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  • 1995-1999  (12)
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  • 1996  (12)
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  • gene transfer
  • 1
    Digitale Medien
    Digitale Medien
    Adenovirus-mediated gene transfer using ex vivo perfusion of the heart graft (1996)
    Springer
    Surgery today 26 (1996), S. 624-628 
    Details
    ISSN: 1436-2813
    Schlagwort(e): gene transfer ; adenovirus ; heart
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Medizin
    Notizen: Abstract A replication-deficient adenovirus was used for ex vivo gene transfer into rat heart grafts under conditions simulating clinical transplantation. The adenoviral vector, AdHCMVsp1LacZ, containing an expression cassette of Escherichiae coli lacZ, was used to perfuse heart grafts during cold ischemia before transplantation. Heart grafts were perfused with University of Wisconsin (UW) solution containing either 0 pfu, 5×1010 pfu, or 1×1011 pfu of viral vector, and were preserved for either 2 or 4 h and then transplanted into syngeneic recipients. The animals were killed at 1, 7, and 14 days after transplantation. The infection rate was assessed by histochemical staining for β-galactosidase. Using polymerase chain reaction (PCR), viral DNA presence was confirmed in every graft perfused with viral vectors. The protein production from the transfected gene was confirmed by a functional protein assay. An efficient gene transfer was achieved with an infection rate of 1%–1.5% for all cardiac myocytes, as assessed by 5-bromo-4-chloro-indolyl-β-d-galactopyranoside (X-gal) staining. All studies were negative in the control grafts. Gene expression persisted for at least 10 days after transplantation. We thus conclude that an efficient adenovirus-mediated gene transfection and expression of gene products can be achieved in ex vivo perfusion of the heart graft during cold preservation.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00311668
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  • 2
    Digitale Medien
    Digitale Medien
    Cancer and gene therapy (1996)
    Springer
    Annals of hematology 73 (1996), S. 207-218 
    Details
    ISSN: 1432-0584
    Schlagwort(e): Key words Cancer ; gene therapy ; gene transfer ; review
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Medizin
    Notizen: Abstract  The authors review the current literature pertaining to gene therapy as related to human cancer, covering gene therapy strategies, techniques of gene transfer, and targeted gene delivery. Gene therapy has various applications. Many technical obstacles in gene delivery need to be overcome. The safe delivery and expression of a gene to its destination are essential for clinical use. A greater understanding of the genetic basis of cancer and tumor immunology is necessary before the goal of cancer gene therapy can be fully realized.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002770050231
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  • 3
    Digitale Medien
    Digitale Medien
    Expression of the murine wild-type tyrosinase gene in transgenic rabbits (1996)
    Springer
    Transgenic research 5 (1996), S. 405-411 
    Details
    ISSN: 1573-9368
    Schlagwort(e): breeding control ; colour marker ; gene transfer ; pigmentation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The tyrosinase gene is known to be essential for melanization and has been shown to rescue pigmentation in albino mice. Previously we have described the strict copy-number-dependent expression of a murine wild-type tyrosinase gene construct over several generations in transgenic mice. In this study, we analysed the same gene construct as a marker gene for the transmission and expression of transgenes in rabbits. Using an albino hybrid strain, we produced transgenic rabbits expressing the murine tyrosinase gene. Strict correlation between integration and expression of the transgene and stable germline transmission of the integrated gene construct according to the Mendelian pattern of inheritance was observed. Thus, breeding control was facilitated by simple phenotypic examination of the transgenic animals. In contrast to mice transgenic for the same gene construct, tyrosinase-transgenic rabbits showed a greater variety in hue, intensity and extent of coat pigmentation, which is caused by the diversity in the loci affecting the melanization. Benefits and limitations of tyrosinase as a marker gene for the detection of homozygous individuals in the albino hybrid strain used are discussed.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01980205
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  • 4
    Digitale Medien
    Digitale Medien
    Selection of transgenic flax plants is facilitated by spectinomycin (1996)
    Springer
    Transgenic research 5 (1996), S. 131-137 
    Details
    ISSN: 1573-9368
    Schlagwort(e): Agrobacterium tumefaciens ; hypocotyl ; gene transfer ; flax ; neomycin ; spectinomycin
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Regeneration of transformed flax shoots after inoculation withAgrobacterium tumefaciens carrying a binary vector with either a neomycin phosphotransferase (nptII) gene and a β-glucuronidase (GUS) reporter gene or a spectinomycin resistance gene was examined. Hypocotyls from 4-day-old seedlings were inoculated with either of the twoA. tumefaciens strains. Selection and regeneration were achieved on a medium containing 0.1 μM thidiazuron, 0.01 μM napthalene acetic acid, 100 mgl−1 kanamycin sulphate or spectinomycin sulphate and 300 mgl−1 cefotaxime. Use of different neomycins for the selection of transformed tissues did select transformed calli but not transformed shoots either directly or via a callus phase. Selection based on spectinomycin resistance allowed the growth of transformed shoots. Transgenic shoots were rooted on a medium containing 100 mgl−1 spectinomycin sulphate. Integration of the spectinomycin resistance gene into the flax genome was confirmed by Southern blot hybridizations and spectinomycin resistance was shown to be inherited as a dominant Mendeliant trait. Therefore, spectinomycin resistance is more suitable for genetic engineering of flax than aminoglycoside resistance.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01969431
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  • 5
    Digitale Medien
    Digitale Medien
    Mammalian artificial chromosomes: A review (1996)
    Springer
    Cytotechnology 21 (1996), S. 253-261 
    Details
    ISSN: 1573-0778
    Schlagwort(e): Centromeres ; telomeres ; molecular vectors ; gene transfer ; gene therapy
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Medizin , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Abstract A mammalian artificial chromosome (MAC) may be assembled through the juxtapposition of three kinds of DNA elements: a centromere, several DNA replication origins, and two telomeric repeats. The resulting structure should be able to carry and express one or more selected genes (transgenes), introduced for specific purposes. The minimal length is unknown, but may be of several Mb. Of its basic elements, the telomeres may present lesser problems, in view of their simple composition and organization. Centromeres could be an issue, given their many unknowns. Mammalian DNA replication origins are at present poorly characterized, but it is expected that at least one may be contained within the MAC components, especially the transgene. Their overall assembly may require a combination of in vivo and in vitro approaches. A promising strategy aims at constructing two telomeric arms of a MAC, one of which may include the transgene. The two novel arms could acquire a functional centromere through recombination with the two arms of a resident chromosome. Alternatively, if the two telomeric constructs are also endowed with properly placed and oriented centromeric sequences, a centromere may be rescued in vivo by homologous recombination with the external parts of the centromere of the resident chromosome. Positive selection for the artificial arms and counterselection against the resident arms should facilitate the assembly process. The assembly of such construct would not change the ploidy number of the host cell. After loading of a transgene, however, the resulting MAC may be isolated and transferred into an expression cell, where it may represent a novel chromosomal element. In this case untoward effects to the host cell may derive from an ensuing dosage effect for the transgene(s) rather than from the presence of a MAC per se. A MAC may contribute to a deeper understanding of the structural requirements for chromosomal function and evolution as well as the mechanism of chromatin formation. It should also help in the development of second generation vectors for transfer of Mb-long DNA sequences, as required for properly regulated mammalian gene function as well as, possibly, for therapy.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00365348
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  • 6
    Digitale Medien
    Digitale Medien
    A Candidate for Cancer Gene Therapy: MIP-lα Gene Transfer to an Adenocarcinoma Cell Line Reduced T\imorigenicity and Induced Protective Immunity in Immunocompetent Mice (1996)
    Springer
    Pharmaceutical research 13 (1996), S. 1896-1901 
    Details
    ISSN: 1573-904X
    Schlagwort(e): gene transfer ; MIP-lα ; chemokine ; protective immunity
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Chemie und Pharmazie
    Notizen: Abstract Purpose. To evaluate the possibility of cancer gene therapy by the gene delivery of chemokine, the effects of human macrophage inflammatory protein lα (hu-MIP-lα), murine-macrophage inflammatory protein lα (mu-MIP-lα), and human-interleukin 8 (hu-IL-8) on tumor progression and immunization were studied. Methods. Cachexia-inducing and highly tumorigenic adenocarcinoma cells (cell line colon 26, clone 20) were transfected with either a control plasmid, hu-MIP-lα, mu-MIP-lα, or hu-IL-8 expression vector. The production of hu-MIP-1α reached 〉1.5 ng/ml in vitro when transfectant cells were cultured at a cell density of 2 × 105 cells in 7 ml for 3 days. Immunocompetent BALB/c mice were inoculated into the footpad with the tumor cells, and then primary tumor growth, morphological analyses, and tumor immunogenicity were studied. Results. The secretion of hu-MIP-lα, mu-MIP-lα, and hu-IL-8 did not affect the growth rate in vitro. Reduced tumorigenicities in vivo were observed in transfected cells with hu-MIP-lα and mu-MIP-lα. Morphologic observation of the site of inoculation of cells transfected with hu-MIP-lα showed infiltration of macrophages and neutrophils on the 5th day after the inoculation. Mice that had rejected cells transfected with hu-MIP-lα gene were immune to a subsequent challenge with the parental cells. Conclusions. The rejection of the cells depends on cytolysis and generates potent and long lasting antitumor immunity. These data suggest that tumor cells transfected with the MIP-lα gene might be useful as an effective therapy for the treatment of certain tumors.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1016057830271
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  • 7
    Digitale Medien
    Digitale Medien
    New Cationic Lipid Formulations for Gene Transfer (1996)
    Springer
    Pharmaceutical research 13 (1996), S. 1856-1860 
    Details
    ISSN: 1573-904X
    Schlagwort(e): gene transfer ; gene therapy ; cationic lipid ; transfection
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Chemie und Pharmazie
    Notizen: Abstract Purpose. To develop appropriate dosage forms of DNA for gene delivery. Methods. 3β[N-(N′, N′ dimethylaminoethane) carbamoyl] cholesterol (DC-Chol) was mixed either with Tween 80 alone, or with additional lipid components including castor oil and phosphatidylcholine (PC) or dioleoylphosphatidylethanolamine (DOPE) to make different lipid formulations. The particle size and the physical stability of the formulations upon mixing with plasmid DNA containing the luciferase cDNA were examined using laser light scattering measurement. The transfection activity of the DNA/lipid complexes was tested in presence or absence of serum using a cell culture system. Results. We demonstrated that many favorable properties as a gene carrier could be achieved by formulating DNA into new dosage forms using Tween 80 as the major emulsifier. Compared to the cationic liposomes, these new formulations transfected different cell lines with an equivalent or higher efficiency. Not only are they resistant to serum, but also form stable DNA complexes which could be stored for longer periods of time without losing transfection activity. Conclusions. Cationic lipids formulated into different lipid formulations using Tween 80 as a surfactant appeared to have more favorable physical and biological activities than traditional cationic liposomes as a carrier for gene delivery.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1016041326636
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  • 8
    Digitale Medien
    Digitale Medien
    Effect of Non-Ionic Surfactants on the Formation of DNA/Emulsion Complexes and Emulsion-Mediated Gene Transfer (1996)
    Springer
    Pharmaceutical research 13 (1996), S. 1642-1646 
    Details
    ISSN: 1573-904X
    Schlagwort(e): emulsions ; gene transfer ; transfection ; gene therapy ; non-ionic surfactant ; cationic lipid
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Chemie und Pharmazie
    Notizen: Abstract Purpose. To study the structure-function relationship of non-ionic surfactants in emulsion-mediated gene delivery. Methods. Four different types of non-ionic surfactants including Tween, Span, Brij and pluronic copolymers were used as co-emulsifiers for preparation of emulsions composed of Castor oil, dioleoylphosphatidylethanolamine (DOPE) and 3β[N-(N′, N′-dimethylaminoethane) carbamoyl] cholesterol (DC-Chol). The effect of different surfactants on the formation of DNA/emulsion complexes and transfection activity were analyzed using plasmid DNA containing luciferase cDNA as a reporter gene. Results. Non-ionic surfactants containing branched polyoxyethylene chains as the hydrophilic head group were more effective in preventing the formation of large DNA/emulsion complexes than those containing one or no polyoxyethylene chain. All emulsion formulations except those containing Brij 700 exhibited high activity in transfecting mouse BL-6 cells in the absence of serum. In the presence of serum, however, transfection activity of each formulation varied significantly. Emulsions containing Tween, Brij 72, pluronic F68 and F127 demonstrated increased activity in transfecting cells in the presence of 20% serum. In contrast to emulsions containing Span, long chain polyoxyethylene of Brij showed decreased transfection activity. The particle size of the DNA/emulsion complexes and their ability to transfect cells are dependent on the concentration of non-ionic surfactant in the formulation. Conclusions. The structure of the hydrophilic head group of the non-ionic surfactants in the emulsion is important in determining how DNA molecules interact with emulsions and the extent to which DNA is transferred inside the cell.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1016480421204
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  • 9
    Digitale Medien
    Digitale Medien
    Immunotherapy I: Cyclosine gene transfer strategies (1996)
    Springer
    Cancer and metastasis reviews 15 (1996), S. 317-328 
    Details
    ISSN: 1573-7233
    Schlagwort(e): cytokine ; tumor ; gene transfer ; immunotherapy
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Medizin
    Notizen: Abstract The cytokine approach to gene therapy of cancer stems from early studies of direct, repeated injection of recombinant cytokines at the tumor site, and extension of the bystander effect that enables a few cytokine gene transduced cells in a tumor to bring about its total destruction. This effect can be extended through the immune system, since cytokine-activated regression of a small mass of tumor cells can afford systemic protection. Transduced cells used as a vaccine provide a local concentration of both cytokine and tumor antigens. Cytokines sustain antigen uptake and presentation by increasing the immunogenic potential of the environment through the recruitment of antigen presenting cells and leukocytes, and activation of a cascade of events which amplify and tone up the efficacy of a vaccine. The promises and difficulties of this approach are discussed by considering what is still missing from experimental studies and what can best be done as soon as possible in animals and humans to reach compelling conclusions.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00046345
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  • 10
    Digitale Medien
    Digitale Medien
    Transgenic insect cells: mosquito cell mutants and the dihydrofolate reductase gene (1996)
    Springer
    Cytotechnology 20 (1996), S. 23-31 
    Details
    ISSN: 1573-0778
    Schlagwort(e): gene transfer ; transgenic insect cells ; Aedes albopticus ; dihydrofolate reductase
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Medizin , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00350386
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  • 11
    Digitale Medien
    Digitale Medien
    Membrane adsorption characteristics determine the kinetics of flow-through transductions (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 260-270 
    Details
    ISSN: 0006-3592
    Schlagwort(e): retrovirus ; gene therapy ; gene transfer ; virus adsorption ; membranes ; Chemistry ; Biochemistry and Biotechnology
    Quelle: Wiley InterScience Backfile Collection 1832-2000
    Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Retrovirus-mediated gene transfer is currently limited by random Brownian motion of the retrovirus. This limitation can be overcome by flowing the retrovirus solution through a porous membrane that supports the target cells, leading to a significant increase in the transduction efficiency. This procedure is termed “flow-through transduction.” In this study, we characterized the effects of the fluid flowrate and the influence that membrane characteristics have on the flow-through transduction procedure. The transduction efficiencies increased with flowrate until a plateau was reached at average flow velocities exceeding 0.3 cm/h for flow times of 3 to 4 h, using a collagen-coated depth (COL) membrane. A correlation between the optimal time for maximal gene transfer using flow-through transductions and the optimal time for maximal virus activity on the membrane was found, suggesting that the membrane adsorption capacity for virus determined the amount of gene transfer that could occur.Membrane adsorption characteristics were further investigated using two different membrane types: a tracketched polyester screen (PE) membrane and the COL membrane. Flow-through transductions using the PE and COL membranes showed that a high level of gene transfer could be attained using the COL membrane while the PE membrane gave much lower transduction efficiencies. The addition of the polycation polybrene (PB) changed these results markedly, making transductions achieved on the PE membrane similar in number to those obtained on the COL membrane. Since PB is believed to influence the virus adsorption to PE membrane, these results further support the conclusion that the increase in gene transfer achieved by the flow-through transduction procedure is due to virus adsorption to the membrane. The flow-through transduction procedure thus leads to co-localization of the viral vector and the target cell that in turn leads to a high transduction efficiency. © 1996 John Wiley & Sons, Inc.
    Zusätzliches Material: 9 Ill.
    Materialart: Digitale Medien
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  • 12
    Digitale Medien
    Digitale Medien
    Release of genetically modified micro-organisms into the environment (1996)
    Chichester : Wiley-Blackwell
    Journal of Chemical Technology AND Biotechnology 65 (1996), S. 5-14 
    Details
    ISSN: 0268-2575
    Schlagwort(e): GMOs ; environment ; release ; genetic engineering ; gene transfer ; Chemistry ; Biochemistry and Biotechnology
    Quelle: Wiley InterScience Backfile Collection 1832-2000
    Thema: Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Biotechnology in general, and recombinant DNA technology in particular, has the capacity to change the health and wealth of every individual, but like other major advances in science and technology such as nuclear power and electronics it can also be exploited to mankind's detriment. For this reason and the fact that recombinant DNA technology involves altering the molecules encoding life itself, the subject has given rise to a high level of public debate. Centuries of experience with the release of conventional micro-organisms for sewage treatment, agriculture and food production have shown that the release of large numbers of foreign organisms into an environment does not necessarily cause ecological damage. In fact few of these organisms survive for long periods. The threat of horizontal gene transfer from recombinant organisms to indigenous ones is however very real and mechanisms exist whereby, at least theoretically, any genetically engineered trait can be transferred to any prokaryotic organism and many eukaryotic ones. The rapid spread of antibiotic-resistant organisms since the widespread introduction of antibiotics graphically demonstrates that such a threat is real. There have now been several experiments to determine the effect of environmental release of micro-organisms, both in enclosed areas and in unenclosed sites. Proposals for the use of genetically modified micro-organisms that contain specific gene deletions have had a relatively smooth passage through government approval agencies and public enquiries and some products have been approved for commercial use. In addition a strain of bakers' yeast with an altered control element has also been approved for commercial use in the UK. Genetically modified viruses, especially those containing foreign genes have not received such favourable treatment and have been the subject of heated national and international debate. Many microbiologists are convinced, however, that the use and release of carefully constructed genetically engineered organisms will result in significant benefit, but with little risk to the environment. Ecologists, however, are not so sanguine and in the current ‘green’ political atmosphere their opinions are very influential. Thus most developed countries now have in force a series of very cautious regulations and guidelines for the release of genetically modified micro-organisms.
    Materialart: Digitale Medien
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  • 13
    Digitale Medien
    Digitale Medien
    Shoot apical meristems as a target for gene transfer by microballistics (1955)
    Springer
    Euphytica 85 (1955), S. 45-51 
    Details
    ISSN: 1573-5060
    Schlagwort(e): meristem ; shoot apex ; ballistic microtargeting ; gene transfer ; wheat
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary The classical approach of gene transfer to a given plant species delivers the foreign gene to transformable cells and then puts the effort into generating plants. This approach is very difficult in many important crop plants, including cereals, and the results of regeneration are very genotype-dependent. In contrast, we use regenerable cells and try to transform them. Shoot apical meristems provide a tissue which regenerates in situ a fertile plant for most given genotypes or species. Transformation of meristem cells may lead to transgenic sectors in chimeras. These sectors may contribute to the gametes and, thus, to transgenic offspring, which then should be homohistonts and not sectorial chimeras like their parents. Our model plant for these studies is wheat. Microtargeting is a ballistic approach which is particularly suitable for the controlled delivery of microprojectiles to meristem cells in situ (Sautter et al., 1991). We summarize in this paper our experience with ballistic microtargeting of transgenes to wheat shoot apical meristem cells in situ.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023929
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  • 14
    Digitale Medien
    Digitale Medien
    Transgenic barley by particle bombardment. Inheritance of the transferred gene and characteristics of transgenic barley plants (1955)
    Springer
    Euphytica 85 (1955), S. 81-88 
    Details
    ISSN: 1573-5060
    Schlagwort(e): gene transfer ; Hordeum vulgare ; neomycin phosphotransferase II ; particle bombardment ; transgenic barley
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Transgenic barley plants (Hordeum vulgare L. cv. Kymppi) were obtained by particle bombardment of various tissues. Immature embryos and microspore-derived cultures were bombarded with gold particles coated with plasmid DNA carrying the gene coding for neomycin phosphotransferase II (NPTII), together with plasmid DNA containing the gene for β-glucuronidase (GUS). Bombarded immature embryos were grown to plants without selection and NPTII activity was screened in small plantlets. One plant proved to be transgenic (T0). This chimeric plant passed the transferred nptII gene to its T1 progeny. The presence of the nptII gene was demonstrated by the PCR technique and enzyme activity was analyzed by an NPTII gel assay. Four T0 spikes and 15 T1 offspring were transgenic. The integration and inheritance was confirmed by Southern blot hybridization. Transgenic T2 and T3 plants were produced by isolating embryos from green grains of transgenic T1 and T2 plants, respectively and growing them to plants. After selfing, the ratio of transgenic to non-transgenic T2 offspring was shown to follow the rule of Mendelian inheritance. The general performance of transgenic plants was normal and no reduction in fertility was observed. Microspore-derived cultures were bombarded one and four weeks after microspore isolation. After bombardment, cultures were grown either with or without antibiotic selection (geneticin R or kanamycin). When cultures were grown without selection and regenerated plants were transferred to kanamycin selection in rooting phase, one out of a total of about 1500 plants survived. This plant both carried and expressed the transferred nptII gene. The integration was confirmed by Southern blot hybridization. This plant was not fertile.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023933
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  • 15
    Digitale Medien
    Digitale Medien
    The sulphur-rich Brazil nut 2S albumin is specifically formed in transgenic seeds of the grain legume Vicia narbonensis (1955)
    Springer
    Euphytica 85 (1955), S. 181-192 
    Details
    ISSN: 1573-5060
    Schlagwort(e): Vicia narbonensis ; gene transfer ; gene expression ; seeds ; 2S albumin ; methionine
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Epicotyl explants were co-cultivated with Agrobacterium tumefaciens EHA101 to transfer a chimeric 2S albumin gene construct carried in the binary Ti plasmid vectors pGSGLUC1 or pGA472 into the grain legume Vicia narbonensis. This gene encoding the sulphur-rich Brazil nut albumin was under the control of either the CaMV 35S promoter which permits gene expression in all organs, or the Vicia faba legumin B4 promoter which elicits seed-specific gene expression. After callus formation and selection for kanamycin resistance, somatic embryos were induced which, in the case of transformation with the vector pGSGLUC1, were screened for GUS activity. Embryos that produced GUS were in addition analysed for 2S albumin formation. Selected transgenic embryos were cloned by multiple shoot regeneration. Rooted and fertile plants were obtained by grafting transgenic shoots on the appropriate seedlings. R1 and R2 generations were raised and analysed for GUS as well as 2S albumin gene expression. Expression of the 35S promoter/2S albumin gene fusion took place in all organs of the transgenic plants including the cotyledons of seeds, whereas seed-specific gene expression was found in transformants with the legumin promoter/2S albumin gene fusion. The 2S albumin accumulated in the 2S protein fraction of transgenic seeds and its primary translation product was processed into the 9 and 3 kDa polypeptide chains. The foreign protein was localised in the protein bodies of the grain legume. Analysis of the R2 plants indicated Mendelian inheritance of the 2S albumin gene. In homozygous V. narbonensis plants the amounts of 2S albumin were twice that present in the corresponding heterozygous plants. Whereas only low level formation of the foreign protein was achieved if the gene was under the control of the 35S promoter, approximately 3.0% of the soluble seed protein was 2S albumin if seed-specific gene expression was directed by the legumin B4 promoter. Some of these transformants exhibited a three-fold increase in the methionine content of the salt-soluble protein fraction extracted from seeds.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023947
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  • 16
    Digitale Medien
    Digitale Medien
    Strategies for variety-independent genetic transformation of important cereals, legumes and woody species utilizing particle bombardment (1955)
    Springer
    Euphytica 85 (1955), S. 13-27 
    Details
    ISSN: 1573-5060
    Schlagwort(e): gene transfer ; crop species ; particle bombardment ; transgenic plants ; cereals ; legumes ; woody plants
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary The limiting component in the creation of transgenic crops has been the lack of effective means to introduce foreign genes into elite germplasm. However, the development of novel direct DNA transfer methodology, by-passing limitations imposed by Agrobacterium-host specificity and cell culture constraints, has allowed the engineering of almost all major crops, including formerly recalcitrant cereals, legumes and woody species. The creation of transgenic rice, wheat, maize, barley, oat, soybean, phaseolus, peanut, poplar, spruce, cotton and others, in an efficient and in some cases, variety-independent fashion, is a significant step towards the routine application of recombinant DNA methodology to the improvement of most important agronomic crops. In this review we will focus on key elements and advantages of particle bombardment technology in order to evaluate its impact on the accelerated commercialization of products based on agricultural biotechnology and its utility in studying basic plant developmental processes and function through transgenesis. Fundamental differences between conventional gene transfer methods, utilizing Agrobacterium vectors or protoplast/suspension cultures, and particle bombardment will be discussed in depth.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023926
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