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  • 1
    Digitale Medien
    Digitale Medien
    A Modular Vector for Agrobacterium Mediated Transformation of Wheat (1999)
    Springer
    Plant molecular biology reporter 17 (1999), S. 323-331 
    Details
    ISSN: 1572-9818
    Schlagwort(e): Agrobacterium ; modular vector ; transformation ; wheat
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Wheat (cv Chinese Spring) tissues were transformed using Agrobacterium tumefasciens and a new plasmid modular vector, pMVTBP. We constructed pMVTBP with unique restriction sites connecting (1) the CaMV 35S promoter, (2) a Kozak sequence, (3) the FLAG epitope, (4) the (His)6 epitope, (5) a coding region (for wheat TATA Binding Protein, wTBP) and (6) the CaMV 35S 3′UTR. This vector thus allows easy exchange of different regulatory or coding sequences. Explants of either germinating mature seeds, or immature embryos, were induced to callus for up to two weeks, treated with virulence-induced bacteria for one hour, then regenerated into plantlets. Transient expression of a GUS reporter gene, assayed at about one week, occurred in 10–12% of calluses. Expression of the FLAG-tagged wTBP was also detected, by immunostaining. Stable expression, by selective growth on geneticin, and by GUS expression at about six weeks, occurred in 1–2% of calluses, quite comparable to that achieved by other methods.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1007686408369
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  • 2
    Digitale Medien
    Digitale Medien
    Transgenic almond (Prunus dulcis Mill.) plants obtained by Agrobacterium-mediated transformation of leaf explants (1999)
    Springer
    Plant cell reports 18 (1999), S. 387-393 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Almond ; Prunus ; Transformation ; Agrobacterium ; Adventitious regeneration
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Almond (Prunus dulcis Mill.) leaves were transformed with the marker genes gusA (β-glucuronidase) and nptII (neomycin phosphotransferase II) via Agrobacterium-mediated transformation. Bacterial strains and preculture of explants affected efficiency of gene transfer evaluated by transient expression assays. Following transformation, shoots were induced from primary explants on medium without kanamycin and exposed to selection 20 days after cocultivation. From 1419 original leaves, four shoots (A, B, C and D) were obtained that showed amplification of the predicted DNA fragments by polymerase chain reaction (PCR). After micropropagation of these shoots, only those cloned from shoot D gave consistently positive results in histochemical GUS detection and PCR amplification. Southern blot hybridisation confirmed stable transgene integration in clone D, which was also negative in PCR amplification of an Agrobacterium gene. Additional molecular analysis suggested that the remaining three shoots (A, B and C) were chimeric.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050591
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  • 3
    Digitale Medien
    Digitale Medien
    Green-fluorescent protein facilitates rapid in vivo detection of genetically transformed plant cells (1999)
    Springer
    Plant cell reports 18 (1999), S. 707-714 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Green-fluorescent protein ; Transformation ; Particle bombardment ; Agrobacterium ; Sugarcane
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Early detection of plant transformation events is necessary for the rapid establishment and optimization of plant transformation protocols. We have assessed modified versions of the green fluorescent protein (GFP) from Aequorea victoria as early reporters of plant transformation using a dissecting fluorescence microscope with appropriate filters. Gfp-expressing cells from four different plant species (sugarcane, maize, lettuce, and tobacco) were readily distinguished, following either Agrobacterium-mediated or particle bombardment-mediated transformation. The identification of gfp-expressing sugarcane cells allowed for the elimination of a high proportion of non-expressing explants and also enabled visual selection of dividing transgenic cells, an early step in the generation of transgenic organisms. The recovery of transgenic cell clusters was streamlined by the ability to visualize gfp-expressing tissues in vitro.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050647
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  • 4
    Digitale Medien
    Digitale Medien
    Stable genetic transformation of white pine (Pinus strobus L.) after cocultivation of embryogenic tissues with Agrobacterium tumefaciens (1999)
    Springer
    Molecular breeding 5 (1999), S. 429-440 
    Details
    ISSN: 1572-9788
    Schlagwort(e): transgenic trees ; scaffold attachment region ; Agrobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract A genetic transformation procedure for white pine has been developed after cocultivation of embryogenic tissues with Agrobacterium tumefaciens. This efficient transformation procedure led to an average of four independent transformed lines per gram of cocultivated embryogenic tissue and up to 50 transformed lines can be obtained in a routine experiment. Constructs bearing the uidA gene or the green fluorescent protein (GFP) gene were introduced and β-glucuronidase (GUS) activity was followed over time. The expression of the uidA gene was lowest with a 35S-gus-intron construct and was 20-fold higher with a 35S-35S-AMVgus::nptII construct. The addition of scaffold attachment region (SAR) sequences surrounding the gus::nptII fusion did not significantly enhance the GUS activity. Transformed mature somatic embryos have been germinated and plantlets are presently being acclimatized.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1009683605841
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  • 5
    Digitale Medien
    Digitale Medien
    Positive-negative selection and T-DNA stability in Arabidopsis transformation (1999)
    Springer
    Plant molecular biology 39 (1999), S. 83-93 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Arabidopsis ; Agrobacterium ; T-DNA ; CodA ; positive-negative selection ; recombination
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract We have analysed the application of positive-negative selection for the selection of homologous recombination interactions between the chromosome and a T-DNA molecule after transformation of plant cells. Two different genomic loci in a cell suspension of Arabidopsis thaliana were chosen to study gene targeting events. One was the chalcone synthase (CHS) gene present as a single copy and the second an hemizygous chromosomally inserted T-DNA containing the hpt gene, conferring resistance to hygromycin, flanked by CHS sequences. The target lines were transformed with replacement-type T-DNA vectors which contained a positive selectable marker flanked by the regions of the CHS gene and a negative selectable marker to counter-select random insertions. As negative marker we used the Escherichia coli codA gene encoding cytosine deaminase, conferring upon the cells sensitivity to 5-flourocytosine (5-FC). Doubly selected transformants represent 1–4% of the primary transformed cells. Targeting events were not found at the chalcone synthase locus nor at the artificial hpt locus in a total of 4379 doubly selected calli, corresponding to at least 109 475 individual primary transformants. We show by PCR and Southern analysis that the 5-FC resistance in the majority of these cells is associated with substantial deletions of the T-DNA molecule from the right-border end.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1006192225464
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  • 6
    Digitale Medien
    Digitale Medien
    Sequence and functional analysis of the left-hand part of the T-region from the nopaline-type Ti plasmid, pTiC58 (1999)
    Springer
    Plant molecular biology 41 (1999), S. 765-776 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Agrobacterium ; Ti plasmid ; oncogenes ; e gene ; 3′ gene
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The Agrobacterium tumefaciens nopaline strain C58 transfers a large, 29 kb T-DNA into plant cells during infection. Part of this DNA (the `common DNA') is also found on the T-DNA of octopine strains, the remaining DNA is nopaline strain-specific. Up to now, only parts of the C58 T-DNA and related T37 T-DNA have been sequenced. We have sequenced the remainder of the nopaline-specific T-DNA (containing genes a to d) and acs to iaaM. Gene c codes for a new unknown T-DNA protein. Gene a is homologous to the agrocinopine synthase gene. Genes b, c′, d and e are part of a larger family: they are related to the T-DNA genes 5, rolB, lso and 3′. Genes 5, rolB and lso induce or modify plant growth and have been called T-DNA oncogenes. Our studies show that gene 3′ (located on the TR-DNA of octopine strains) is also oncogenic. Although the b–e T-DNA fragment from C58 and its individual genes lack growth-inducing activity, an a-acs deletion mutant was distinctly less virulent on Kalanchoe daigremontiana and showed reduced shoot formation on Kalanchoe tubiflora. Shoot formation could be restored by genes c and c′ in co-infection experiments. Contrary to an earlier report, a C58 e gene deletion mutant was fully virulent on all plants tested.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1006370207379
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  • 7
    Digitale Medien
    Digitale Medien
    Evidence of Migration and Endophytic Presence of Agrobacterium tumefaciens in Rose Plants (1999)
    Springer
    European journal of plant pathology 105 (1999), S. 39-50 
    Details
    ISSN: 1573-8469
    Schlagwort(e): Agrobacterium ; crown gall ; systemic infection ; rose ; endophyte
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Agrobacterium tumefaciens was isolated from stem tumors of several rose cultivars showing that the bacterium is the causal agent of aerial galls in rose plants. No differences were observed in the characteristics of the Agrobacterium isolates from crown or aerial galls. Stem inoculation of ten rose cultivars showed that all of them were susceptible to A. tumefaciens but differences in the size of the resulting tumors were observed. The movement of A. tumefaciens in rose plants was demonstrated using two wild type strains and two antibiotic resistant mutants. Three months after inoculation, the inoculated strains were recovered in the roots, crown and below and above the inoculation site but low numbers of pathogenic Agrobacterium cells were isolated. New tumors appeared in 5% of the noninoculated wounds. A. tumefaciens was isolated from the stem at different distances from the tumor in naturally infected plants. In symptomless commercial plants, the isolation from the roots, crown and at different stem levels demonstrated the existence of systemic and latent infections in rose. Direct isolation using a nonselective and selective media with or without a previous enrichment step were efficient methods for isolating tumorigenic Agrobacterium from the different parts of rose plants.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1008660500107
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  • 8
    Digitale Medien
    Digitale Medien
    Origin and evolution of plasmids (1998)
    Springer
    Antonie van Leeuwenhoek 73 (1998), S. 117-126 
    Details
    ISSN: 1572-9699
    Schlagwort(e): riboplasmids ; encapsidation ; pseudovirions ; selfish plasmids ; replicons ; ribozyme ; Agrobacterium ; Rhizobium ; grapevines ; L-tartrate ; lignin ; methoxyphenols ; satellite viruses ; opines ; crown gall ; T-DNA ; origin of replication
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Studies on the origin and evolution of plasmids may provide valuable insights on the promiscuous nature of DNA. The first examples of the selfish nature of nucleic acids are exemplified by primordial oligoribonucleotides which evolved into primitive replicons. The propagation of these molecules were likely patterned after the current viral RNA ribozymes, which have been recently shown to possess RNA synthesizing and template mediated polymerizing capabilities in the absence of proteins. The parasitic nature of nucleic acids is depicted by satellite nucleic acid molecules associated with viruses. The satellites of adenovirus and tobacco ringspot virus serve as established examples: they contain no open reading frames. Comparative analysis of the replication origins of virions and plasmids show them to be conserved, originating from the simplest autocatalytic replicon to highly complex and evolved plasmids, replicating by a rolling circle mechanism. The eventual association of proteins with nucleic acids provided added efficiency and protective advantages for molecular perpetuation. The promiscuous and selfish nature of plasmids is demonstrated by their ability to genetically engineer their host so that the host cell is best able to cope and survive in hostile environments. Survival of the host ensures survival of the plasmid. Sequestering of genes by plasmids occurs when the environmental conditions negatively affect the host. The sequestering mechanism is fundamental and forms the outreach mechanisms to generate and propagate macromolecules of increasing size when necessary for survival. The level of sophistication of plasmids increases with the addition of new genes such as those that allow the host to occupy a specific environment normally inhospitable to the host cell. The vast range of plasmid types which have obtained genes interchangeably reflect the levels of sophistication achieved by these macromolecules. The Ti plasmid in Agrobacterium tumefaciens and the pSym and accessory plasmids in Rhizobium illustrate the level of complexity attained by replicons.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1000652513822
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  • 9
    Digitale Medien
    Digitale Medien
    Transformation and regeneration of Lobelia erinus using Agrobacterium tumefaciens (1998)
    Springer
    Plant cell reports 18 (1998), S. 308-311 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Campanulaceae ; Lobelia erinus ; Transformation ; Agrobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract A transformation/regeneration system was developed for the common garden Lobelia (Lobelia erinus). Using an Agrobacterium-based protocol, over 40 transformants have been generated with four different binary vectors. The explant source was hypocotyl-root sections from axenically grown seedlings. Stable transformation was demonstrated by Southern hybridization analysis, β-glucuronidase staining, and transmission of the T-DNA to progeny. This extends the ever-widening range of transformable plant species to the Campanulaceae and will allow molecular studies of development and physiology in this easily cultured and popular garden plant.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050577
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  • 10
    Digitale Medien
    Digitale Medien
    Factors affecting soybean cotyledonary node transformation (1998)
    Springer
    Plant cell reports 18 (1998), S. 180-186 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Soybean ; SAAT (sonication assisted Agrobacterium-mediated transformation) ; Agrobacterium ; Transformation ; KYRT1
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Cotyledonary node transformation efficiency was evaluated using a sonication assisted Agrobacterium-mediated transformation (SAAT) protocol, three dissimilar A. tumefaciens strains, and explants derived from 28 diverse cultivars and/or genotypes of soybean [Glycine max (L.) Merr.]. The explants were evaluated at 10 and 45 days after co-cultivation for transformation with a binary vector containing both a GUS-intron gene and an NPTII selectable marker. The best overall strain of A. tumefaciens was determined to be KYRT1 based on stable GUS transformation of soybean cotyledonary node explants measured at the terminal 45 day evaluation point. SAAT did not increase stable transformation at 45 days post co-cultivation. SAAT was determined to significantly decrease shoot proliferation of some genotypes, but it is unclear what effect this may have on the recovery of transformed shoots. Significant differences were also detected between genotypes for transformation and shoot proliferation frequency.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050553
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  • 11
    Digitale Medien
    Digitale Medien
    Transgenic peppermint (Mentha×piperita L.) plants obtained by cocultivation with Agrobacterium tumefaciens (1998)
    Springer
    Plant cell reports 17 (1998), S. 165-171 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Peppermint ; Transformation ; Agrobacterium ; GUS ; Transgenic
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The first transgenic peppermint (Mentha×piperita L. cultivar Black Mitcham) plants have been obtained by Agrobacterium-mediated transformation by cocultivation with morphogenically responsive leaf explants. Basal leaf explants with petioles, from leaves closest to the apex of in-vitro-culture-maintained shoots (5 cm), exhibited optimal shoot organogenetic responsiveness on medium supplemented with thidiazuron (8.4 µm). Shoot formation occurred at sites of excision on the leaf blade and petiole either directly from cells of the explant or via a primary callus. Analyses of transient GUS activity data indicated that DNA delivery by microprojectile bombardment was more effective than Agrobacterium infection. However, no transgenic plants were obtained from over 22,000 leaf explants after particle bombardment. Cocultivation of leaf explants with Agrobacterium strain EHA 105 and kanamycin selection produced transgenic plants. Greater transient and stable -glucuronidase (GUS) activities were detected in explants or propagules transformed with the construct where gusA was driven by the pBISN1 promoter rather than a CaMV 35S promoter. Eight plants were subsequently regenerated and verified as transgenic based on detection of the nptII transgene by PCR and Southern blot analyses. The Southern analyses indicated that the plants were derived from eight unique transformation events. All transgenic plants appeared morphologically normal. Analyses of GUS activities in leaves sampled from different portions of these transgenic plants, 10 months after transfer to the greenhouse, indicated that six out of the eight original regenerants were uniformly transformed, i.e., did not exhibit chimeric sectors.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050372
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  • 12
    Digitale Medien
    Digitale Medien
    Agrobacterium tumefaciens-mediated transformation of Eucalyptus camaldulensis and production of transgenic plants (1998)
    Springer
    Plant cell reports 17 (1998), S. 675-680 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Genetic transformation ; Agrobacterium ; Eucalyptus ; Regeneration
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract An efficient system for Agrobacterium-mediated transformation of Eucalyptus camaldulensis and production of transgenic plants was developed. Transformation was accomplished by cocultivation of hypocotyl segments with Agrobacterium tumefaciens containing a binary Ti-plasmid vector harboring chimeric neomycin phosphotransferase and β-glucuronidase (GUS) genes. A modified Gamborg's B5 medium used in this study was effective for both callus induction and regeneration of transgenic shoots. This medium could also effectively maintain the organogenic capability of callus for more than a year. Culturing transgenic shoots in Murashige and Skoog medium supplemented with 0.1 mg ⋅ l–1 benzylaminopurine prior to root induction in rooting medium markedly increased the rootability of shoots that were recalcitrant to rooting. Histochemical assay revealed the expression of the GUS gene in leaf, stem, and root tissues of transgenic plants. Insertion of the GUS gene in the nuclear genome of transgenic plants was verified by genomic Southern hybridization analysis, further confirming the integration and expression of T-DNA in these plants.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050464
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  • 13
    Digitale Medien
    Digitale Medien
    Activity of the CaMV 35S promoter in various parts of transgenic early flowering birch clones (1998)
    Springer
    Plant cell reports 18 (1998), S. 243-248 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key wordsBetula pendula ; Transformation ; Agrobacterium ; Gene expression
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Early flowering together with small size would be useful for various biotechnical or genetic studies on trees. We report here the selection and micropropagation of early flowering birch (Betula pendula) clones (BPM1–12) obtained from seeds of birches bred elsewhere for early flowering. Under conditions that accelerate flowering (a high CO2 level, strong and continuous illumination), the first male inflorescences emerged in 3–5 months, the trees then being 20–80 cm high. Transgenic lines (CaMV 35S-GUS INT) were produced through Agrobacterium-mediated gene transfer from BPM2, BPM5 and JR1/4 (a normally flowering birch). β-Glucuronidase (GUS) activities in the different lines, assayed 1–1.5 years after transformation, varied greatly. During further in vitro culture for 10 months, the activities decreased to 0.3–7% of the original values. GUS activities were detected in all organs studied, including the developing male inflorescences; the highest activity was in the roots.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050564
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  • 14
    Digitale Medien
    Digitale Medien
    Transformation of arctic bramble (Rubus arcticus L.) by Agrobacterium tumefaciens (1998)
    Springer
    Plant cell reports 17 (1998), S. 822-826 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Rosaceae ; β-Glucuronidase ; Regeneration ; Agrobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Genetic transformation of arctic bramble (Rubus arcticus L.) was achieved utilizing a Ti-plasmid vector system of Agrobacterium tumefaciens. Internodal stem segments were inoculated with Agrobacterium strain EHA101 carrying a T-DNA with the CaMV 35 S promoter-gus-int marker gene from which β-glucuronidase (GUS) is expressed only in plants. Regenerants were produced on Murashige and Skoog medium. Growth of Agrobacterium was inhibited with cefotaxime. Kanamycin was used as the selective agent for the transformants. Regenerants were assayed by histochemical GUS staining, and by Southern analysis using a gus-int probe. Transgenic arctic bramble plants containing gus-int and expressing GUS were recovered. Expression has been stable for 3 years in micropropagation.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050491
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  • 15
    Digitale Medien
    Digitale Medien
    CaMV 35S promoter directs β-glucuronidase expression in the laticiferous system of transgenic Hevea brasiliensis (rubber tree) (1998)
    Springer
    Plant cell reports 17 (1998), S. 621-625 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key wordsHevea brasiliensis ; Agrobacterium ; CaMV 35S ; β-Glucuronidase ; Latex
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Hevea brasiliensis anther calli were genetically transformed using Agrobacterium GV2260 (p35SGUSINT) that harboured the β-glucuronidase (gus) and neomycin phosphotransferase (nptII) genes. β-Glucuronidase protein (GUS) was expressed in the leaves of kanamycin-resistant plants that were regnerated, and the presence of the gene was confirmed by Southern analysis. GUS was also observed to be expressed in the latex and more importantly in the serum fraction. Transverse sections of the leaf petiole from a transformed plant revealed GUS expression to be especially enhanced in the phloem and laticifers. GUS expression was subsequently detected in every one of 194 plants representing three successive vegetative cycles propagated from the original transformant. Transgenic Hevea could thus facilitate the continual production of foreign proteins expressed in the latex.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002990050454
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  • 16
    Digitale Medien
    Digitale Medien
    Genetic transformation and regeneration of mature tissues of woody fruit plants bypassing the juvenile stage (1998)
    Springer
    Transgenic research 7 (1998), S. 51-59 
    Details
    ISSN: 1573-9368
    Schlagwort(e): sweet orange ; Citrus ; woody ; transformation ; Agrobacterium ; mature
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Regeneration and transformation systems from mature plant material of woody fruit species have to be achieved as a necessary requirement for the introduction of useful genes into specific cultivars and the rapid evaluation of resulting horticultural traits. We report here, for the first time, a procedure for genetic transformation and regeneration of mature tissues of woody plants that overcomes the long juvenile periods and high heterozygosity that are characteristic of most of these species. An improved regeneration frequency from mature explants was obtained by invigoration of the plant material through grafting of mature buds on juvenile seedlings. Co-cultivation of the explants in feederplates after inoculation with Agrobacterium tumefaciens resulted in enhanced transformation frequencies. Furthermore, in vitro shoot-tip grafting of the regenerated mature shoots on seedling rootstocks provided a rapid and efficient system for plant production. Citrus is the most extensivel y grown fruit crop worldwide and sweet orange (Citrus sinensis L. Osbeck) accounts for approximately 70% of the Citrus total production. Mature transgenic sweet orange plants have been obtained, which flowered and bore fruit in 14 months
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1008855922283
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  • 17
    Digitale Medien
    Digitale Medien
    An efficient protocol for sugarcane (Saccharum spp. L.) transformation mediated by Agrobacterium tumefaciens (1998)
    Springer
    Transgenic research 7 (1998), S. 213-222 
    Details
    ISSN: 1573-9368
    Schlagwort(e): Saccharum ; Agrobacterium ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract This is the first successful report of the recovery of morphologically normal transgenic sugarcane plants from co-cultivation of calluses with Agrobacterium tumefaciens. Transformation frequencies (total of transgenic plants/number of cell clusters) were between 9.4 × 10−3 and 1.15 × 10−2. In our experiments, both LBA4404 (pTOK233) and EHA101 (pMTCA3IG), carrying a super-binary vector or supervirulent strain, respectively, were successful for sugarcane transformation. We found that three main factors: (1) the use of young regenerable calluses as target explants; (2) induction and/or improvement of the A. tumefaciens virulence system with sugarcane cell cultures and (3) pre-induction of organogenesis or somatic-embryogenesis-like sexual embryos, seem to be crucial in order to increase the cells competence for T-DNA transfer process. Patterns generated by Southern hybridization confirmed that T-DNAs were randomly integrated into sugarcane genome without th e persistence of A. tumefaciens in the transgenic plants
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1008845114531
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  • 18
    Digitale Medien
    Digitale Medien
    Agrobacterium tumefaciens-mediated transformation of cauliflower, Brassica oleracea var. botrytis (1998)
    Springer
    Molecular breeding 4 (1998), S. 531-541 
    Details
    ISSN: 1572-9788
    Schlagwort(e): Agrobacterium ; Brassica oleracea ; cauliflower ; regeneration ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract We have developed an efficient and simpler method for genetic transformation and regeneration of cauliflower, Brassica oleracea var. botrytis plants. Explants from 4-day old seedlings were inoculated and cocultivated with Agrobacterium tumefaciens strain LBA4404 harbouring a binary vector with the neomycin phosphotransferase-II gene under the regulatory control of nopaline synthase promoter and terminator sequences, permitting transformed shoots to be selected on kanamycin containing medium. After three months rooted transformed plantlets were successfully transferred and grown under glasshouse conditions. Higher numbers of transformed plants were obtained from cotyledon than hypocotyl explants, presumably indicating cotyledons of cauliflower are more amenable to genetic transformation. Integration and expression of the introduced transgene were analysed by DNA gel blot and PCR analysis and NPT-II expression assay. Factors influencing transformation efficiency include explant age, concentration of bacterium used for infection, duration of infection and cocultivation with Agrobacterium. Transgenic plants of three commercial genotypes of cauliflower were produced using this method. We also show that introduction of antisense Bcp1 (pollen-specific gene) linked to a pollen-specific promoter (Lat52) resulted in the expected sterility of 50% pollen carrying this transgenic construct.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1009658614579
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  • 19
    Digitale Medien
    Digitale Medien
    Investigation of Agrobacterium-mediated transformation of apple using green fluorescent protein: high transient expression and low stable transformation suggest that factors other than T-DNA transfer are rate-limiting (1998)
    Springer
    Plant molecular biology 37 (1998), S. 549-559 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Agrobacterium ; apple ; GFP ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract To investigate early events of Agrobacterium-mediated transformation of apple cultivars, a synthetic green fluorescent protein gene (SGFP) was used as a highly sensitive, vital reporter gene. Leaf explants from four apple cultivars (‘Delicious’, ‘Golden Delicious’, ‘Royal Gala’ and ‘Greensleeves’) were infected with Agrobacterium EHA101 harboring plasmid pDM96.0501. Fluorescence microscopy indicated that SGFP expression was first detected 48 h after infection and quantitative analysis revealed a high T-DNA transfer rate. Plant cells with stably incorporated T-DNA exhibited cell division and developed transgenic calli, followed by formation of transgenic shoots at low frequencies. The detection of SGFP expression with an epifluorescence stereomicroscope confirmed the effectiveness of SGFP as a reporter gene for detection of very early transformation events and for screening of putative transformants. The efficiency of the transformation and regeneration process decreased ca. 10000-fold from Agrobacterium infection to transgenic shoot regeneration, suggesting that factors other than Agrobacterium interaction and T-DNA transfer are rate-limiting steps in Agrobacterium-mediated transformation of apple.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1006041313209
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  • 20
    Digitale Medien
    Digitale Medien
    The use of visual marker genes as cell-specific reporters of Agrobacterium-mediated T-DNA delivery to wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.). (1998)
    Springer
    Euphytica 99 (1998), S. 17-25 
    Details
    ISSN: 1573-5060
    Schlagwort(e): Agrobacterium ; barley ; C1/Lc ; GFP ; GUS ; wheat
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Transfer of T-DNA from Agrobacterium tumefaciens and A. rhizogenes to cells of wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) is demonstrated following the inoculation of immature embryos and immature embryo-derived callus. Agrobacterium T-DNA vectors containing the C1/Lc anthocyanin-biosynthesis regulatory genes, the gusA gene or a synthetic green fluorescent protein gene (sgfp-S65T) were constructed from original binary vectors. The visual T-DNA markers were used as cell-autonomous reporters of early Agrobacterium-mediated transformation events in the wheat and barley cells. This localization of the transformed cells revealed a non-random distribution throughout each embryo and callus piece.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1018303102488
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  • 21
    Digitale Medien
    Digitale Medien
    Cre/lox-mediated site-specific integration of Agrobacterium T-DNA in Arabidopsis thaliana by transient expression of cre (1998)
    Springer
    Plant molecular biology 38 (1998), S. 393-406 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Agrobacterium ; Arabidopsis ; Cre/lox ; recombination ; site-specific integration ; T-DNA
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The Cre/lox system was used to obtain targeted integration of an Agrobacterium T-DNA at a lox site in the genome of Arabidopsis thaliana. Site-specific recombinants, and not random events, were preferentially selected by activation of a silent lox-neomycin phosphotransferase (nptII) target gene. To analyse the effectiveness of Agrobacterium-mediated transfer we used T-DNA vectors harbouring a single lox sequence (this vector had to circularize at the T-DNA left- and right-border sequences prior to site-specific integration) or two lox sequences (this vector allowed circularization at the lox sequences within the T-DNA either prior to or after random integration, followed by targeting of the circularized vector), respectively. Furthermore, to control the reversibility of the integration reaction, Cre recombinase was provided transiently by using a cotransformation approach. One precise stable integrant was found amongst the recombinant calli obtained after transformation with a double-lox T-DNA vector. The results indicate that Agrobacterium-mediated transformation can be used as a tool to obtain site-specific integration.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1006024500008
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  • 22
    Digitale Medien
    Digitale Medien
    Transgenic Populus tremula: a step-by-step protocol for its Agrobacterium-mediated transformation (1997)
    Springer
    Plant molecular biology reporter 15 (1997), S. 219-235 
    Details
    ISSN: 1572-9818
    Schlagwort(e): Agrobacterium ; Populus ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract In recent years, Populus species have acquired an important place in basic and applied research of woody plants. The practical role of Populus species in world forestry and their importance to research as a woody-plant model have led to increasing interest in tissue-culture and molecular techniques, as well as the development of transformation procedures for this genus. A simple technical procedure is described here step-by-step, for the first time, as a routine method for transforming Populus tremula using a disarmed Agrobacterium tumefaciens hypervirulent strain. The procedure begins with the inoculation of stem explants with bacterial suspension, followed by a short period of co-cultivation on a highly regenerative medium. Transformed shoots are selected on regeneration medium containing antibiotics and the presence of the inserted target genes is checked using a rapid and efficient PCR test. Selected shoots are transferred to a rooting medium, under the same selection pressure, and propagated via stem cuttings. Selected plants can be hardened and transferred to the green-house within 4 months of inoculation. The method has proven efficient for several gene constructs, selection on Kan or Hyg, and three different Agrobacterium strains.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1007484917759
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  • 23
    Digitale Medien
    Digitale Medien
    Role of the host cell cycle in theAgrobacterium-mediated genetic transformation ofPetunia: Evidence of an S-phase control mechanism for T-DNA transfer (1997)
    Springer
    Planta 201 (1997), S. 160-172 
    Details
    ISSN: 1432-2048
    Schlagwort(e): Agrobacterium ; Leaf mesophyll cells ; Petunia ; Transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Chimeric β-glucuronidase (GUS) gene expression in an efficientAgrobacterium-mediated transformation system utilising mesophyll cells ofPetunia hybrida synchronized with cell cycle phase-specific inhibitors (mimosine and colchicine) was used to show the absolute requirement of S-phase for transfer and/or integration of the transferred DNA (T-DNA). Flow-cytometric analysis of nuclear DNA content and immunohistological detection of bromodeoxyuridine (BrdUrd) incorporation showed that, prior to phytohormone treatment, most (98%) mesophyll cells were at GO-Gl-phase (quiescent phase) and no cell division was occurring. After 48 h and 72 h of phytohormone treatment, there was a rapid increase in S-G2-M-phase populations (〉 75%) and a concomitant decrease (down to 24%) in G0–-G1-phase cells. Assays of GUS showed that maximum transformation (〉 95% of explants) also occurred after this period. Our data showed that mimosine and colchicine blocked the mesophyll cells at late Gl-phase and M-phase, respectively. No transformation (= GUS expression) was observed in phytohormone-treated cells inhibited in late G1 by mimosine. However, after removal of mimosine, 82% of the explants were transformed, indicating the non-toxic and reversible effect of the inhibitor. On the other hand, a relatively high transformation frequency (65% of explants) was observed after blocking the cell cycle at M-phase with colchicine. However, only transient, but no stable, gene expression (= kanamycin-resistant callus formation) was observed in colchicine-treated M-phase-arrested cells. Similarly, endoreduplication of nuclear DNA, which occurred during the 48 h of phytohormone treatment in some mesophyll cells and cells located along the minor veins in the leaf explants, resulted in transient GUS expression only. These observations indicate a direct correlation between endoreduplication and transient GUS gene expression. Obviously, for stable GUS gene expression, cell division and proliferation are required, indicating that both DNA duplication (S-phase) and cell division (M-phase) are strongly related to stable transformation. We propose that the present system should facilitate further dissection of the process of T-DNA integration in the host genome and therefore should aid in developing new strategies for transformation of recalcitrant plants.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01007700
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  • 24
    Digitale Medien
    Digitale Medien
    Expression and inheritance of foreign genes in transgenic peanut plants generated byAgrobacterium-mediated transformation (1997)
    Springer
    Plant cell reports 16 (1997), S. 541-544 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Transgenic peanut ; Agrobacterium ; Transformation ; Transgene expression ; Transgene inheritance
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract To evaluate and characterize the stability of traits transferred viaAgrobacterium transformation, foreign gene expression must be examined in sexually derived progeny. The objective of this study was to analyze three transgenic peanut plants, 1-10, 12-1, and 17-1, for the inheritance and expression of their foreign genes. Segregation ratios for the introduced genes in T2 plants gave either 100% or 3:1 expression of the β-glucuronidase (GUS) gene, demonstrating recovery of both homozygous and heterozygous T1 plants. Fluorometric GUS assay in T1 and T2 generations of all three plants showed that the GUS gene was stably expressed in the progeny. DNA analyses showed 100% concordance between the presence of the foreign gene and enzyme activity. Our results demonstrate that transgenes in peanut introduced byAgrobacterium can be inherited in a Mendelian manner.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01142320
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  • 25
    Digitale Medien
    Digitale Medien
    Isolation of new 6-methylnicotinic-acid-degrading bacteria, one of which catalyses the regioselective hydroxylation of nicotinic acid at position C2 (1997)
    Springer
    Archives of microbiology 168 (1997), S. 355-361 
    Details
    ISSN: 1432-072X
    Schlagwort(e): Key words 6-Methylnicotinic acid ; 2-Hydroxy-6-methylnicotinic acid ; Nicotinic acid ; 2-Hydroxynicotinic acid ; Ralstonia ; Burkholderia ; Paenibacillus ; Agrobacterium ; Rhizobium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract 2-Hydroxynicotinic acid is an important building block for herbicides and pharmaceuticals. Enrichment strategies to increase the chances of finding microorganisms capable of hydroxylating at the C2 position and to avoid the degradation of nicotinic acid via the usual intermediate, 6-hydroxynicotinic acid, were used. Three bacterial strains (Mena 23/3–3c, Mena 25/4–1, and Mena 25/ 4–3) were isolated from enrichment cultures with 6-methylnicotinic acid as the sole source of carbon and energy. Partial characterization of these strains indicated that they represent new bacterial species. All three strains completely degraded 6-methylnicotinic acid, and evidence is presented that the first step in the degradation pathway of strain Mena 23/3–3c is hydroxylation at the C2 position. Resting cells of this strain grown on 6-methylnicotinic acid also hydroxylated nicotinic acid at the C2 position, but did not further degrade the product. Strain Mena 23/ 3–3c showed the highest degree of 16S rRNA sequence similarity to members of the genera Ralstonia and Burkholderia.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/s002030050509
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  • 26
    Digitale Medien
    Digitale Medien
    Excision of the maize transposable element Ac in periclinal chimeric leaves of 35S-Ac-rolC transgenic aspen-Populus (1997)
    Springer
    Plant molecular biology 33 (1997), S. 1097-1103 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Ac ; Agrobacterium ; periclinal chimera ; rolC ; transgenic Populus
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The transposable element Ac from maize, in combination with the phenotypic selectable marker rolC, was employed in transformation experiments of a hybrid aspen clone. A number of transgenic clones exhibited light-green sectors on green leaves. In vitro regeneration from leaves showing a high number of light-green spots resulted in R2 plants, which also showed light-green sectored leaves. However, only one out of 385 regenerated plants obtained showed green leaves. Both PCR and northern analysis indicated Ac excision and restoration of rolC expression. In Southern blot analysis of this green plant additional bands were observed as compared to the original R1 plant. The occurrence of these bands and a suggested Ac excision in the non-green L1-epidermal layer leading to periclinal chimerism of this plant is discussed.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1005788706864
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  • 27
    Digitale Medien
    Digitale Medien
    Tapetum-specific expression of theOsg6B promoter-β-glucuronidase gene in transgenic rice (1997)
    Springer
    Plant cell reports 16 (1997), S. 363-367 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Agrobacterium ; Tapetum-specific promoter ; Transformation ; Rice
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The promoter of an anther tapetum-specific gene,Osg6B, was fused to aβ-glucuronidase (GUS) gene and introduced into rice byAgrobacterium-mediated gene transfer. Fluorometric and histochemical GUS assay showed that GUS was expressed exclusively within the tapetum of anthers from the uninucleate microspore stage (7 days before anthesis) to the tricellular pollen stage (3 days before anthesis). This is the first demonstration of an anther-specific promoter directing tapetum-specific expression in rice.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01146774
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  • 28
    Digitale Medien
    Digitale Medien
    Expression and inheritance of foreign genes in transgenic peanut plants generated by Agrobacterium-mediated transformation (1997)
    Springer
    Plant cell reports 16 (1997), S. 541-544 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Key words Transgenic peanut ; Agrobacterium ; Transformation ; Transgene expression ; Transgene inheritance
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract To evaluate and characterize the stability of traits transferred via Agrobacterium transformation, foreign gene expression must be examined in sexually derived progeny. The objective of this study was to analyze three transgenic peanut plants, 1-10, 12-1, and 17-1, for the inheritance and expression of their foreign genes. Segregation ratios for the introduced genes in T2 plants gave either 100% or 3:1 expression of the β-glucuronidase (GUS) gene, demonstrating recovery of both homozygous and heterozygous T1 plants. Fluorometric GUS assay in T1 and T2 generations of all three plants showed that the GUS gene was stably expressed in the progeny. DNA analyses showed 100% concordance between the presence of the foreign gene and enzyme activity. Our results demonstrate that transgenes in peanut introduced by Agrobacterium can be inherited in a Mendelian manner.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01142320
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  • 29
    Digitale Medien
    Digitale Medien
    Genetic transformation of Populus genotypes with different chimaeric gene constructs: transformation efficiency and molecular analysis (1997)
    Springer
    Transgenic research 6 (1997), S. 111-121 
    Details
    ISSN: 1573-9368
    Schlagwort(e): Ac ; Agrobacterium ; aspen ; Populus ; transformation ; transposition
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Aspen (Populus tremula) and hybrid aspen (P. tremula × P. tremuloides) were transformed with different gene constructs using two types of promoter. The aim was to determine the influence of the reporter gene rolC, controlled by promoters of viral or plant origin, on genetic and morphologic expression of different transgenic aspen clones. An improved transformation method using leaf discs was developed, by which putative transgenic plantlets were regenerated at high efficiencies (up to 34%) on kanamycin-containing medium. Transgenic aspen carrying the rolC gene from Agrobacterium rhizogenes under control of the cauliflower-35S-promoter are reduced in size with smaller leaves, whereas aspen transgenic for the same rolC gene, but under control of the light inducible rbcS promoter from potato, are only slightly reduced in size compared to untransformed controls. However, all clones carrying 35S-rolC and rbcS-rolC genes revealed light-green colouration of leaves when compared to untransformed aspen. Owing to this special feature, constructs were used in which expression of the rolC gene was inhibited by insertion of a transposable element, Ac, from maize. Transgenic aspen transformed with the 35S-Ac-rolC and rbcS-Ac-rolC genes were morphologically similar to untransformed aspen, but out of 54 independently regenerated 35S-Ac-rolC transgenic aspen clones, 30 clones showed light-green/dark green variegated leaves. In contrast, out of 19 independently transformed rbcS-Ac-rolC aspen clones, only two clones revealed light-green/dark green variegated leaves. The role of bacterial strains in transformation, and molecular genetics of transgenic aspen plants (including the function of the transposable element, Ac, in the aspen genome) are discussed
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1018421620040
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  • 30
    Digitale Medien
    Digitale Medien
    Agrobacterium rhizogenes and A. tumefaciens co-transformation to obtain grapevine hairy roots producing the coat protein of grapevine chrome mosaic nepovirus (1997)
    Springer
    Plant cell, tissue and organ culture 49 (1997), S. 53-62 
    Details
    ISSN: 1573-5044
    Schlagwort(e): Agrobacterium ; coat protein ; grapevine ; hairy root ; nepovirus ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Hairy root cultures of grapevine were obtained from plantlets co-inoculated by virulent Agrobacterium rhizogenes strains and disarmed A. tumefaciens strains harbouring the binary vectors pKHG4 and pKVHG 2+. These plasmids contain the nptII, hpt and gus genes and differ for the presence of the gene encoding for the grapevine chrome mosaic virus coat protein. For the cultivar ‘Gravesac’, 72% of the excised root tips initiated hairy root cultures on growth regulator-free media. According to the nature of the strains used in co-inoculation, co-transformation frequencies of the hairy root clones ranged from 4 to 16%. Co-transformed roots showed resistance to kanamycin and hygromycin but responses varied from clone to clone. Fluorometric GUS expression and GCMV coat protein production showed a large variability among hairy root clones co-transformed by pKHVG2+. Though the presence of gus, nptII and GCMV coat protein genes was checked by polymerase chain reaction and Southern blotting, it was difficult to establish a clear relationship between expression of the different transgenes. The regeneration of plants was not achieved, but the possibility to graft in vitro transgenic roots to non transformed shoot systems could permit rapid testing of the resistance induced by nepovirus coat protein in roots of cultivars that are recalcitrant to A. tumefaciens-mediated transformation.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1005854212592
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  • 31
    Digitale Medien
    Digitale Medien
    Isolation and identification of N2-fixing microorganisms from the rhizosphere of Capparis spinosa (L.) (1997)
    Springer
    Plant and soil 197 (1997), S. 19-23 
    Details
    ISSN: 1573-5036
    Schlagwort(e): Agrobacterium ; Capparis spinosa ; Comamonas ; N2 fixation ; Pseudomonas ; rhizosphere ; Sphingobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Four bacterial strains, Pseudomonas stutzeri var. mendocina, Comamonas sp., Agrobacterium tumefaciens biovar. 2 and Sphingobacterium sp., isolated from the rhizosphere of wild-grown caper (Capparis spinosa L.) plants were able to fix N2 as shown by their growth in nitrogen-free medium and by the acetylene reduction test. P. stutzeri var. mendocina and Comamonas sp. contained DNA homologous to the Klebsiella pneumoniae M5a1 nifHDK genes. No hybridization was found with total DNA from either A. tumefaciens biovar. 2 or Sphingobacterium sp. using nifHDK probes from either K. pneumoniae or Rhizobium meliloti.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1004211909641
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  • 32
    Digitale Medien
    Digitale Medien
    SAAT: sonication-assisted Agrobacterium-mediated transformation (1997)
    Springer
    Transgenic research 6 (1997), S. 329-336 
    Details
    ISSN: 1573-9368
    Schlagwort(e): Agrobacterium ; SAAT ; sonication ; transformation ; wounding
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Plant transformation via Agrobacterium can be limited by both host specificity and the inability of Agrobacterium to reach the proper cells in the target tissue. Described here is a new and efficient Agrobacterium-based transformation technology that overcomes these barriers and enhances DNA transfer in such diverse plant groups as dicots, monocots, and gymnosperms. This new technology, called sonication-assisted Agrobacterium-mediated transformation (SAAT), involves subjecting the plant tissue to brief periods of ultrasound in the presence of Agrobacterium. Scanning electron and light microscopy reveal that SAAT treatment produces small and uniform fissures and channels throughout the tissue allowing the Agrobacterium easy access to internal plant tissues. Unlike other transformation methods, this system has the potential to transform meristematic tissue buried under several cell layers. SAAT increases transient transformation efficiency in several different plant tissues including leaf tissue, immature cotyledons, somatic and zygotic embryos, roots, stems, shoot apices, embryogenic suspension cells and whole seedlings. A 100- to 1400-fold increase in transient β- glucuronid ase expression has been demonstrated in various tissues of soybean, Ohio buckeye, cowpea, white spruce, wheat and maize. Stable transformation of both soybean and Ohio buckeye has been obtained using SAAT of embryogenic suspension culture tissues. For soybean, SAAT treatment was necessary to obtain stable transformation with this tissue
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1018470930944
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  • 33
    Digitale Medien
    Digitale Medien
    Matrix attachment regions (MARs) enhance transformation frequency and transgene expression in poplar (1997)
    Springer
    Transgenic research 6 (1997), S. 415-420 
    Details
    ISSN: 1573-9368
    Schlagwort(e): GUS ; matrix attachment regions ; Populus ; transformation ; transgene expression ; Agrobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract We tested the value of a matrix attachment region (MAR) fragment derived from a tobacco gene for increasing the frequency of Agrobacterium-mediated transformation. A binary vector that carried a GUS reporter gene containing an intron and an nptII gene was modified to contain flanking MAR elements within the T-DNA borders. Vectors containing or lacking MARs were then used to transform tobacco, a readily transformabl e poplar clone (Populus tremula × P. alba), and a recalcitrant poplar clone (Populus trichocarpa × P. deltoides). MARs increased GUS gene expression approximately 10-fold in the two hybrid poplar clones and twofold in tobacco one month after cocultivation with Agrobacterium; MARs also increased the frequency of kanamycin-resistant poplar shoots recovered
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1023/A:1018439518649
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  • 34
    Digitale Medien
    Digitale Medien
    Transgenic plant production mediated by Agrobacterium in Indica rice (1996)
    Springer
    Plant cell reports 15 (1996), S. 727-730 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Acetosyringone ; Agrobacterium ; Indica rice ; Oryza sativa L. ; Transgenic plants
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Summary A reproducible system has been developed for the production of transgenic plants in indica rice using Agrobacterium-mediated gene transfer. Three-week-old scutella calli served as an excellent starting material. These were infected with an Agrobacterium tumefaciens strain EHA101 carrying a plasmid pIG121Hm containing genes for β-glucuronidase (GUS) and hygromycin resistnace (HygR). Hygromycin (50 mg/l) was used as a selectable agent. Inclusion of acetosyringone (50μM) in the Agrobacterium suspension and co-culture media proved to be indispensable for successful transformation. Transformation efficiency of Basmati 370 was 22% which was as high as reported in japonica rice and dicots. A large number of morphologically normal, fertile transgenic plants were obtained. Integration of foreign genes into the genome of transgenic plants was confirmed by Southern blot analysis. GUS and HygR genes were inherited and expressed in R1 progeny. Mendelian segregation was observed in some R1 progeny.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00232216
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  • 35
    Digitale Medien
    Digitale Medien
    Transient expression of uidA constructs in in vitro onion (Allium cepa L.) cultures following particle bombardment and Agrobacterium-mediated DNA delivery (1996)
    Springer
    Plant cell reports 15 (1996), S. 958-962 
    Details
    ISSN: 1432-203X
    Schlagwort(e): Transformation ; particle bombardment ; Agrobacterium ; Allium cepa.
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Particle bombardment and Agrobacterium-mediated DNA delivery into immature embryos and microbulbs were used to investigate the expression of the uidA gene in in vitro onion cultures. Both methods were successful in delivering DNA and subsequent uidA expression was observed. Optimal transient β-glucuronidase activity was observed in immature embryos that had been pre-cultured for three days and bombarded at a distance of 3 cm from the stopping plate, under 25 in Hg vacuum, using 900–1300 psi rupture discs. The CaMV35S-uidA gene construct gave five fold higher transient β-glucuronidase activity than the uidA gene construct regulated by any of four other promoters initially chosen for high experession in monocotyledonous tissues.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00231596
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  • 36
    Digitale Medien
    Digitale Medien
    Production of an extracellular polysaccharide byAgrobacterium sp DS3 NRRL B-14297 isolated from soil (1996)
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 129-133 
    Details
    ISSN: 1476-5535
    Schlagwort(e): extracellular polysaccharide ; Agrobacterium ; viscous polysaccharide
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie , Werkstoffwissenschaften, Fertigungsverfahren, Fertigung
    Notizen: Abstract A bacterium isolated from soil and identified asAgrobacterium sp produced a water-soluble extracellular polysaccharide capable of producing highly viscous solutions. Gas chromatographic analysis revealed a sugar composition of glucose, galactose and mannose in the molar ratio of 7.5∶2.4∶1, together with 3.7% (w/w) pyruvic acid. Methylation analyses showed the presence of (1→3)-, (1→4)- and (1→6)-linked glucose, (1→3)- and (1→4, 1→6)-linked galactose and a small portion of (1→3)-linked mannose residues. Succinic acid was not present. The molecular weight of the polysaccharide was estimated by light scattering to be 2×106 Da. The viscosity of solutions containing the polysaccharide remained constant from pH 3 to 11, and decreased by 50% when heated from 5 to 55°C. Maximum yield of the polysaccharide, 20 g L−1, was reached in 48 h at 30°C incubation.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01570073
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  • 37
    Digitale Medien
    Digitale Medien
    Agrobacterium tumefaciens-mediated transformation of japonica and indica rice varieties (1996)
    Springer
    Planta 199 (1996), S. 612-617 
    Details
    ISSN: 1432-2048
    Schlagwort(e): Agrobacterium ; indica rice ; Inheritance ; Japonica rice ; Oryza ; Transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Genetic transformation of rice (Oryza sativa L.) mediated by Agrobacterium ttumefaciens has been confirmed for japonica varieties and extended to include the more recalcitrant indica varieties. Immature embryos were inoculated with either A. tumefaciens At656 (pCNL56) or LBA4404 (pTOK233). Experimental conditions were developed initially for immature embryos treated with strain At656, based upon both transient and stable β-glucuromdase (GUS) activities. However, plant regeneration following selection on G418 (pCNL56 contained the nptII gene) did not occur. Using the same basic protocol, but inoculating immature embryos of rice with LBA4404 (pTOK233), resulted in efficient (about 27%) production of transgenic plants of the japonica variety, Radon, and an acceptable efficiency (from 1–5%) for the indica varieties IR72 and TCS10. Transformation was based upon resistance to hygromycin (pTOK233 contains the hpt gene), the presence of GUS activity (from the gusA gene), Southern blots for detection of the integrated gusA gene, and transmission of GUS activity to progeny in a Mendelian 3:1 segregation ratio. Southern blots indicated two to three copies of the gene integrated in most transformants. Transgenic plants of both the japonica and indica varieties were self-fertile and comparable in this respect to seed-grown plants. Key factors facilitating the transformation of rice by Agrobacterium tumefaciens appeared to be the use of embryos as the expiant, the use of hygromycin as the selection agent (which does not interfere with rice regeneration), the presence of extra copies of certain vir genes on the binary vector of pTOK233, and maintaining high concentrations of acetosyringone for inducing the vir genes during co-cultivation of embryos with Agrobacterium.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00195194
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  • 38
    Digitale Medien
    Digitale Medien
    Sequence and characterisation of a ribosomal RNA operon from Agrobacterium vitis (1996)
    Springer
    Molecular genetics and genomics 251 (1996), S. 99-107 
    Details
    ISSN: 1617-4623
    Schlagwort(e): Key words Ribosomal genes ; Agrobacterium ; Evolution ; 23S rRNA
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract  One of the four ribosomal RNA operons (rrnA) from the Agrobacterium vitis vitopine strain S4 was sequenced. rrnA is most closely related to the rrn operons of Bradyrhizobium japonicum and Rhodobacter sphaeroides and carries an fMet-tRNA gene downstream of its 5S gene, as in the case of R. sphaeroides. The 16S rRNA sequence of S4 differs from the A. vitis K309 type strain sequence by only one nucleotide, in spite of the fact that S4 and K309 have very different Ti plasmids. The predicted secondary structure of the S4 23S rRNA shows several features that are specific for the alpha proteobacteria, and an unusual branched structure in the universal B8 stem. The 3′ ends of the three other rrn copies of S4 were also cloned and sequenced. Sequence comparison delimits the 3′ ends of the four repeats and defines two groups: rrnA/rrnB and rrnC/rrnD.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF02174350
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  • 39
    Digitale Medien
    Digitale Medien
    Transformation ofBrassica oleracea L.: a critical review (1996)
    Springer
    Molecular breeding 2 (1996), S. 185-210 
    Details
    ISSN: 1572-9788
    Schlagwort(e): Brassica oleracea ; Agrobacterium ; transformation ; direct gene transfer ; regeneration ; virulence ; flowering ; rol genes ; transgene expression ; transgene inactivation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Brassica oleracea is a highly polymorphic species encompassing a wide range of important vegetable and fodder crops. Gene transfer into cultivated forms of this species requires reproducible and efficient methods for genetic transformation and plant regeneration. In this review, we have collated the research experience on transformation ofB. oleracea to highlight the problems encountered. Most research effort has been directed at developingAgrobacterium-mediated transformation methods with relatively little emphasis to date on direct gene transfer techniques. Common procedures for the transformation ofB. oleracea have not emerged, due to the inherent variability between and amongst genotypes. Future progress would be facilitated by the use of genetically fixed material, such as double-haploid or inbred lines, to reduce variation of response within genotypes and would avoid the need for cultivar-specific transformation protocols if responsive lines amenable to crossing with cultivated forms could be identified. The principal difficulties relate to combining efficient plant regeneration with gene transfer. Methods that enhance bacterial virulence and increase the proportion of cells susceptible to transformation and competent for regeneration are discussed. Inefficient selection is a major cause of poor transformation frequencies inB. oleracea and has resulted in the regeneration of chimeric plants uponAgrobacterium tumefaciens-mediated transformation. Promising results have been obtained withAgrobacterium rhizogenes-mediated transformation but the impact of therol genes on flowering of primary transformants has not yet been fully assessed. Strategies to reduce the deleterious effects of therol genes on flowering are discussed. Few agronomically useful characters have been introduced, the majority of research having been confined to the introduction of marker and reporter genes; possible candidate genes are discussed.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00564197
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  • 40
    Digitale Medien
    Digitale Medien
    Sequence and characterisation of a ribosomal RNA operon fromAgrobacterium vitis (1996)
    Springer
    Molecular genetics and genomics 251 (1996), S. 99-107 
    Details
    ISSN: 1617-4623
    Schlagwort(e): Ribosomal genes ; Agrobacterium ; Evolution ; 23S rRNA
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract One of the four ribosomal RNA operons (rrnA) from theAgrobacterium vitis vitopine strain S4 was sequenced.rrnA is most closely related to therrn operons ofBradyrhizobium japonicum andRhodobacter sphaeroides and carries an fMet-tRNA gene downstream of its 5S gene, as in the case ofR. sphaeroides. The 16S rRNA sequence of S4 differs from theA. vitis K309 type strain sequence by only one nucleotide, in spite of the fact that S4 and K309 have very different Ti plasmids. The predicted secondary structure of the S4 23S rRNA shows several features that are specific for the alpha proteobacteria, and an unusual branched structure in the universal B8 stem. The 3′ ends of the three otherrrn copies of S4 were also cloned and sequenced. Sequence comparison delimits the 3′ ends of the four repeats and defines two groups:rrnA/rrnB andrrnC/rrnD.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF02174350
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  • 41
    Digitale Medien
    Digitale Medien
    Corresponding 16S rRNA gene segments in Rhizobiaceae and Aeromonas yield discordant phylogenies (1996)
    Springer
    Plant and soil 186 (1996), S. 69-74 
    Details
    ISSN: 1573-5036
    Schlagwort(e): Agrobacterium ; population genetics ; Rhizobium ; systematics ; taxonomy
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Previous evidence has indicated that the 16S rRNA genes in certain species of Aeromonas may have a history of lateral transfer and recombination. A comparative analysis of patterns of 16S nucleotide sequence polymorphism among species of Rhizobium and Agrobacterium was conducted to determine if there is similar evidence for chimeric 16S genes in members of the Rhizobiaceae. Results from phylogenetic analyses and comparison of patterns of nucleotide sequence polymorphism in portions of rhizobial 16S genes revealed the same type of segment-dependent polymorphic site partitioning that was previously reported for Aeromonas. These results support the hypothesis that certain 16S segments in rhizobia may have a history of lateral transfer and recombination.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00035057
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  • 42
    Digitale Medien
    Digitale Medien
    Identification of Agrobacterium and Rhizobium species based on cellular fatty acid composition (1996)
    Springer
    Plant and soil 184 (1996), S. 143-158 
    Details
    ISSN: 1573-5036
    Schlagwort(e): Agrobacterium ; FAME ; fatty acid analysis ; rapid identification ; Rhizobium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract The increasing number of phylogenetically defined species in the genera Agrobacterium, Rhizobium and Sinorhizobium suggests a need for a rapid identification method which will distinguish between these species. We have examined 65 strains of Agrobacterium representing: A. tumefacies, (34); A. rhizogenes, (16) A. vitis, (10) A. rubi (2) and some unclassified strains, and 150 strains of Rhizobium and Sinorhizobium representing: R. etli (21); R. galegae (20); R. huakuii (17); R. leguminosarum (20); R. loti (16); R. topici (18); S. fredii (19); and S. meliloti (20). Fatty acid methyl esters (FAME) were obtained from each strain, as previously described, and analysed by gas-chromatography using the MIDI Hewlett-Packard Microbial Identification System (MIS). Fatty acid profiles were recorded, characteristic fatty acids noted and the overall similarities between fatty acid profiles for each species calculated. Relationships between species were also derived from the fatty acid data by principal component analysis. This showed overlapping clusters for strains of R. leguminosarum and R. etli, R. topici and A. rhizogenes and S. fredii and S. meliloti within one supercluster. Strains of A. tumefaciens, A rubi, A. vitis and R. galegae formed a second supercluster while R. loti and R. huakuii strains formed a third cluster well separated from all the other strains. The fatty acid profiles were used to correctly identify at least 94% of the strains representing each species in the collection except R. etli. R. etli strains (23.8%) were misidentified as R. leguminosarum. This was attributed to the high similarity (44.7%) between R. etli and R. leguminosarum. It is concluded that whole cell fatty acid analysis should form part of the polyphasic description of new species of root nodule bacteria, with the proviso that growth conditions and analytical methods be carefully standardized. It is suggested that FAME-MIS system and the database we have compiled provide a basis for future development.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00029284
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  • 43
    Digitale Medien
    Digitale Medien
    Agrobacterium-mediated transformation of Javanica rice (1996)
    Springer
    Molecular breeding 2 (1996), S. 267-276 
    Details
    ISSN: 1572-9788
    Schlagwort(e): Agrobacterium ; Javanica rice ; regeneration ; rice ; TAIL-PCR ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract Difficulties frequently encountered using direct DNA transfer methods for transformation of Javanica varieties of rice (Oryza sativa L.) have limited the application of biotechnology to these varieties. We now reportAgrobacterium-mediated transformation of Javanica cultivars Gulfmont and Jefferson that are, respectively, widely used or about to enter commercial cultivation in the southern USA. Vigorous, phenotypically normal, fertile plants expressing both the selectable marker and the gene of interest were obtained. Southern analysis showed that only one or two copies of the T-DNA insert were present. Sequence analysis of right border fragments of one line confirmed that insertion was into a coding region of rice nuclear DNA. This analysis also revealed the presence of relatively short regions of permuted T-DNA border sequences, similar to those found afterAgrobacterium-mediated transformation of dicots. Progeny analysis of lines bearing two copies showed co-segregation, indicating that they were located relatively closely on the same chromosome. The introduced genes were transmitted to the R1 and R2 generations in a Mendelian fashion, confirming the suitability of this approach for biotechnological improvement of elite rice cultivars.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00564204
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  • 44
    Digitale Medien
    Digitale Medien
    5′-Regulatory region of Agrobacterium tumefaciens T-DNA gene 6b directs organ-specific, wound-inducible and auxin-inducible expression in transgenic tobacco (1995)
    Springer
    Plant molecular biology 29 (1995), S. 1299-1304 
    Details
    ISSN: 1573-5028
    Schlagwort(e): Agrobacterium ; gene 6b ; phytohormonal regulation of expression ; T-DNA promoter
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The regulatory activity of a 826 bp DNA fragment located upstream of the pTiBo542 TL-DNA gene 6b coding region was analysed in transgenic tobacco, using β-glucuronidase (gus) as a reporter gene. The region was shown to drive organ-specific, wound- and auxin-inducible expression of the reporter, the effect being dependent on the type and concentration of auxin.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00020470
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  • 45
    Digitale Medien
    Digitale Medien
    The three-dimensional structure of vascular tissues in Agrobacterium tumefaciens-induced crown galls and in the host stems of Ricinus communis L. (1995)
    Springer
    Planta 196 (1995), S. 597-605 
    Details
    ISSN: 1432-2048
    Schlagwort(e): Agrobacterium ; Crown gall ; Fiber ; Phloem anastomoses ; Phloem and xylem differentiation ; Phytohormones (auxin, cytokinin, ethylene, gibberellin) ; Ray differentiation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The three-dimensional pattern of phloem and xylem in 10-d-to two-month-old tumors induced by Agrobacterium tumefaciens (C58) and in adjacent Ricinus communis L. stem tissues was studied in thick sections by clearing with lactic acid and by staining with lacmoid. The crown galls contained two types of vascular strands: treelike branched bundles, which developed towards the tumor surface in fast-growing regions, and globular bundles in the slowly developing parts. Both types of vascular bundles contained xylem and phloem and were continuous with the vascular system of the host plant. The tumor bundles were interconnected by a dense net of phloem anastomoses, consisting of sieve tubes but no vessels. These vascular patterns reflect the apparent synthesis sites, concentration gradients and flow pathways of the plant hormones additionally produced in the tumors upon expression of the T-DNA-encoded genes. The A. tumefaciens-induced crown gall affected vascular differentiation in the host stem. In the basipetal direction, the tumor induced more xylem differentiation directly below it, where the crown-gall bundles joined the vascular system of the host. In the centripetal direction, the crown gall caused the development of pathologic xylem characterized by narrow vessels, giant rays and absence of fibers. On the other hand, most probably as a consequence of its gibberellic acid content, the host plant stimulated a local differentiation of regenerative phloem and xylem fibers with unique ramifications, only at the base of the tumor. However, fibers were absent from the main body of the crown gall. The study shows that A. tumefaciens-induced crown galls are characterized by a sophisticated network of vascular tissues in the tumor and are accompanied by a perturbated vessel system in the host. The hormonal mechanisms controlling vascular differentiation in the tumor and neighboring host tissues are discussed. In addition, the “gall constriction hypothesis” is proposed for explaining the mechanism which gives priority in water supply to the growing gall over the host shoot.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00203661
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  • 46
    Digitale Medien
    Digitale Medien
    The effect of auxin on cytokinin levels and metabolism in transgenic tobacco tissue expressing an ipt gene (1995)
    Springer
    Planta 196 (1995), S. 84-94 
    Details
    ISSN: 1432-2048
    Schlagwort(e): Agrobacterium ; Auxin ; Cytokinin metabolism ; Cytokinin oxidase ; ipt gene ; Nicotiana (cytokinin)
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The ipt gene from the T-DNA of Agrobacterium tumefaciens was transferred to tobacco (Nicotiana tabacum L.) in order to study the control which auxin appears to exert over levels of cytokinin generated by expression of this gene. The transgenic tissues contained elevated levels of cytokinins, exhibited cytokinin and auxin autonomy and grew as shooty calli on hormone-free media. Addition of 1-naphthylacetic acid to this culture medium reduced the total level of cytokinins by 84% while 6-benzylaminopurine elevated the cytokinin level when added to media containing auxin. The cytokinins in the transgenic tissue were labelled with 3H and auxin was found to promote conversion of zeatin-type cytokinins to 3H-labelled adenine derivatives. When the very rapid metabolism of exogenous [3H]zeatin riboside was suppressed by a phenylurea derivative, a noncompetitive inhibitor of cytokinin oxidase, auxin promoted metabolism to adenine-type compounds. Since these results indicated that auxin promoted cytokinin oxidase activity in the transformed tissue, this enzyme was purified from the tobacco tissue cultures. Auxin did not increase the level of the enzyme per unit tissue protein, but did enhance the activity of the enzyme in vitro and promoted the activity of both glycosylated and non-glycosylated forms. This enhancement could contribute to the decrease in cytokinin level induced by auxin. Studies of cytokinin biosynthesis in the transgenic tissues indicated that trans-hydroxylation of isopentenyladenine-type cytokinins to yield zeatin-type cytokinins occurred principally at the nucleotide level.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00193221
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  • 47
    Digitale Medien
    Digitale Medien
    Variability in the response toPseudomonas solanacearum of transgenic lines of potato carrying a cecropin gene analogue (1995)
    Springer
    Potato research 38 (1995), S. 371-378 
    Details
    ISSN: 1871-4528
    Schlagwort(e): bacterial wilt ; resistance ; transformation ; Agrobacterium ; S. tuberosum L. ; transgenic plant
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Transgenic potato plants of cv. Désirée carrying an antibacterial gene, coding for a cecropin lytic peptide analogue, were inoculated with a virulent strain ofPseudomonas solanacearum under controlled conditions. The disease index scored during three repeated infection trials indicated an increased variability in plant response among the transgenic lines which gave either a more susceptible or a more resistant response to the pathogen when compared with untransformed Désirée. Immunity toP. solanacearum was not observed, but it was possible to select a group of transgenic lines that showed resistance levels and disease development curves comparable to the field resistant cv. Cruza 148.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF02357742
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  • 48
    Digitale Medien
    Digitale Medien
    Rapid micropreps and minipreps of Ti plasmids and binary vectors fromAgrobacterium tumefaciens (1995)
    Springer
    Transgenic research 4 (1995), S. 349-351 
    Details
    ISSN: 1573-9368
    Schlagwort(e): Agrobacterium ; Ti plasmid ; binary vector ; minipreparation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract A simple and versatile procedure has been developed for the isolation of both large helper/Ti plasmids and binary vectors fromAgrobacterium tumefaciens. Using a slightly modified alkaline lysis protocol, intact plasmid can be recovered from cultures grown in standard micro-centrifuge tubes or culture tubes in sufficient yield and purity to allow for restriction analysis on ethidium bromide stained gels of the 〉200 kb Ti plasmid DNA. Contamination by chromosomal DNA is minimal and there is thus no need for isopycnic gradient purification. This same procedure can be combined with a high temperature treatment (37°C) and antibiotic selection to generate preparations containing binary vector DNA that are virtually free of interfering Ti plasmid DNA. Restriction patterns produced from these binary vector DNA preparations are unambiguous and therefore preliminary screening by Southern hybridization can be eliminated.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01972532
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  • 49
    Digitale Medien
    Digitale Medien
    Cell competence forAgrobacterium-mediated DNA transfer inPisum sativum L. (1995)
    Springer
    Transgenic research 4 (1995), S. 184-191 
    Details
    ISSN: 1573-9368
    Schlagwort(e): competence ; transient ; transformation ; Agrobacterium ; grain legume ; histochemical GUS-detection
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract Distribution and properties of pea (Pisum sativum L.) cells, competent forAgrobacterium-mediated transformation were analysed byin situ histochemical detection of GUS (β-glucuronidase) activity, 4 d after inoculation with engineeredAgrobacterium tumefaciens. The vector system consisted of the hypervirulent disarmed strain EHA101 and the binary plasmid pIBGUS, carrying an intron-containing, 35S-promotor drivengusA (oruidA) gene and two selectable marker genes. Cells competent for transformation were mainly restricted to the dedifferentiating cells neighbouring the vascular system of cotyledon and epicotyl explants. A standardized assay was developed, allowing determination and quantification of factors influencing number and distribution of competent cells. In etiolated seedlings, competence for transformation decreased with the distance of the epicotyl explant from the shoot apex and was specifically induced by the exogenous application of auxins. Transient expression ofgusA afterAgrobacterium-mediated DNA transfer was dramatically reduced upon application of cell-cycle and DNA replication inhibitors aphidicolin, colchicine and nalidixic acid. GUS expression after direct DNA transfer of double-stranded plasmid DNA (via PEG into protoplasts or via particle bombardment of epicotyl segments) was independent of cell-division/DNA replication. A GUS-positive mutant of EHA101 was constructed to allowin situ analysis of attaching bacteria within the plant tissue. Attachment and invasion was inhibited by well-developed cuticula but was restored after chloroform treatment of the tissue surface. Moreover, no correlation was found between distribution of attaching bacteria and the pattern of transformation-competent cells.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01968783
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  • 50
    Digitale Medien
    Digitale Medien
    Ti plasmid-encoded octopine and nopaline catabolism in Agrobacterium: specificities of the LysR-type regulators OccR and NocR, and protein-induced DNA bending (1995)
    Springer
    Molecular genetics and genomics 249 (1995), S. 102-110 
    Details
    ISSN: 1617-4623
    Schlagwort(e): Agrobacterium ; Ti plasmid ; Activator ; Opines ; LysR family
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Biologie
    Notizen: Abstract The occ and noc regions in octopine and nopaline Ti plasmids, respectively, are responsible for the catabolism of octopine and nopaline in Agrobacterium. The functions are activated in the presence of the opines by OccR and NocR, two related regulatory proteins, and the promoters contain common sequence motifs. We have investigated heterologous interactions between the regulators and the promoters. Previous experiments using all possible heterologous combinations of opines, regulators, and promoters in vivo had demonstrated that only the combination of nopalme, NocR, and the occ promoter led to limited promoter activation. We now show that OccR and NocR bind to the heterologous promoters in vitro and in vivo. The weak or non-existent promoter activation actually observed could be explained by the assumption that OccR and NocR use different activation mechanisms; we investigated protein-induced DNA bending because of reports that the two regulators differ in this respect. Analysis with a bending vector showed that both OccR and NocR induced a DNA bend that is relaxed in the presence of the respective opine. The data suggest that subtle differences in regulator/promoter interactions are responsible for the inactivity of the heterologous combinations. Investigations with a chimeric NocR/OccR protein indicated that it induced a DNA bend in both promoters. No opine-induced relaxation was detectable with the hybrid, and the inducible promoter was not activated. These findings suggest that bend relaxation may be an integral part of promoter activation.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00290241
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  • 51
    Digitale Medien
    Digitale Medien
    New plant promoter and enhancer testing vectors (1995)
    Springer
    Molecular breeding 1 (1995), S. 419-423 
    Details
    ISSN: 1572-9788
    Schlagwort(e): binary vectors ; gus ; T-DNA ; Agrobacterium
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract We describe here a set of binary vectors suitable forAgrobacterium-mediated gene transfer and specially designed for studying plant promoters. These vectors are based on the use of thegus reporter gene, contain multiple unique restriction sites upstream of thegus gene, and minimal promoters for testing the effect of enhancers or activator elements. In addition, an intron-containinggus (uidA) gene was introduced into one of these vectors in order to examine reporter gene activity in tissues whereAgrobacterium contamination may be a problem or in transient expression assays.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF01248419
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  • 52
    Digitale Medien
    Digitale Medien
    Efficient transformation and regeneration of carnation cultivars usingAgrobacterium (1995)
    Springer
    Molecular breeding 1 (1995), S. 283-293 
    Details
    ISSN: 1572-9788
    Schlagwort(e): Agrobacterium ; carnation ; Dianthus caryophyllus ; regeneration ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Abstract We have developed an efficient method for transformation and regeneration of plants from carnation,Dianthus caryophyllus L. Whole leaves fromin vitro shoot cultures were mixed withAgrobacterium, cocultivated for 5 days and then plated on 2 µg/l chlorsulfuron (CS). Regenerated shoots and shoot clusters were divided into smaller sections and plated on 3 µg/l CS for selection to produce fully transformed shoots. Geneticin (G418) and kanamycin used were not as effective selective agents as CS. All regenerated shoots were vitrified. These were normalized, rooted and transferred to the greenhouse. 100% of regenerated plants were transformed based on rooting assay, GUS assay, PCR and Southern analysis.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF02277428
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  • 53
    Digitale Medien
    Digitale Medien
    Transformation of lily by Agrobacterium (1955)
    Springer
    Euphytica 85 (1955), S. 97-100 
    Details
    ISSN: 1573-5060
    Schlagwort(e): Agrobacterium ; transformation ; lily ; β-glucuronidase
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Lily cv. Harmony was inoculated with several Agrobacterium strains to study its susceptibility to Agrobacterium infection and transformation. Tumorous tissue formation on inoculated stem internodes of sterile-grown plantlets, as well as expression of a β-glucuronidase marker gene interrupted by an intron in cells of inoculated stem nodes, indicate that the monocotyledon Lilium is a host for Agrobacterium.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023935
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  • 54
    Digitale Medien
    Digitale Medien
    A study of different (CaMV 35S and mas) promoter activities and risk assessment of field use in transgenic rapeseed plants (1955)
    Springer
    Euphytica 85 (1955), S. 411-416 
    Details
    ISSN: 1573-5060
    Schlagwort(e): Agrobacterium ; Brassica napus ; CaMV 35S promoter ; mas promoter ; gene expression ; risk assessment
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Gene fusions between the β-glucuronidase (GUS) reporter gene and the promoters of the cauliflower mosaic virus 35S RNA transcript (CaMV 35S) and the mannopine synthase (mas) genes were introduced into rapeseed varieties via Agrobacterium-mediated transformation. Fluorometric assay of β-glucuronidase activity indicated different expression patterns for the two promoters. In seedlings, the CaMV 35S promoter had maximum activity in the primary roots, while the mas promoter was most active in the cotyledons. Etiolated seedlings cultured in the dark showed reduced activity of the mas promoter. Before vernalization at the rosette stage, both promoters were more active in older plant parts than in younger ones. At this stage the highest activity was recorded in cotyledons. After the plants had bolted reduced promoter function was detected in the upper parts of the transformed plants. Both promoters were found to be functional in the majority of the studied organs of transgenic rapeseed plants, but the promoter activity varied considerably between the organs at different developmental stages. The ability of pollen to transfer the introduced genes to other varieties and related species (e.g. Brassica napus and Diplotaxus muralis) by cross-pollination was studied in greenhouse experiments, and field trials were carried out to estimate the distance for biologically-relevant gene dispersal. In artificial crossing, the introduced marker gene was transferable into other varieties of Brassica napus. In field trials, at a distance of 1 metre from the source of transgenic plants, the frequency of an outcrossing event was relatively high (10-3). Resistant individuals were found at 16 and 32 metres from the transgenic pollen donors, but the frequency of an outcrossing event dropped to 10-5.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023974
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  • 55
    Digitale Medien
    Digitale Medien
    An assessment of morphogenic and transformation efficiency in a range of varieties of potato (Solanum tuberosum L.) (1955)
    Springer
    Euphytica 85 (1955), S. 101-108 
    Details
    ISSN: 1573-5060
    Schlagwort(e): Agrobacterium ; plant regeneration ; potato ; Solanum tuberosum ; tissue culture ; transformation
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary To provide a truly genotype-independent transformation system, it is necessary to be able to transform a wide range of potato genotypes. The ability to regenerate shoots in vitro was determined for 34 potato varieties using tuber disc explants. Following a culture regime used extensively in previous studies with the variety Desiree, half of the varieties could be regenerated from tuber discs and half could not. From a sample of varieties that could be regenerated from tuber discs, all but one variety gave transgenic plants. Twelve varieties were evaluated for the capacity to regenerate shoots from leaf and internode explants excised from in vitro grown plants. All of the varieties tested regenerated adventitious shoots. Leaf and internode explants from 5 varieties were subsequently used for transformation, and transgenic plants were produced from two potato varieties that did not give transgenic plants from tuber disc explants. Some varieties could not be transformed by either method, and will require modification of the in vitro regeneration and transformation system to be successful.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023936
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  • 56
    Digitale Medien
    Digitale Medien
    Some methodological aspects of apple transformation by Agrobacterium (1955)
    Springer
    Euphytica 85 (1955), S. 131-134 
    Details
    ISSN: 1573-5060
    Schlagwort(e): apple ; transformation ; Agrobacterium ; preculture ; azacytidine
    Quelle: Springer Online Journal Archives 1860-2000
    Thema: Land- und Forstwirtschaft, Gartenbau, Fischereiwirtschaft, Hauswirtschaft
    Notizen: Summary Leaf explants of apple cvs Gala and Golden Delicious were infected with the Agrobacterium tumefaciens strain AGL0(pMOG410). The effects of a 2 d preculture of the explants before infection and the addition of 5-azacytidine to the selection medium were studied. The percentages of GUS-positive explants after 5 w did not significantly alter due to these treatments. One of the ‘Gala’ shoots, which was removed from a leaf explant cultured for 8 w on selection medium, proved to be GUS-positive and will be analyzed further. In general, however, it should be concluded that regeneration of transgenic shoots directly from leaf tissue was not very effective.
    Materialart: Digitale Medien
    URL: http://dx.doi.org/10.1007/BF00023941
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