermentation. In the last fifteen years, numerous studies have focused on this species due to its important role in food microbiology. The present paper reviews current knowledge of this emerging species in the fields of taxonomy, phylogeny and physiology, and metabolism. Recent developments in genetic tools and molecular genetics will also be emphasized to evaluate future prospects.

Food Microbiology↗