Biological weapons and genetic engineering.
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Biological Warfare↗
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Results for “Genetic Engineering”
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Genetic counseling and genetic engineering.
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Ethics, Medical↗
[Characteristics of atrazine-degrading genetically engineered microorganism (GEM) labeled with green fluorescent protein (GFP)].
The atrazine-degrading genetically engineered microorganism (GEM) was labeled by transforming a plasmid containing green fluorescent protein (GFP) gene. GFP gene could be expressed well in the cell of GEM after transforming. In the LB medium containing antibiotic, the expressing level of GFP was higher than in pure LB medium and in the basic medium. In the stable phase containing antibiotic, the expressing level of GFP was higher than in pure LB medium and in the basic medium. In the stable phase of cell growth, the expressing level of GFP was higher than in the lag phase and in the logarithmic phase. The transformation and expression of GFP gene wouldn't affect the degrading ability of GEM. The level of GFP expression was almost linear with the degrading activity. There exist two states of cells after inoculating GEM labeled with GFP into activated sludge: suspended state and adhered state to sludge floc, and the amount of suspended cells is higher than that adhered to sludge floc.
Atrazine↗
[Expression of recombinant human serum albumin in genetically engineered Pichia pastoris in high-density fermentation].
The optimum culture conditions of genetically engineered Pichia pastoris in shake-flask cultivation and in fed-batch fermentation were investigated respectively in this paper. It showed that the cultural period induced with 5 g/L methanol is 96 h, optimum methanol concentration is 10 g/L, and pH range is 5.72-6.59 in shake-flask culture. Although the seed inoculum amount increased, the target protein productivity per cell optical density was decreased. Their relationship fit the equeation Y = 12.941x(-0.5059) (r = 0.9789, where x is inoculating OD600, Y is protein productivity per cell optical density), we postulated that the restricting factor may be dissolved oxygen (DO) at shake-flask culture. With the 10% inoculum and 20 OD600 of seed, the lag phase of cell growth is 2.11 h in batch cultivation, and the relationship between cell optical density (Y) and culture time (t) is Y = 0.7841e0.2319t (r = 0.9936); The cell dry weight of broth reached 115-160 g/L and the maximum rHSA concentration was 3.6 g/L at the 120th at the fed-batch fermentation phase.
Fermentation↗
A genetically engineered, nonthrombogenic cellular lining for LVADs: in vitro preconditioning before in vivo implantation.
Because of the clinical success of left ventricular assist devices (LVADs) used for short-term "bridge to transplant" and the limited availability of donor organs, heart assist devices are being considered for long-term implantation as an alternative to heart transplantation. In an effort to improve biocompatibility, our laboratory has developed a nonthrombogenic cellular lining from genetically engineered smooth muscle cells (GE-SMC) for the Thermocardiosystems Heartmate LVAD. Smooth muscle cells have been transduced with the gene for endothelial nitric oxide synthase (NOS III) and produce NO at concentrations that reduce platelet deposition and smooth muscle cell proliferation when tested in vitro. In this investigation, the adhesive capabilities of GE-SMC linings were examined. An in vitro circulatory loop was designed to expose cell lined LVADs to in vivo operating conditions. Cumulative cell loss from cell lined LVADs was less than 10% after 24 hours of flow. Using a protocol for "preconditioning" the cell lining within the mock circulatory loop, the first implantation of an LVAD containing a genetically engineered SMC lining was successfully implemented in a bovine model. Results from this 24 hour study indicate that the flow-conditioned cellular lining remained intact with no evidence of thromboembolization and only minimal changes in coagulation studies.
Animals↗
Transfer of genetically engineered cells to the glomerulus.
In the rat, cultured cells injected into the renal circulation are entrapped in the glomerulus. This peculiar property allows to create chimeric glomeruli in which genetically engineered cells are populated. Using glomerular cells engineered in vitro, it is feasible to generate glomeruli that produce recombinant gene products. This approach would be useful for identification of local function of a certain gene product in the glomerulus and for therapeutic intervention in glomerular disease. Transfer of activated leukocytes to the glomerulus is useful to elucidate pathologic actions of infiltrating cells on the glomerular structure and function. Use of leukocytes in which certain gene function is selectively reinforced or deleted should enable to disclose exact roles of leukocyte-associated genes in glomerular pathophysiology. Transfer of engineered leukocytes also allows to investigate how resident cells modulate the activity of infiltrating cells in normal and pathologic circumstances. This article summarizes current experience with adoptive transfer of engineered cells to the glomerulus and addresses its potential application to kidney research.
Animals↗
Genetic engineering of the biosynthesis of glycinebetaine enhances thermotolerance of photosystem II in tobacco plants.
Genetically engineered tobacco (Nicotiana tabacum L.) with the ability to accumulate glycinebetaine was established. The wild type and transgenic plants were exposed to heat treatment (25-50 degrees C) for 4 h in the dark and under growth light intensity (300 mumol m(-2) s(-1)). The analyses of oxygen-evolving activity and chlorophyll fluorescence demonstrated that photosystem II (PSII) in transgenic plants showed higher thermotolerance than in wild type plants in particular when heat stress was performed in the light, suggesting that the accumulation of glycinebetaine leads to increased tolerance to heat-enhanced photoinhibition. This increased tolerance was associated with an improvement on thermostability of the oxygen-evolving complex and the reaction center of PSII. The enhanced tolerance was caused by acceleration of the repair of PSII from heat-enhanced photoinhibition. Under heat stress, there was a significant accumulation of H(2)O(2), O (2) (-) and catalytic Fe in wild type plants but this accumulation was much less in transgenic plants. Heat stress significantly decreased the activities of catalase, ascorbate peroxidase, glutathione reductase, dehydroascorbate reductase, and monodehydroascorbate reductase in wild type plants whereas the activities of these enzymes either decreased much less or maintained or even increased in transgenic plants. In addition, heat stress increased the activity of superoxide dismutase in wild type plants but this increase was much greater in transgenic plants. Furthermore, transgenic plants also showed higher content of ascorbate and reduced glutathione than that of wild type plants under heat stress. The results suggest that the increased thermotolerance induced by accumulation of glycinebetaine in vivo was associated with the enhancement of the repair of PSII from heat-enhanced photo inhibition, which might be due to less accumulation of reactive oxygen species in transgenic plants.
Adaptation, Physiological↗
Genetically engineered plants producing opines alter their biological environment.
Little is known about the consequences of releasing genetically engineered plants (GEP) into the environment. Using opine-producing GEP, we show that transgenic plants alter their biological environment, more precisely the root-associated bacterial populations. The alterations were both transgene-specific and target population-specific. Therefore, assessment studies on the introduction of a given transgene into a GEP will be valid on the given transgene. Evidence of any transgene-associated biological effect will depend on the determination of the pertinent target populations, the identification of which is a key step of such studies.
Bacteria↗
Can we genetically engineer safer and more effective immunotherapy reagents?
PURPOSE OF REVIEW: Progress in allergen-specific immunotherapy, the only causative form of allergy treatment, was limited by the lack of defined allergen molecules for vaccine formulation. Today the genetic informations for the most common allergens have been obtained. Here we review recombinant allergen-based technologies for the improvement of diagnosis and therapy of allergy. RECENT FINDINGS: Numerous strategies, including the genetic engineering of allergens for reduction of allergenic activity, have been developed to improve allergen-specific immunotherapy. Genetically modified allergen derivatives with reduced allergenic activity, preserved T cell epitope repertoire and retained immunogenicity have been characterized in vitro and in vivo. SUMMARY: Based on the review of the recently published data we argue that it is possible to genetically engineer safer and more effective immunotherapy reagents.
Antigens↗
Germans mix support and scepticism for genetic engineering.
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Genetic Engineering↗
Evolving strategies for the genetic engineering of herbicide resistance in plants.
Developments in plant genetic engineering technology will shortly permit the commercial introduction of transgenic crop varieties resistant to a number of non-selective herbicides. High levels of tolerance have been achieved both by overexpression of a target protein and by modification of that target to an insensitive form. However the results of preliminary trials suggest that in some instances the yield penalty for such genetic alterations will be prohibitive. An alternative strategy, based on the transfer and expression of a gene encoding a herbicide-detoxifying enzyme, appears to offer high resistance levels at low metabolic cost and is expected to assume increasing importance, although it may not prove suitable for all herbicides.
Journal Article↗
Reduction in allergenicity of grass pollen by genetic engineering.
BACKGROUND: Hay fever and allergic asthma triggered by grass pollen allergens affect approximately 20% of the population in cool temperate climates. Ryegrass is the dominant source of allergens due to its prodigious airborne pollen production. Lol p 5 or group 5 is among the most important and widespread grass pollen allergen because it reacts with IgE antibodies of more than 90% of grass pollen-allergic patients, contains most of the grass pollen-specific IgE epitopes and elicits strong biological responses. Significant efforts have been made in developing diagnostic and therapeutic reagents for designing new and more effective immunotherapeutic strategies for treatment of allergic diseases. An alternative approach to this problem could be to reduce the amount of allergen content in the source plant. METHODS: High velocity microprojectile bombardment was used to genetically engineer ryegrass. Antisense construct targeted to one of major allergen, Lol p 5, was introduced. The expression of antisense RNA was regulated by a pollen-specific promoter. Pollen was analysed for IgE reactivity. RESULTS: Analysis of proteins with allergen-specific monoclonal and polyclonal antibodies did not detect Lol p 5 in the transgenic pollen. The transgenic pollen showed remarkably reduced allergenicity as reflected by low IgE binding capacity of pollen extract as compared to control pollen. The transgenic ryegrass plants in which Lol p 5 gene expression is perturbed showed normal fertile pollen development. CONCLUSIONS: Our studies showed that it is possible to selectively 'switch off' allergen production in pollen of ryegrass demonstrating feasibility of genetic engineering of plants for reduced allergenicity.
Allergens↗
Science, law, and politics in the Food and Drug Administration's genetically engineered foods policy: FDA's 1992 policy statement.
The US Food and Drug Administration's (FDA's) 1992 policy statement was developed in the context of critical gaps in scientific knowledge concerning the compositional effects of genetic transformation and severe limitations in methods for safety testing. FDA acknowledged that pleiotropy and insertional mutagenesis may cause unintended changes, but it was unknown whether this happens to a greater extent in genetic engineering compared with traditional breeding. Moreover, the agency was not able to identify methods by which producers could screen for unintended allergens and toxicants. Despite these uncertainties, FDA granted genetically engineered foods the presumption of GRAS (Generally Recognized As Safe) and recommended that producers use voluntary consultations before marketing them.
Allergens↗
Genetic engineering of stimuli-sensitive silkelastin-like protein block copolymers.
Differentially charged analogues of block copolymers containing repeating sequences from silk (GAGAGS) and elastin (GVGVP) were synthesized using genetic engineering techniques by replacing a valine residue with glutamic acid. The sensitivity to pH and temperature was examined at various polymer concentrations, ionic strengths, and polymer lengths. The polymers transitioned from soluble to precipitate state over narrow temperature ranges. The transition temperature T(t) (the temperature at which half-maximal spectrophotometric absorption was observed) increased with increasing pH up to pH 7.0 and leveled off above this value for the Glu-containing polymer (17E)(11). T(t) was independent of pH for the Val-containing polymer (17V)(11). It decreased with increasing ionic strength, polymer concentration, and polymer length for both polymers. These results suggest that by substituting charged amino acids for neutral amino acids at strategic locations in the polymer backbone and by control of the length of silkelastin-like block copolymers using genetic engineering techniques, it is possible to precisely control sensitivity to pH, temperature, and ionic strength.
Amino Acid Sequence↗
Genetic engineering of structural protein polymers.
Genetic and protein engineering are components of a new polymer chemistry that provide the tools for producing macromolecular polyamide copolymers of diversity and precision far beyond the current capabilities of synthetic polymer chemistry. The genetic machinery allows molecular control of chemical and physical chain properties. Nature utilizes this control to formulate protein polymers into materials with extraordinary mechanical properties, such as the strength and toughness of silk and the elasticity and resilience of mammalian elastin. The properties of these materials have been attributed to the presence of short repeating oligopeptide sequences contained in the proteins, fibroin, and elastin. We have produced homoblock protein polymers consisting exclusively of silk-like crystalline blocks and elastin-like flexible blocks. We have demonstrated that each homoblock polymer as produced by microbial fermentation exhibits measurable properties of crystallinity and elasticity. Additionally, we have produced alternating block copolymers of various amounts of silk-like and elastin-like blocks, ranging from a ratio of 1:4 to 2:1, respectively. The crystallinity of each copolymer varies with the amount of crystalline block interruptions. The production of fiber materials with custom-engineered mechanical properties is a potential outcome of this technology.
Amino Acid Sequence↗
Genetic engineering and coagulation factors.
It is unfortunate that we cannot report, in the area of coagulation, advances that have been seen in related fields such as thrombolytic therapy. The reported progress (Gold et al, 1984; Van de Werf et al, 1984) with human recombinant tissue plasminogen activator (Pennica et al, 1983) augers well for the application of recombinant technology to the problems faced by patients with coagulation defects. While plasminogen activator is being assessed in an acute therapeutic setting, its use signals a beginning of the application of the technology to abnormalities of the haemostatic mechanism. Chronic administration of coagulation factors for prophylaxis and replacement therapy would appear to be just one more step down the pathway illuminated by the biochemists, microbiologists and cell biologists who have preceded the clinicians in this promising area. There is no record of the use of genetically engineered materials in the treatment of coagulation defects, primarily because the body of knowledge and refined techniques have only recently been acquired. For this reason we have had to project developments in other areas onto the problems that exist for the haemostatically compromised patient. In describing the potential usefulness of these technologies, it is difficult to ascertain where the logical projection, from a fully investigated model system, diverges from flights of imaginative fancy. Cloning projects considered overly ambitious and grandiose at the beginning of this decade are already accomplished feats. The feasibility of gene therapy in the mammalian system has been demonstrated, and trade publications now discuss governmental approval for investigative use of this procedure in 1985. Panels of physicians, scientists and even politicians now seriously contemplate and promulgate views and regulations pertaining to the efficacy and ethics of the use of genetic engineering in the treatment of human disease. The haemophilias will certainly be among the first genetic diseases to be approached with the techniques of recombinant DNA technology. Diagnostic testing, using cloned DNA, is already underway and therapeutic trials are predicted for the near future. Every observer of this rapidly growing field has to define for himself when the future is. For the potential carrier of haemophilia B, the future is now. For the physician managing the patient with a haemophilic inhibitor, the future can't come soon enough. And, for those who are concerned with man's tampering with the gene pools of living things, from viruses to humans, the future comes too quickly to be dealt with in a rational and understanding way.
Animals↗
Skin test evaluation of genetically engineered hypoallergenic derivatives of the major birch pollen allergen, Bet v 1: results obtained with a mix of two recombinant Bet v 1 fragments and recombinant Bet v 1 trimer in a Swedish population before the birch pollen season.
BACKGROUND: More than 95% of birch pollen-allergic subjects react with the major birch pollen allergen, Bet v 1, and almost 60% of them are sensitized exclusively to this allergen. OBJECTIVE: The aim of this study was to compare the in vivo biologic activity of genetically engineered hypoallergenic derivatives of Bet v 1 (an equimolar mixture of 2 recombinant [r] Bet v 1 fragments and of rBet v 1 trimer) with that of rBet v 1 wild-type by skin prick and intradermal testing. METHODS: Birch pollen-allergic patients who had not received immunotherapy (n = 23), a group of allergic patients without birch pollen allergy (n = 12), and nonatopic persons (n = 8) from northern Europe (Sweden) underwent skin prick and intradermal testing with different concentrations of the recombinant allergens and commercial birch pollen extract before the birch pollen season. Immediate and late-phase reactions were recorded and allergen-specific IgE and IgG subclass responses were determined by CAP radioallergosorbent test and ELISA, respectively. RESULTS: Atopic persons without birch pollen allergy and nonatopic individuals did not have skin reactions to rBet v 1 wild-type and genetically engineered hypoallergenic derivatives. By intradermal testing, 8 of 23 and 13 of 23 birch pollen-allergic patients did not react with the highest concentration (1 microg/mL) of the rBet v 1 fragment mix and rBet v 1 trimer, respectively, compared with 1 with rBet v 1 wild type. Likewise, the highest concentration (100 microg/mL) of fragment mix or trimer failed to elicit a positive skin prick test in 18 of 23 and 15 of 23 patients in comparison with 0/23 with the monomer. No late reactions were observed. CONCLUSION: The recombinant hypoallergenic birch pollen allergens can probably be used for patient-tailored immunotherapy with a reduced risk to induce anaphylactic reactions.
Adolescent↗
Streptavidin-based containment systems for genetically engineered microorganisms.
The use of genetically modified microorganisms for environmental remediation continues to be debated. Conditional lethal systems with tightly regulated gene expression can be used to contain released microorganisms and ameliorate some of the concerns about horizontal gene transfer. We have described streptavidin-based suicide systems to address these concerns and evaluated their function in Pseudomonas putida containing the TOL plasmid for aromatic hydrocarbon metabolism. Tight regulation of expression of a truncated streptavidin gene was required to avoid premature production of the toxic protein. Streptavidin expression was induced by the absence of 3-methyl benzoate (hydrocarbon substrate) which resulted in the elimination of 99.9% of the bacterial culture within eight hours. Low mutant escape rates at 10(-7) per cell per generation were also realized.
Bacteria↗