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  • 1
    Electronic Resource
    Electronic Resource
    Constitutive versus seed specific expression in transgenic wheat: temporal and spatial control (1999)
    Springer
    Transgenic research 8 (1999), S. 73-82 
    Details
    ISSN: 1573-9368
    Keywords: low molecular weight glutenin promoter ; particle bombardment ; transgene expression ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transgenic wheat plants from specific cultivars can now be routinely engineered in many laboratories. However, our understanding of the factors controlling transgene expression and stability in wheat compared to other cereals is rather limited. Only a few promoters have been tested in transgenic wheat, and relatively little is known of their relative activities and expression parameters. In the present study, the spatial and temporal properties of one heterologous constitutive promoter and one seed‐specific wheat promoter were investigated. We generated constructs with the reporter gene gusA (β‐glucuronidase) driven by: (a) the constitutive maize ubiquitin‐1 (ubi‐1) promoter, and (b) two different‐sized fragments of the seed‐specific low molecular weight glutenin (LMWG1D1) promoter from wheat. The activities of all three promoter constructs were comparable in endosperm tissue. A detailed analysis of spatial and temporal properties of the promoters is described. Heat shock treatment of transgenic plants carrying the ubi‐1: gusA construct resulted in a significant elevation in the levels of GUS activity. The inheritance of transgene expression levels and stability was evaluated over four generations, as a function of transgene integration patterns and copy number.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008801929494
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  • 2
    Electronic Resource
    Electronic Resource
    Multiple traits of agronomic importance in transgenic indica rice plants: analysis of transgene integration patterns, expression levels and stability (1999)
    Springer
    Molecular breeding 5 (1999), S. 471-480 
    Details
    ISSN: 1572-9788
    Keywords: Basmati 370 and M7 varieties ; δ-endotoxins (Cry1Ac and Cry2A) ; particle bombardment ; snowdrop lectin (GNA) ; transgenic rice
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract We cotransformed indica rice (Oryza sativa L. cvs. Basmati 370 and M7) with three plasmids, carrying a total of four genes, by particle bombardment. The Bacillus thuringiensis (Bt) δ-endotoxin genes cry1Ac and cry2A were carried on separate vectors, while the gna (snowdrop lectin) and hpt (hygromycin phosphotransferase) genes were linked on the same, cointegrate vector. We evaluated the molecular and expression profiles of 29 independently derived transgenic lines over two generations. The gna and hpt genes cointegrated with a frequency of 100% as expected. Furthermore, 60% of the transgenic plants carried all four genes. Southern blot analysis showed that transgene copy number ranged from 1 to 15. We used western blots to determine protein expression levels in R0 and R1 plants. We observed wide variation in the expression levels of the nonselected transgenes among independently-derived lines, but expression levels within lines derived from the same clone were similar. Consistent with previous reports, we observed no correlation between transgene copy number and the level or stability of protein expression. We show that the introduction of multiple agronomically valuable genes into the rice genome by cotransformation is a practical strategy for engineering elite rice varieties.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009634226797
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  • 3
    Electronic Resource
    Electronic Resource
    Characterization of the regulatory elements of the maize P-rr gene by transient expression assays (1999)
    Springer
    Plant molecular biology 39 (1999), S. 11-19 
    Details
    ISSN: 1573-5028
    Keywords: enhancer ; flavonoids ; particle bombardment ; pericarp ; promoter ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The maize P-rr gene conditions floral-specific flavonoid pigmentation, especially in the kernel pericarp and cob. We analyzed the P-rr promoter by transient expression assays, in which segments of the P-rr promoter were fused to the GUS reporter gene and introduced into maize cells by particle bombardment. A basal P-rr promoter fragment (−235 to +326) gave low, but significant, levels of GUS reporter gene expression. Interestingly, two widely spaced segments containing enhancer-like activity were found. When tested individually, both the proximal (−1252 to −236) and distal (−6110 to −4842) segments boosted expression of the basal P-rr promoter::GUS construct about five-fold. A 1.6 kb segment of the P-rr promoter (−1252 to +326) containing the proximal enhancer and the 5′-untranslated leader driving the GUS reporter gene showed preferential expression in BMS and embryogenic suspension cell cultures vs. endosperm-derived suspension cell cultures. These results demonstrate the application of transient assay techniques for the identification of regulatory elements responsible for floral-specific regulation of the complex P-rr gene promoter in maize.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006172815663
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  • 4
    Electronic Resource
    Electronic Resource
    Expression of fungal thermotolerant endo-1,4-β-glucanase in transgenic barley seeds during germination (1999)
    Springer
    Plant molecular biology 41 (1999), S. 777-783 
    Details
    ISSN: 1573-5028
    Keywords: cellulase ; heterologous expression ; Hordeum vulgare L. ; particle bombardment ; thermotolerant endo-1.4-β-glucanase ; transgenic barley
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The malting quality of two barley cultivars, Kymppi and Golden Promise, was modified to better meet the requirements of the brewing process. The egl1 gene, coding for fungal thermotolerant endo-1,4-β-glucanase (EGI, cellulase), was transferred to the cultivars using particle bombardment, and transgenic plants were regenerated on bialaphos selection. Integration of the egl1 gene was confirmed by Southern blot hybridization. The transgenic seeds were screened for the expression of the heterologous EGI. Under the high-pI α-amylase promoter, the egl1 gene was expressed during germination. The heterologous enzyme was thermotolerant at 65 °C for 2 h, thus being suitable for mashing conditions. The amount of heterologous EGI produced by the seeds (ca. 0.025% of soluble seed protein), has been shown to be sufficient to reduce wort viscosity by decreasing the soluble β-glucan content. A decrease in the soluble β-glucan content in the wort improves the filtration rate of beer.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006318206471
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  • 5
    Electronic Resource
    Electronic Resource
    Parameters influencing the regeneration capacity of calluses derived from mature indica and japonica rice seeds after microprojectile bombardment (1999)
    Springer
    Euphytica 107 (1999), S. 115-122 
    Details
    ISSN: 1573-5060
    Keywords: gus expressing cells ; histological analysis ; japonica and indica rice ; particle bombardment ; primary callus ; regeneration capacity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The microprojectile bombardment procedure has allowed the stable transformation of indica and japonica rice varieties, although at different frequencies of transformation depending mainly on their regeneration capacity and on the specific parameters of the transformation protocol. A study of the process of regeneration to whole plants from primary calli derived from mature indica and japonica rice seeds, via embryogenesis, has shown that somatic embryos are produced by division and differentiation of the external cell layers of callus tissues. Adjusting the bombardment conditions to optimize gene delivery to those regenerable cells, we have evaluated the influence of parameters such as the target distance, particle penetration and the effect of osmotic treatment on the regeneration capacity of bombarded cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026434228146
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  • 6
    Electronic Resource
    Electronic Resource
    Genetic transformation via particle bombardment of Catharanthus roseus plants through adventitious organogenesis of buds (1999)
    Springer
    Biotechnology letters 21 (1999), S. 997-1002 
    Details
    ISSN: 1573-6776
    Keywords: β-glucuronidase ; Catharanthus roseus ; genetic transformation ; green fluorescent protein ; particle bombardment
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In vitro propagation of Catharanthus roseus was achieved using nodal explants. Bud induction was best on medium containing 1.0 mg benzyl aminopurine l−1. Hardening of rooted shoots to soil was very successful with 98% survival. Genetically transformed C. roseus plantlets were obtained after bombardment of nodal explants, which were then micropropagated, with DNA coated particles with green fluorescent protein (GFP) or β-glucuronidase (GUS) reporter genes. Histological studies showed that the gene insertion method proved effective with many cells and different tissues displaying the reporter gene signals, showing that gene expressions were rather stable.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005622317333
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  • 7
    Electronic Resource
    Electronic Resource
    Green fluorescent protein (GFP) as a marker during pollen development (1999)
    Springer
    Transgenic research 8 (1999), S. 279-294 
    Details
    ISSN: 1573-9368
    Keywords: Antirrhinum ; Arabidopsis ; green fluorescent protein (GFP) ; in vitro ; microspores ; particle bombardment ; tobacco
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The transient expression of three mutant forms of green fluorescent protein (GFP) genes, GFP4, GFP5ER, and GFP4S65C, under several constitutive and pollen‐specific promoters throughout pollen development in Nicotianatabacum, thaliana and Antirrhinummajus is described. Immature pollen of tobacco, Arabidopsis and snapdragon, isolated at different developmental stages, were bombarded with plasmids containing the GFP and cultured in vitro for several days until maturity. The expression of GFP was monitored every day during in vitro maturation, germination and pollination, as well as after in situ pollination. The expression pattern of each construct was compared in parallel experiments to that of ß‐glucuronidase (GUS) constructs expressed by the same promoters. The results show that the expression level of all three GFP mutant forms was dependent on the strength of the promoter used. The strongest promoter was the DC3 promoter, and no notable differences in the intensity and brightness of all three versions of GFP were observed. GFP‐expressing pollen from tobacco and snapdragon developed in vitro for several days until maturity and germinated in vitro as well as on the surface of stigmata, strongly suggesting that all three GFPs are not toxic for the development of functional pollen. Furthermore, stably transformed tobacco plants expressing GFP under the control of the strong pollen‐expressed DC3 and LAT52 promoters were not impaired in reproductive function, confirming that GFP can be used as a non‐destructive marker for plant reproductive biology and development.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008938728051
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  • 8
    Electronic Resource
    Electronic Resource
    An Efficient Rice Transformation System Utilizing Mature Seed-derived Explants and a Portable, Inexpensive Particle Bombardment Device (1998)
    Springer
    Transgenic research 7 (1998), S. 289-294 
    Details
    ISSN: 1573-9368
    Keywords: Oryza sativa ; particle bombardment ; transformation ; transgenic rice
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We developed a practical and efficient gene transfer system for indica rice utilizing mature-seed derived explants and a simple bombardment device which uses compressed helium for accelerating DNA-coated metal particles. Unlike instruments which have been described in the literature previously, this new bombardment device, which is an improvement of the particle inflow concept, does not require vacuum. This attribute simplifies the transformation procedure significantly and it makes rice transformation technology accessible to laboratories which may not have the resources to invest in more expensive particle bombardment instruments. We determined experimentally that we could recover transgenic rice plants utilizing three different particle bombardment instruments at comparable frequencies.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008870012568
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  • 9
    Electronic Resource
    Electronic Resource
    Molecular Characteristics of Transgenic Wheat and the Effect on Transgene Expression (1998)
    Springer
    Transgenic research 7 (1998), S. 463-471 
    Details
    ISSN: 1573-9368
    Keywords: particle bombardment ; transgene expression ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A population of R0 transgenic wheat plants, generated by particle bombardment, was analyzed to define molecular, genetic and phenotypic properties resulting from transformation with a cointegrate vector, or cotransformation with two separate plasmids. By evaluating the progeny of 70 independently-derived transgenic plants, we also identified rare events such as chimerism and transgene elimination, which provide valuable information concerning the development of transgenic cereal plants following bombardment experiments. The frequency of chimerism in our transgenic wheat plants was very low. Furthermore, while transgene elimination did occur, this was also a very rare event. We determined the copy numbers of integrated transgenes and the levels of transgene expression. Comparisons to transgenic rice plants generated in the same manner demonstrated some similarities, but also important differences in transgene behavior. Whereas in rice there is no evidence for any direct relationship between transgene copy number and transgene expression or stability, multicopy populations in wheat demonstrated a bias towards higher levels of expression for the two genes and the maize ubiquitin promoter evaluated in the present study.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008833324193
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  • 10
    Electronic Resource
    Electronic Resource
    Molecular analysis of the genome of transgenic rice (Oryza sativa L.) plants produced via particle bombardment or intact cell electroporation (1998)
    Springer
    Molecular breeding 4 (1998), S. 99-109 
    Details
    ISSN: 1572-9788
    Keywords: transgenic rice ; particle bombardment ; cell electroporation ; RAPD ; AFLP ; AFRP ; RAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract In the present work we utilised some of the most discriminative molecular tools, such as RAPD, AFLP, AFRP and RAMP, to analyse the genome of independently derived transgenic plants from three elite Italian cultivars (cv. Lido, Carnaroli and Thaibonnet) and found that two methods for direct gene transfer, namely particle bombardment and intact cell electroporation (the latter being a procedure set up in this work), result in transgenic rice (Oryza sativa L.) plants that exhibit negligible genomic changes. This is in contrast with recently published results showing relevant changes in the DNA of transgenic rice plants generated through protoplasts electroporation and of transgenic poplar plants engineered through Agrobacterium tumefaciens infection. Implications of these findings are discussed in the context of selecting appropriate gene transfer methodologies to produce transgenic plants expressing genes of interest while retaining their genomic integrity and, thus, their superior agronomic and/or industrial traits.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009627409668
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  • 11
    Electronic Resource
    Electronic Resource
    Effective control of yellow stem borer and rice leaf folder in transgenic rice indica varieties Basmati 370 and M 7 using the novel δ-endotoxin cry2A Bacillus thuringiensis gene (1998)
    Springer
    Molecular breeding 4 (1998), S. 501-507 
    Details
    ISSN: 1572-9788
    Keywords: Bacillus thuringiensis (Bt) ; cry2A ; particle bombardment ; rice leaf folder ; transgenic rice ; yellow stem borer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Transgenic rice indica varieties Basmati 370 and M 7 expressing the novel cry2A (Bt) insecticidal gene were generated by particle bombardment. Molecular and biochemical analyses in R0 and R1 populations confirmed stable integration and expression of this novel Bt transgene. We estimated that the gene product was expressed up to 5% of total leaf protein. Insect feeding bioassays demonstrated that the Cry2A protein was effective against the yellow stem borer and the rice leaf folder, two major rice pests in the Indian Subcontinent. This is the first report of the control of major rice pests using this specific Bt gene. The cry2A gene can now be used in combination with other insecticidal genes for pyramiding resistance against insect pests. This will delay, or perhaps in combination with integrated pest management practices, prevent evolution of insect populations resistant to single insecticidal genes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009660315970
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  • 12
    Electronic Resource
    Electronic Resource
    Transgenic beans (Phaseolus vulgaris L.) engineered to express viral antisense RNAs show delayed and attenuated symptoms to bean golden mosaic geminivirus (1998)
    Springer
    Molecular breeding 4 (1998), S. 491-499 
    Details
    ISSN: 1572-9788
    Keywords: BGMV ; common bean ; particle bombardment ; plant transformation ; virus resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The genes Rep-TrAP-REn and BC1 from the Brazilian isolate bean golden mosaic geminivirus (BGMV-BR) were cloned in antisense orientation under the transcriptional control of the CaMV 35S promoter. This construct was used to transform common bean (Phaseolus vulgaris L.) using the biolistic method. Transgenic plants from the R3 and R4 generations were challenged by inoculation with a BGMV-BR viruliferous whitefly population. Of the four transgenic lines tested, two had both delayed and attenuated viral symptoms. Un-transformed plants or plants transformed with a construct containing only the gus-neo gene developed typical BGMV-BR symptoms 10–15 days after inoculation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009613607559
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  • 13
    Electronic Resource
    Electronic Resource
    Tissue-specificity of maize sucrose synthase gene promoters in maize tissues after particle bombardment (1998)
    Springer
    Euphytica 103 (1998), S. 17-21 
    Details
    ISSN: 1573-5060
    Keywords: particle bombardment ; promoter ; tissue-specificity sucrose synthase ; transient expression ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The reporter gene encoding β-glucuronidase (GUS) driven by either of the two maize sucrose synthase gene (Sh1 and Sus 1) promoters was introduced and expressed in various maize tissues via particle bombardment. Transient gene expression was examined by histochemical assays. It was found that the two SS promoters directed differential GUS expression. In the developing kernel, the Sh1 promoter was active only in the upper and central parts of the endosperm. In contrast, strong GUS activity controlled by the Sus1 promoter was detected in various types of cells, including the aleurone cells, the subaleurone endosperm cells, the scutellar cells of the embryo and the pericarp cells. Both promoters showed similar expression patterns in vegetative tissues.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1018326915934
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  • 14
    Electronic Resource
    Electronic Resource
    Expression and Immunolocalisation of the Snowdrop Lectin, GNA in Transgenic Rice Plants (1998)
    Springer
    Transgenic research 7 (1998), S. 371-378 
    Details
    ISSN: 1573-9368
    Keywords: rice transformation ; GNA ; particle bombardment ; immunolocalisation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transgenic rice (Oryza sativa L.) plants generated through particle bombardment expressed high levels of an insecticidal protein (the snowdrop lectin, GNA) directed against sap-sucking insects. Engineered plants expressed GNA either constitutively or in a tissue specific manner, depending on the nature of the promoter used to drive expression of the gene. We used specific antibodies raised against GNA to localize its expression in phloem tissue in plants engineered with the rice sucrose synthase promoter driving GNA expression. We report here molecular, biochemical and immunological analyses for fifteen independently-derived transformants out of more than 200 plants we generated.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008856703464
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  • 15
    Electronic Resource
    Electronic Resource
    Comparison of the activities of CaMV 35S and FMV 34S promoter derivatives in Catharanthus roseus cells transiently and stably transformed by particle bombardment (1997)
    Springer
    Plant molecular biology 33 (1997), S. 943-946 
    Details
    ISSN: 1573-5028
    Keywords: CaMV 35S promoter ; Catharanthus roseus ; FMV 34S promoter ; particle bombardment ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Activities of several CaMV 35S and FMV 34S promoter derivatives fused to the gusA reporter gene were compared in suspension-cultured Catharanthus roseus cells that were transiently and stably transformed using particle bombardment. Our data demonstrate that the 35S and a deletion derivative of the 34S promoter combined with particle bombardment form useful tools for genetic engineering of C. roseus cells. Our results disagree on several points with activities of 35S and 34S promoter derivatives reported for tobacco, indicating that absolute and relative promoter activities can differ between plant species.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005763605355
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  • 16
    Electronic Resource
    Electronic Resource
    Transgenic cotton resistant to herbicide bialaphos (1997)
    Springer
    Transgenic research 6 (1997), S. 385-392 
    Details
    ISSN: 1573-9368
    Keywords: transgenic cotton ; bialaphos ; particle bombardment ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Resistance to bialaphos, a non-selective herbicide, was intro duced into cotton through genetic engineering. A gene encoding phosphinothric in acetyltransferase (bar) from Streptomyces hygroscopicus was inserted into elite varieties of cotton through particle bombardment. Based on the marker gene, β-glucuronidase (gus) expression, a total of 18 Pima (Gossypium barbadense), 45 DP50 (G. hirsutum L.), 20 Coker 312 (G. hirsutum) and 2 El Dorado (G. hirsutum) transgenic plants were recovered. Integration of the bar gene into cotton genomic DNA was confirmed by Southern blot analysis and gene expression was confirmed by northern blot and enzyme assays. Herbicide (Basta®) tolerance up to 15 000 ppm was demonstrated in greenhouse trials. The newly introduced herbicide tolerance trait is inherited in a Mendelian fashion in the progenies of germline transformants. This study demonstrates the potential for particle bombardment to introduce commerically important genes directly into elite varieties of cotton. This mode of gene transfer can expedite the introduction of transgenic cotton products into world markets
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1018483300902
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  • 17
    Electronic Resource
    Electronic Resource
    The application of gene transfer techniques to marine resource management: recent advances, problems and future directions (1997)
    Springer
    Hydrobiologia 352 (1997), S. 263-278 
    Details
    ISSN: 1573-5117
    Keywords: transgenic fish ; gene transfer ; growth enhancement ; lopifection ; particle bombardment ; electroporation ; fish sperm ; fish embryos
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Recent advantages in transgenic fish research are reviewed, with special reference to the methods for gene transfer. These include microinjection, electroporation, particle bombardment, and lipofection. The success and problems associated with each of these methods, and the possible applications of transgenic fish research to aquaculture are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1003000127867
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  • 18
    Electronic Resource
    Electronic Resource
    Rice transformation: bombardment (1997)
    Springer
    Plant molecular biology 35 (1997), S. 197-203 
    Details
    ISSN: 1573-5028
    Keywords: genetic engineering ; particle bombardment ; plant biotechnology ; transgenic rice ; Oryza sativa
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Bombardment-based methodology is responsible for the effective genetic manipulation of major cereals including rice. Many groups reported significant advances on various aspects of rice molecular biology and genetic engineering using procedures based on bombardment technology. Molecular and genetic characterization of large numbers of these plants (more than 500 independent transgenic plants) provided information on structure, expression and stability of integrated DNA through multiple generations. Such evaluations were carried out in the greenhouse and in the field. Stability of expression was found to be dependent on the nature of the promoter and the transgene, and in specific cases on gene copy number. Direct DNA transfer utilizing particle bombardment for the delivery of foreign DNA into rice tissue results in the recovery of large numbers of independently derived transgenic plants in a variety-independent fashion. Gene copy number, level and stability of expression of transgenes can be compared to other DNA delivery methods, direct or indirect, including Agrobacterium-mediated gene transfer. In this paper, the technology is summarized and discussed in terms of present and future applications, including field trials and potential commercialization of transgenic rice expressing a number of genes of agronomic interest such as pest and herbicide resistance.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005791230345
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  • 19
    Electronic Resource
    Electronic Resource
    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
    Keywords: Transformation ; particle bombardment ; Agrobacterium ; Allium cepa.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: 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.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00231596
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  • 20
    Electronic Resource
    Electronic Resource
    Structural characterization of genes corresponding to cotton fiber mRNA, E6: reduced E6 protein in transgenic plants by antisense gene (1996)
    Springer
    Plant molecular biology 30 (1996), S. 297-306 
    Details
    ISSN: 1573-5028
    Keywords: cotton fiber ; transgenics ; particle bombardment ; fiber properties ; cDNA and genes ; nucleotide sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Two genes, each corresponding to fiber mRNA E6, were isolated from cotton cultivars Coker 312 (Gossypium hirsutum L.) and Sea Island (G. barbadense L.). E6 is one of the predominant fiber-specific mRNAs present during early fiber development. The distinguishing feature of the nucleotide-derived E6 protein is the presence of a motif where a dimer, Ser-Gly, is repeated several times. Two of the Sea Island genes contained a pentameric motif, Ser-Gly, while one of the Coker genes had one and the other had four motifs each. cDNA clones containing one or five Ser-Gly motifs were also identified. Thus, it appears that the strict conservation of this motif may not be critical to E6 protein function. Sequence characterizations of the genes and cDNAs showed that multiple members of the E6 family are transcribed in fiber and may result in proteins 238 to 246 amino acids long. The 3′ ends of the genes and cDNAs showed considerable heterology among them. Transgenic plants containing antisense genes were generated to decipher E6 function. Transgenic cotton with reduced E6 protein levels in the range of 60 to 98% were identified. However, no discernible phenotypic changes in fiber development or properties were apparent. This result leads to the conclusion that E6 is not critical to the normal development or structural integrity of cotton fibers.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00020115
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  • 21
    Electronic Resource
    Electronic Resource
    Stable transformation of cultured cells of the liverwortMarchantia polymorpha by particle bombardment (1996)
    Springer
    Transgenic research 5 (1996), S. 337-341 
    Details
    ISSN: 1573-9368
    Keywords: cultured liverwort cells ; hygromycin phosphotransferase (HPT) ; Marchantia polymorpha ; particle bombardment ; stable transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Suspension-cultured cells (A-18 line) of the liverwortMarchanta polymorpha were bombarded by a pneumatic particle gun with plasmid pCH harbouring the hygromycin phosphotransferase (HPT) gene (hpt) under the control of the cauliflower mosaic virus (CaMV) 35 S promoter and the nopaline synthase polyadenylation region. Nine weeks after bombardments, 128 hygromycin-resistant calluses were obtained from an approximate total of 7×106 cells. Ten cell lines chosen randomly were analysed further. Southern blot analysis showed that all of the ten lines contain thehpt gene in the genome, demonstrating that these lines are transformants. An HPT enzyme activity assay confirmed the expression of the gene in all of the transformant lines.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01968943
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  • 22
    Electronic Resource
    Electronic Resource
    Development, field evaluation, and agronomic performance of transgenic herbicide resistant rice (1996)
    Springer
    Molecular breeding 2 (1996), S. 359-368 
    Details
    ISSN: 1572-9788
    Keywords: bar gene ; glufosinate ; herbicide tolerance ; particle bombardment ; Oryza sativa L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The commerical cultivars ‘Gulfmont’, ‘IR72’ and ‘Koshihikari’ were genetically engineered using electric discharge particle bombardment to express the bar gene which confers resistance to the broad-range herbicide glufosinate. Southern and northern blot analyses of transgenics material revealed stable integration and expression of introduced transgenes in the lines evaluated. In a few plants, silencing of the uidA marker gene was detected at the transcriptional level. Field studies were conducted in 1993 and 1994 at the Rice Research Station near Crowley, LA. This report summarizes results from the first two years of field trial for transgenic Gulfmont and Koshihikari. Transgenic cultivar IR72 was tested in 1995 and preliminary results are similar to those reported for transgenic Gulfmont. All 11 independently derived transgenic lines produced fertile, normal looking seed at maturity. Significant differences were observed in the absence of the herbicide between parental cultivars and transgenic Gulfmont-and Koshihikari-derived lines for days to 50% heading (20% of transgenic lines), plant height (13%), and grain yield (7%). Foliar application of glufosinate had little or no effect on agronomic performance of all transgenic Gulfmont and IR72 lines, while herbicide applications affected grain, yield and plant height of some transgenic Koshhikari. Non-transgenic plants of all three cultivars at the 4-leaf stage were killed within 7 days after 1.12 or 2.24 kg/ha glufosinate applications. Significant differences among certain transgenic lines were observed for agronomic traits after herbicide applications. These results demonstrate that the bar gene was effective in conferring field-level resistance to glufosinate in rice. Variation among transgenic lines required traditional breeding selection procedures to identify superior agronomic types with high levels of herbicide resistance and showed the necessity to generate several independent transgenic lines of each cultivar.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00437914
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  • 23
    Electronic Resource
    Electronic Resource
    Production of transgenic cassava (Manihot esculenta Crantz) plants by particle bombardment using luciferase activity as selection marker (1996)
    Springer
    Molecular breeding 2 (1996), S. 339-349 
    Details
    ISSN: 1572-9788
    Keywords: cassava ; genetic modification ; luciferase ; particle bombardment
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Cassava embroids derived from friable embryogenic callus of the genotype TMS60444 were bombarded with DNA of the constructs pJIT100 or pJIT64. Both constructs contain the non-invasive reporter gene luciferase from firefly driven by the CaMV 35S promoter. The influence of several particle gun machine parameters and pretreatment of plant material on transient luciferase activity were studied to determine the most essential conditions for stable transformation. Two weeks after bombardment pieces of friable calli with luciferase activity were selected. In total, 67 independent selected calli with luciferase activity (spots), derived from five different experiments, were further cultured either in liquid or on solid medium. Per plate or flask one spot was cultured. In subsequent selection rounds all spots of one individual plate or flask were cultured as one individual group. In this way different transformation events were separated and multiplied. Eight weeks after bombardment 34 cultures still contained luciferase activity. The mean number of luciferase spots per culture had increased from 1 to 4.6 spots in liquid and to 2.5 spots on solid medium. After two more months of subsequent culture and luciferase selection presence of the construct in these cultures was confirmed at the molecular level using the polymerase chain reaction assay and Southern analysis. Friable embryos derived from four transformation events were cultured for maturation. Between 3% and 21% of the mature embryos of the different transformation events were luciferase-positive. After multiplication of the luciferase-positive mature embryos by secondary somatic embryogenesis they were germinated. The plantlets analysed contained one to several copies of the inserted DNA. The method presented enables the transformation of this particular cassava genotype, thus allowing the genetic improvement of this important tropical crop by transgenesis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00437912
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  • 24
    Electronic Resource
    Electronic Resource
    Production of fertile transgenic barley (Hordeum vulgare L.) plant using the hygromycin-resistance marker (1995)
    Springer
    Plant cell reports 14 (1995), S. 329-334 
    Details
    ISSN: 1432-203X
    Keywords: Barley ; Hordeum vulgare ; immature embryo ; particle bombardment ; particle gun ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Fertile transgenic barley (Hordeum vulgare L.) plants were obtained by high velocity particle bombardment. The plasmid pBCl was used to deliver the selectable hph gene and reporter Gus gene into immature embryo. After the selection culture 18 hygromycin resistant plants were obtained. Samples for Southern hybridization and enzymatic Gus assay were obtained from 11 plants. Southern hybridization confirmed the presence of the hph gene in the 11 hygromycin resistant plants(T0). Enzymatic assay indicated that all the t0 plants that showed hph positive in Southern analysis possessed detectable amount of Gus activity. To date all the 11 t0 plants reached maturity and mature seeds were obtained Transmission of the hph gene to progeny(T1) of two independent t0 plants was confirmed by Southern hybridization.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00232038
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  • 25
    Electronic Resource
    Electronic Resource
    Promoter analysis of seed storage protein genes from Canavalia gladiata D.C. (1995)
    Springer
    Plant molecular biology 27 (1995), S. 729-741 
    Details
    ISSN: 1573-5028
    Keywords: Canavalia gladiata ; canavalin ; concanavalin A ; particle bombardment ; transgenic tobacco ; 5′-upstream sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A number of A/T-rich sequences and a CATGCAT/A sequence are contained in the 5′-upstream regions of the genes encoding concanavalin A (Con A) and canavalin, two major seed storage proteins of Canavalia gladiata D.C. To study the role of these sequences in the seed-specific gene expression, we constructed 5′-deletion mutants and examined the transient expression of β-glucuronidase reporter gene by particle bombardment and the stable expression by Agrobacterium-mediated transformation of tobacco plants. Positive regulatory elements were located in the −894/−602 and −602/−74 regions of the Con A gene, and in the −428/−376, −281/−155 and −155/−50 regions of the canavalin gene. In addition, the results suggested that the A/T-rich sequences in the 5′-upstream region of the Con A gene play a role in transcriptional activation, but that those of the canavalin gene have little effect on the gene expression. The CATGCAT/A sequence was not sufficient by itself for high levels of expression of both the Con A and canavalin genes. The canavalin polypeptide amounted to about 1% of the total extractable protein in the transgenic tobacco seeds, but the Con A polypeptide was not detected in the extractable protein.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00020226
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  • 26
    Electronic Resource
    Electronic Resource
    Successful expression in pollen of various plant species of in vitro synthesized mRNA introduced by particle bombardment (1995)
    Springer
    Plant molecular biology 28 (1995), S. 337-341 
    Details
    ISSN: 1573-5028
    Keywords: Lilium longiflorum ; Freesia refracta ; Tulipa gesneriana ; GUS mRNA ; particle bombardment ; pollen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Gold particles coated with β-glucuronidase (GUS) mRNA with a 5′ cap structure that had been synthesized in vitro were introduced, by use of a pneumatic particle gun, into pollen grains of lily (Lilium longiflorum), freesia (Freesia refracta) and tulip (Tulipa gesneriana). A fluorometric assay for the GUS activity indicated that in vitro synthesized GUS mRNA introduced into these pollen cells by particle bombardment was successfully expressed. GUS activity in extracts of the bombarded lily pollen became detectable fluorometrically within 30 min after bombardment, peaked at 6 h, then gradually decreased. This activity changed as a function of the developmental stage of the pollen cell of lily.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00020252
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  • 27
    Electronic Resource
    Electronic Resource
    Activity of constitutive promoters in various species from the Liliaceae (1995)
    Springer
    Plant molecular biology 28 (1995), S. 949-955 
    Details
    ISSN: 1573-5028
    Keywords: Liliaceae ; tulip ; lily ; leek ; monocot intron ; particle bombardment ; promoter activity ; transient expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In this paper we first review literature on the performance of various promoters in monocotyledonous species. In general, promoters isolated from monocots show a higher activity in monocot species. Moreover, the presence of an intron between the promoter and reporter gene increases transcription levels. We used the same approach to study gene expression in Liliaceae. The activities of the CaMV 35S, maize Adh1-based pEmu, rice Act1 and maize Ubi promoters, coupled to the β-glucuronidase (gus) reporter gene, were evaluated for transient gene expression upon particle bombardment of tissues of tobacco, rice, tulip, lily and leek. Although monocot promoters performed very well in rice tissues, the results of this study show that this cannot be generalized for other monocot species. The transcription inducing effects of monocot promoters were less pronounced or even absent in tissues of Liliaceae, while the presence of an intron between promoter and gus gene reduced promoter activity.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00042079
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  • 28
    Electronic Resource
    Electronic Resource
    The impact of selection parameters on the phenotype and genotype of transgenic rice callus and plants (1995)
    Springer
    Transgenic research 4 (1995), S. 44-51 
    Details
    ISSN: 1573-9368
    Keywords: Oryza sativa ; transgenic rice plants ; particle bombardment ; selection parameters ; hygromycin resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transgenic rice embryogenic callus and plants were recovered from experiments involving electric discharge particle acceleration (AccellTM technology). Critical parameters influencing successful delivery and stable integration of exogenous DNA into organized rice tissue had been identified previously. We report here on the effects of one selective agent (hygromycin B) on the phenotype and genotype of recovered callus and plants. The nature, timing and culture practices appeared to be more critical for the successful recovery of transgenic callus and plants than the level of selection used. By utilizing the procedures described in this report, transformation frequencies well in excess of 10% were obtained routinely in all varieties of rice tested. The combination of AccellTM technology with a selectable and prolific regeneration culture system resulted in the development of a variety-independent and highly efficient method for the routine introduction of any gene into any rice variety.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01976501
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  • 29
    Electronic Resource
    Electronic Resource
    Transgenic haploid plants ofNicotiana rustica produced by bombardment-mediated transformation of pollen (1995)
    Springer
    Transgenic research 4 (1995), S. 341-348 
    Details
    ISSN: 1573-9368
    Keywords: transgenic haploid plants ; pollen culture ; particle bombardment ; β-glucuronidase ; neomycin phosphotransferase II
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Immature pollen fromNicotiana rustica was bombarded with gold particles coated with plasmid DNA encoding neomycin phosphotransferase II (NPTII) and β-glucuronidase (GUS) genes which, respectively, are under the control of the cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (NOS) terminator in the plasmid. Kanamycin-resistant pollen embryoids were selected from the bombarded pollen cells and two independent lines of transgenic plants were regenerated. Enzyme assays showed that one has both NPTII and GUS activities and the other only weak NPTII activity. Southern blot analyses indicated that the former has a DNA fragment corresponding to the intact expression cassettes for both genes in its genome; whereas the latter lacks intact expression cassettes for both genes and has only the intactnptII coding sequence in its genome. The transgenic plants of both lines have 24 chromosomes, confirming haploidy, and they are infertile. These results indicate that transgenic haploid plants can be produced directly by the bombardment-mediated transformation of immature pollen.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01972531
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  • 30
    Electronic Resource
    Electronic Resource
    Transformation studies in Hordeum vulgare using a highly regenerable microspore system (1955)
    Springer
    Euphytica 85 (1955), S. 113-118 
    Details
    ISSN: 1573-5060
    Keywords: Hordeum vulgare ; isolated microspores ; particle bombardment ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary A highly regenerable, isolated microspore system for barley, Hordeum vulgare L. cv. Igri, has been developed which is amenable to transformation studies using particle bombardment. The system allows DNA to be delivered to microspores at the single cell stage and both transient and stable transformation events have been demonstrated. The potential advantages of using isolated microspores as the target tissue in routine transformation systems are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023938
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  • 31
    Electronic Resource
    Electronic Resource
    Transgenic barley by particle bombardment. Inheritance of the transferred gene and characteristics of transgenic barley plants (1955)
    Springer
    Euphytica 85 (1955), S. 81-88 
    Details
    ISSN: 1573-5060
    Keywords: gene transfer ; Hordeum vulgare ; neomycin phosphotransferase II ; particle bombardment ; transgenic barley
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Transgenic barley plants (Hordeum vulgare L. cv. Kymppi) were obtained by particle bombardment of various tissues. Immature embryos and microspore-derived cultures were bombarded with gold particles coated with plasmid DNA carrying the gene coding for neomycin phosphotransferase II (NPTII), together with plasmid DNA containing the gene for β-glucuronidase (GUS). Bombarded immature embryos were grown to plants without selection and NPTII activity was screened in small plantlets. One plant proved to be transgenic (T0). This chimeric plant passed the transferred nptII gene to its T1 progeny. The presence of the nptII gene was demonstrated by the PCR technique and enzyme activity was analyzed by an NPTII gel assay. Four T0 spikes and 15 T1 offspring were transgenic. The integration and inheritance was confirmed by Southern blot hybridization. Transgenic T2 and T3 plants were produced by isolating embryos from green grains of transgenic T1 and T2 plants, respectively and growing them to plants. After selfing, the ratio of transgenic to non-transgenic T2 offspring was shown to follow the rule of Mendelian inheritance. The general performance of transgenic plants was normal and no reduction in fertility was observed. Microspore-derived cultures were bombarded one and four weeks after microspore isolation. After bombardment, cultures were grown either with or without antibiotic selection (geneticin R or kanamycin). When cultures were grown without selection and regenerated plants were transferred to kanamycin selection in rooting phase, one out of a total of about 1500 plants survived. This plant both carried and expressed the transferred nptII gene. The integration was confirmed by Southern blot hybridization. This plant was not fertile.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023933
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  • 32
    Electronic Resource
    Electronic Resource
    Genetic engineering of cereal crop plants: a review (1955)
    Springer
    Euphytica 85 (1955), S. 35-44 
    Details
    ISSN: 1573-5060
    Keywords: cereals ; protoplast transformation ; tissue electroporation ; particle bombardment
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Many aspects of basic and applied problems in plant biology can be investigated by transformation techniques. In dicotyledonous species, the ability to generate transgenic plants provides the tools for an understanding of plant gene function and regulation as well as for the directed transfer of genes of agronomic interest. For many dicotyledonous plants Agrobacterium tumefaciens can be routinely used to introduce foreign DNA into their genome. However, cereals seem to be recalcitrant to Agrobacterium-mediated transformation. In cereals, many efforts have been made in recent years to establish reliable transformation techniques. Several transformation techniques have been developed but to date only three methods have been found to be suitable for obtaining transgenic cereals: transformation of totipotent protoplasts, particle bombardment of regenerable tissues and, more recently, tissue electroporation. The current state of transformation methods used for cereals will be reviewed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023928
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  • 33
    Electronic Resource
    Electronic Resource
    Strategies for variety-independent genetic transformation of important cereals, legumes and woody species utilizing particle bombardment (1955)
    Springer
    Euphytica 85 (1955), S. 13-27 
    Details
    ISSN: 1573-5060
    Keywords: gene transfer ; crop species ; particle bombardment ; transgenic plants ; cereals ; legumes ; woody plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The limiting component in the creation of transgenic crops has been the lack of effective means to introduce foreign genes into elite germplasm. However, the development of novel direct DNA transfer methodology, by-passing limitations imposed by Agrobacterium-host specificity and cell culture constraints, has allowed the engineering of almost all major crops, including formerly recalcitrant cereals, legumes and woody species. The creation of transgenic rice, wheat, maize, barley, oat, soybean, phaseolus, peanut, poplar, spruce, cotton and others, in an efficient and in some cases, variety-independent fashion, is a significant step towards the routine application of recombinant DNA methodology to the improvement of most important agronomic crops. In this review we will focus on key elements and advantages of particle bombardment technology in order to evaluate its impact on the accelerated commercialization of products based on agricultural biotechnology and its utility in studying basic plant developmental processes and function through transgenesis. Fundamental differences between conventional gene transfer methods, utilizing Agrobacterium vectors or protoplast/suspension cultures, and particle bombardment will be discussed in depth.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023926
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