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  • 2020-2022
  • 1995-1999  (17)
  • 1955-1959
  • 1920-1924
  • 1840-1849
  • 1995  (17)
  • particle bombardment
  • protoplasts
  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell reports 14 (1995), S. 738-742 
    ISSN: 1432-203X
    Keywords: Hyoscyamus muticus ; plant regeneration ; protoplasts ; transformed roots
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Successful plant regeneration was achieved for the first time from hairy root-derived protoplasts of Hyoscyamus muticus. High yields (7 × 106 / g fresh weight) of protoplasts were isolated directly from the transformed roots of Hyoscyamus muticus using an enzyme mixture comprising 1 % macerozyme and 2 % cellulase in an osmoticum consisting of 0.2 M CaCl2 and 0.6 M mannitol. Protoplasts were first cultured in liquid NT/PRO I medium and further on semi-solid NT/PRO II agar medium. The procedure permits highly efficient formation of colonies. The plating efficiency varied from 1–9 %. The small individual colonies regenerated easily into shoots and roots at frequencies of 18 % and 70 %, respectively. The time required for the development of small plantlets from protoplasts was 8–11 weeks. The regenerated plants contained rolB from Ri-T-DNA and exhibited an altered phenotype compared to the control plants.
    Type of Medium: Electronic Resource
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  • 2
    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.
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 28 (1995), S. 123-136 
    ISSN: 1573-5028
    Keywords: Agrobacterium rhizogenes ; auxin biosynthesis genes ; plant cell division ; protoplasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Plant oncogenes aux1 and aux2 carried by the TR-DNA of Agrobacterium rhizogenes strain A4 encode two enzymes involved in the auxin biosynthesis pathway in transformed plant cells. The short divergent promoter region between the two aux-coding sequences contains the main regulatory elements. This region was fused to the uidA reporter gene and introduced into Nicotiana tabacum in order to investigate the regulation and the tissue specificity of these genes. Neither wound nor hormone induction could be detected on transgenic leaf discs. However, phytohormone concentration and auxin/cytokinin balance controlled the expression of the chimaeric genes in transgenic protoplasts. The expression was localised in apical meristems, root tip meristems, lateral root primordia, in cells derived from transgenic protoplasts and in transgenic calli. Histological analysis showed that the expression was located in cells reactivated by in vitro culture. Experiments using cell-cycle inhibitors such as hydroxyurea or aphidicolin on transgenic protoplast cultures highly decreased the β-glucuronidase activity of the chimaeric genes. These results as well as the histological approach suggest a correlation between expression of the aux1 and aux2 genes and cell division.
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  • 4
    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.
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  • 5
    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.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 28 (1995), S. 949-955 
    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.
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  • 7
    ISSN: 1573-5028
    Keywords: Agrobacterium tumefaciens ; extensin ; hydroxyproline-rich cell wall protein ; Nicotiana sylvestris ; protoplasts ; wounding
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA clone (6PExt 1.2) encoding a novel extensin was isolated from a cDNA library made from 6 h old mesophyll protoplasts of Nicotiana sylvestris. The screening was performed with a heterologous probe from carrot. The encoded polypeptide showed features characteristic of hydroxyproline-rich glycoproteins such as Ser-(Pro)4 repeats and a high content in Tyr and Lys residues. The presence of four Tyr-X-Tyr-Lys motifs suggests the possibility for intramolecular isodityrosine cross-links whereas three Val-Tyr-Lys motifs may participate in intermolecular cross-links. The analysis of genomic DNA gel blots using both the N. sylvestris and the carrot clones as probes showed that the 6PExt 1.2 gene belongs to a complex multigene family encoding extensin and extensin-related polypeptides in N. sylvestris as well as in related Nicotianeae including a laboratory hybrid. This was confirmed by the analysis of RNA gel blots: a set of mRNAs ranging in size from 0.3 kb to 3.5 kb was found by the carrot extensin probe. The 6PExt 1.2 probe found a 1.2 kb mRNA in protoplasts and in wounded tissues as well as a 0.9 kb mRNA which seemed to be stem-specific. The gene encoding 6PExt 1.2 was induced by wounding in protoplasts, in leaf strips and after Agrobacterium tumefaciens infection of stems.
    Type of Medium: Electronic Resource
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  • 8
    ISSN: 1432-203X
    Keywords: garlic ; Allium sativum L. ; shoot primordia ; protoplasts ; plant regeneration
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts derived from tissue-cultured shoot primordia of garlic (Allium sativum L.) initiated successive cell divisions within 4 days and formed small individual calli (0.2mm in diameter) after 5 weeks of culture on Gamborg's B5 medium supplemented with 0.1% casein hydrolysate, 1mg/1 1-naphthaleneacetic acid and 1mg/1 6-benzylaminopurine. Plating efficiency was roughly 5% at the density of 1x104 protoplasts/ml of medium. Adventitious buds developed from the calli during subsequent subculture on Gamborg's B5 medium supplemented with 40mg/l adenine and 10% coconut milk. When transferred to the same medium without supplements, these buds grew into shoots and rooted. The regenerated garlic plantlets were successfully transferred to the greenhouse and grew into whole plants.
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Transgenic research 4 (1995), S. 44-51 
    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.
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  • 10
    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.
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  • 11
    ISSN: 1573-5176
    Keywords: protoplasts ; flow cytometry ; seaweed ; biotechnology
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The ability to rapidly distinguish viable sub-populations of cells within populations of macroalgal protoplast isolations was demonstrated using flow cytometry. Viable protoplasts from Ulva sp. and Porphyra perforata J. Ag. were distinguished from non-viable protoplasts based on differential fluorescein accumulation. The identities of cortical and epidermal protoplasts from Macrocystis pyrifera (L.) C. Ag. were inferred based on light-scattering and chlorophyll a autofluorescence. Three cell types could be distinguished among protoplasts released from thalli of P. perforata based on chlorophyll a and phycoerythrin autofluorescence. Mixed protoplast populations of Ulva sp. and P. perforata were also discernable based on relative chlorophyll a and phycoerythrin autofluorescence. The ability to screen heterogenous protoplast populations rapidly, combined with the cell sorting capabilities of many flow cytometers, should prove valuable for seaweed biotechnology.
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  • 12
    ISSN: 1573-5044
    Keywords: Hordeum vulgare ; plasmid uptake ; polyethylene glycol (PEG) ; protoplasts ; transient gene expression ; stable transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cat gene expression has been investigated following PEG-mediated plasmid uptake into barley protoplasts. The uptake conditions optimised for transient expression were employed for stable transformation. Transformed protoplast-derived calli of the cvs. Dissa and Igri, were selected on medium containing G418 at 40 μg ml−1 or kanamycin sulphate at 250 μg ml−1. Absolute transformation frequencies of 28.9×10−5 and 21.3×10−5 were recorded for Dissa with kanamycin sulphate and G418 selection, respectively. The frequency for Igri was 11.5×10−5 with G418 selection. Antibiotic resistant protoplast-derived colonies expressed NPTII activity; Southern hybridisation confirmed integration of the nptII gene into barley genomic DNA.
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  • 13
    ISSN: 1573-5036
    Keywords: Al tolerance ; 1 ; 3-β-glucan ; K leakage ; plasma membrane ; protoplasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Experiments were carried out to identify the primary site for aluminium (Al) toxicity in roots. Al accumulated in large amounts in the younger and outer cells in roots of pea and was retarded when the ionic strength of the Al solution was high. Cell destruction was extensive in the regions with high Al accumulation. The accumulation of Al in, and potassium (K) leakage from, the root tip were in the order pea〉maize〉rice, the same order as their sensitivity to Al. The protoplasts from the root tip portion of pea incubated with Al showed a wrinkled and uneven surface. The protoplasts progressively shrank and eventually collapsed. Viability decreased in this process. In the control protoplasts of maize, β-glucan formation was uniform on the spherical surfaces, whereas it was spotty in the Al-treated protoplasts; the cell wall material of the latter contained partly 1, 3-β-glucan which is known to be synthesised by 1, 3-β-glucan synthase embedded in the plasma membrane. These results suggest that the specific site for Al toxicity is the plasma membrane of younger and outer cells in roots and that Al tolerance depends largely on the integrity of the plasma membrane.
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  • 14
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell, tissue and organ culture 41 (1995), S. 41-48 
    ISSN: 1573-5044
    Keywords: callus ; Medicago littoralis ; plant regeneration ; protoplasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Plant regeneration from explant and protoplast derived callus has been achieved in Medicago littoralis cv. Harbinger 1886, an annual legume resistant to the fungus Pseudopeziza medicaginis. Callus was induced from different tissue explants and the fastest growth rate was observed for hypocotyls in B5 medium with 2 mg l−1 2,4-dichlorophenoxyacetic acid and 0.5 mg l−1 N6-benzyladenine. Protoplasts were isolated from cotyledons and leaves of sterile plants and from callus; the first two kinds of protoplasts showed a plating efficiency of 5.6% and 5%, respectively, when embedded in agarose. Plant regeneration occurred on media containing % MathType!MTEF!2!1!+-% feaafiart1ev1aaatCvAUfeBSjuyZL2yd9gzLbvyNv2CaerbuLwBLn% hiov2DGi1BTfMBaeXatLxBI9gBaerbd9wDYLwzYbItLDharqqtubsr% 4rNCHbGeaGqiVu0Je9sqqrpepC0xbbL8F4rqqrFfpeea0xe9Lq-Jc9% vqaqpepm0xbba9pwe9Q8fs0-yqaqpepae9qq-f0-yqaqVeLsFr0-vr% 0-vr0db8meaabaqaciGacaGaaeqabaWaaeaaeaaakeaacaqGobWaaW% baaSqabeaacaqG2aaaaOGaaeOVfiaabs5adaahaaWcbeqaaiaaikda% aaGccaqG+waaaa!3F97!\[{\text{N}}^{\text{6}} {\text{\Delta }}^2 {\text{}}\]isopentenyl-adenine combined with indole-3-acetic acid or 1,2-benzisoxazole-3-acetic acid, and on media with N6-benzyladenine plus α-naphtaleneacetic acid; a cytokinin/auxin ratio higher than 1 induced embryos while a ratio around 1 stimulated shoot formation. Embryo development and rooting of shoots were performed in RL medium without growth regulators.
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  • 15
    Electronic Resource
    Electronic Resource
    Springer
    Journal of industrial microbiology and biotechnology 15 (1995), S. 397-400 
    ISSN: 1476-5535
    Keywords: mycolytic enzymes ; Trichoderma harzianum ; protoplasts ; Rhizopus nigricans ; induction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The effect of the growth form of the inducing microorganism on specificTrichoderma harzianum mycolytic enzyme production was studied. The pelleted form ofRhizopus nigricans gave a better product concerning protoplast formation ability. The maximum yield of protoplasts from the target fungusCochliobolus lunatus was 1×108 ml−1. Analysis of individual specific enzyme activities inTrichoderma mycolytic enzyme preparations confirms the importance of high chitinase and low protease activity for high protoplast yields. Supplementation of the production medium with chitin increased the chitinase activity in theTrichoderma exoenzyme mixture.
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  • 16
    ISSN: 1573-5044
    Keywords: potato ; protoplasts ; regeneration ; Solanum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The response to different in vitro methods for use in potato breeding has been evaluated in 11 genotypes of 5 Solanum species, S. etuberosum, S. lycopersicoides, S. maglia, S. rickii, and S. tuberosum. Callus induction and growth, and shoot regeneration were strongly influenced by the genotype, explant source, and medium utilized. Furthermore, considerable differences among the 11 genotypes were found both in plating efficiency and shoot regeneration from protoplast culture. Some interesting correlations were found between different tissue culture responses, suggesting linkage and/or pleiotropic effect of genes. The potential application to potato breeding of the in vitro techniques analyzed is discussed.
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  • 17
    Electronic Resource
    Electronic Resource
    Springer
    World journal of microbiology and biotechnology 11 (1995), S. 409-415 
    ISSN: 1573-0972
    Keywords: Breeding ; embryo culture ; haploids ; micropropagation ; protoplasts ; synthetic seed ; transformation ; wide hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Plant tissue culture comprises a set of in vitro techniques, methods and strategies that are part of the group of technologies called plant biotechnology. Tissue culture has been exploited to create genetic variability from which crop plants can be improved, to improve the state of health of the planted material and to increase the number of desirable germplasms available to the plant breeder. Tissue-culture protocols are available for most crop species, although continued optimization is still required for many crops, especially cereals and woody plants. Tissueculture techniques, in combination with molecular techniques, have been successfully used to incorporate specific traits through gene transfer. In vitro techniques for the culture of protoplasts, anthers, microspores, ovules and embryos have been used to create new genetic variation in the breeding lines, often via haploid production. Cell culture has also produced somaclonal and gametoclonal variants with crop-improvement potential. The culture of single cells and meristems can be effectively used to eradicate pathogens from planting material and thereby dramatically improve the yield of established cultivars. Large-scale micropropagation laboratories are providing millions of plants for the commercial ornamental market and the agricultural, clonally-propagated crop market. With selected laboratory material typically taking one or two decades to reach the commercial market through plant breeding, this technology can be expected to have an ever increasing impact on crop improvement as we approach the new millenium.
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