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  • 1
    Electronic Resource
    Electronic Resource
    Transformation of lily by Agrobacterium (1955)
    Springer
    Euphytica 85 (1955), S. 97-100 
    Details
    ISSN: 1573-5060
    Keywords: Agrobacterium ; transformation ; lily ; β-glucuronidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: 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.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023935
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    Paper (German National Licenses)
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  • 2
    Electronic Resource
    Electronic Resource
    Conditional inhibition of β-glucuronidase expression by antisense gene fragments in petunia protoplasts (1993)
    Springer
    Plant molecular biology 23 (1993), S. 45-55 
    Details
    ISSN: 1573-5028
    Keywords: Key words ; antisense RNA ; β-glucuronidase ; protoplasts ; transient gene expression ; PCR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Antisense RNA-mediated inhibition of gene expression is a valuable tool to induce mutant phenotypes. We are interested in the application of antisense gene fragments with the aim to improve the efficiency of inhibition and to be able to selectively suppress gene family members in plants. Protoplasts may provide a rapid system to screen the efficiency of antisense gene segments. As a first step, we set up a transient expression system for leaf protoplasts of Petunia hybrida and used as a model system the inhibition of β-glucuronidase (uidA) expression by uidA antisense gene segments. Both GUS enzyme activities and uidA RNA levels were measured. Co-introducing equal amounts of a full-length uidA antisense gene and a uidA sense gene reduced GUS activity by 60–70%. Various uidA antisense fragments also inhibited expression although with different efficiencies and we show that strong antisense fragments can be retrieved from weak antisense gene fragments. A promoter-less antisense gene did not reduce uidA expression indicating that the inhibition is mediated by antisense transcripts. Using quantitative PCR on first-strand cDNA we show that expression of functional antisense genes lead to reduced levels of uidA mRNA. This suggests that the mechanism of antisense RNA inhibition in protoplasts is similar to that in transgenic plants and that the protoplast system in combination with PCR can be used to preselect antisense fragments of any gene.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00021418
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  • 3
    Electronic Resource
    Electronic Resource
    Transformation of four pathogenic Phytophthora spp by microprojectile bombardment on intact mycelia (1993)
    Springer
    Current genetics 23 (1993), S. 42-46 
    Details
    ISSN: 1432-0983
    Keywords: Phytophthora ; Transformation ; Biolistic ; β-glucuronidase ; Plant promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Phytophthora capsici, P. citricola, P. cinnamomi and P. citrophthora were transformed without the removal of cell walls by particle acceleration with plasmids containing the β-glucuronidase gene and hygromycin B resistance. Transformants were detected by histochemical and fluorometric β-glucuronidase assays and confirmed by Southern-blot hybridization. It was found that the promoter of a plant virus is functional in Phytophthora. In addition, a method was designed to visually identify homogeneous transformed colonies, derived from zoospores of transformed multinucleated Phytophthora mycelia, based on blue color development on plates containing X-Gluc.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336748
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  • 4
    Electronic Resource
    Electronic Resource
    The maize zein gene zE19 contains two distinct promoters which are independently activated in endosperm and anthers of transgenic Petunia plants (1990)
    Springer
    Plant molecular biology 15 (1990), S. 81-93 
    Details
    ISSN: 1573-5028
    Keywords: double promoter ; β-glucuronidase ; cell type specificity ; gene regulation ; seed storage protein gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The activity, tissue specificity and temporal expression of the tandem promoter region preceding a maize zein gene (zE19, encoding a 19 kDa zein protein) were tested in transgenic Petunia plants. To simplify the analysis, the tandem promoter as well as each of the two separate promoter regions were fused to the β-glucuronidase (GUS) reporter gene. All of the three constructs directed the synthesis of GUS in the endosperm of transformed seeds indicating that both separate promoters are independently activated and show the same tissue and cell type specificity observed for zein genes in maize. The kinetics of accumulation and the localization of GUS activity are not coordinated with those of Petunia endogenous seed storage proteins during the development of transformed seeds. Unexpectedly, we detected high levels of GUS activity in anthers of transformed Petunia plants for all three constructs. This appears to reflect the expression pattern of zein genes in maize, since we detect zein transcripts in anthers. Finally, we discuss the possible origin and function of the tandem promoter arrangement on the basis of these data.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00017726
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  • 5
    Electronic Resource
    Electronic Resource
    Levels and location of expression of the Agrobacterium tumefaciens pTiA6 ipt gene promoter in transgenic tobacco (1993)
    Springer
    Plant molecular biology 21 (1993), S. 1011-1021 
    Details
    ISSN: 1573-5028
    Keywords: Agrobacterium tumefaciens ; β-glucuronidase ; cytokinin ; ipt ; isopentenyl transferase ; Nicotiana tabacum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The location of gene expression of the Agrobacterium tumefaciens ipt gene promoter in transgenic tobacco plants was examined using the β-glucuronidase (GUS) reporter gene. Expression of GUS was detected in every organ and most cell types examined. The highest levels of GUS activity were found in roots. To further examine the transcriptional basis of this broad expression pattern, deletions in the 5′ noncoding region of the gene were translationally fused to two promoterless reporter genes, encoding the enzymes chloramphenicol acetyl transferase (CAT) and β-glucuronidase (GUS). Reporter enzyme assays revealed the existence of an upstream segment required for maximal promoter function, the 5′ end of which is between-442 and-408 of the Pipt ATG codon. This upstream segment is required for maximal levels of GUS expression in roots, but not in other organs, and a tobacco suspension-cultured cell line. The implications of broad ipt expression on the process of crown gall tumorigenesis are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023599
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  • 6
    Electronic Resource
    Electronic Resource
    Expression of GUS in primary transformants and segregation patterns of GUS, TL- and TR-DNA in the T1 generation of hairy root transformants ofLotus corniculatus (1994)
    Springer
    Transgenic research 3 (1994), S. 232-240 
    Details
    ISSN: 1573-9368
    Keywords: Agrobacterium rhizogenes ; Lotus corniculatus ; TL-DNA ; TR-DNA ; β-glucuronidase ; segregation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Agrobacterium rhizogenes was assessed as a vehicle for transformation ofLotus corniculatus. Plants were co-transformed usingA. rhizogenes strain LBA 9402 harbouring the bacterial plasmid pRi1855 and the binary transformation vector pJit 73. pRi 1855 transfers both TL and TR sequences, while pJit 73 encodes β-glucuronidase (GUS) and also two selectable marker genes giving resistance to the antibiotics kanamycin and hygromycin. Three primary transformants (lines 1,6 and 12) were subjected to detailed morphological and biochemical analysis and lines 6 and 12 were also analysed at the molecular level. Tissues of both lines 6 and 12 were resistant to hygromycin and expressed GUS. Analysis of various tissues of each line showed a significantly lower GUS activity in line 6 than in line 12. Genetical analysis of progeny produced between control plants and lines 6 and 12 indicated that line 6 had one dose of theuid gene while line 12 had two or more independently segregating doses of the gene. Both lines 6 and 12 contained multiple copies of TL-DNA, while only line 6 was TR positive. In the progeny of lines 6 and 12 there was no evidence for linkage of TL-DNA withuid, while in the progeny of line 6, TR-DNA was under-represented. GUS-positive progeny which were free of both TL and TR sequences were identified from both lines.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02336776
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  • 7
    Electronic Resource
    Electronic Resource
    Construction of an intron-containing marker gene: Splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacterium-mediated plant transformation (1990)
    Springer
    Molecular genetics and genomics 220 (1990), S. 245-250 
    Details
    ISSN: 1617-4623
    Keywords: Agrobacterium tumefaciens ; β-glucuronidase ; Portable intron ; Splicing ; Transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Agrobacterium tumefaciens is a commonly used tool for transforming dicotyledonous plants. The underlying mechanism of transformation however is not very well understood. One problem complicating the analysis of this mechanism is the fact that most indicator genes are already active in Agrobacterium, thereby preventing the precise determination of timing and localisation of T-DNA transfer to plant cells. In order to overcome this obstacle a modified prokaryotic indicator gene was constructed. The expression of this indicator gene and its use in analysing early events in Agrobacterium-mediated plant transformation are described. A portable intron, derived from a plant intron, was introduced into the β-glucuronidase (GUS) gene. In transgenic plants containing this chimaeric gene the intron is spliced efficiently, giving rise to GUS enzymatic activity. Mapping of the splice junction indicates the exact removal of the intron. No GUS activity is detected in agrobacteria containing this construct due to the lack of a eukaryotic splicing apparatus in prokaryotes. Early phases after transformation of Arabidopsis cotyledon explants were analysed using this GUS-intron chimaeric gene showing that as early as 36 h after Agrobacterium infection significant GUS activity is detected. In vivo GUS staining of transformed cells clearly shows that quickly proliferating calli expressing GUS activity are formed, mainly at the cut surface. Minor transformation events occur however throughout the whole cotyledon. These data indicate that Agrobacterium-mediated T-DNA transfer to plants is much more efficient than has been judged from experiments where selection is applied immediately. The intron-containing GUS gene can be used as an optimised marker gene in transient and stable transformation experiments.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00260489
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  • 8
    Electronic Resource
    Electronic Resource
    Microprotoplast fusion technique: a new tool for gene transfer between sexually-incongruent plant species (1955)
    Springer
    Euphytica 85 (1955), S. 255-268 
    Details
    ISSN: 1573-5060
    Keywords: microprotoplast fusion ; partial genome transfer ; monosomic additions ; kanamycin resistance ; β-glucuronidase ; gene expression ; potato
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Various aspects of a microprotoplast fusion technique and the strategies followed for intergeneric partial genome transfer (one or a few chromosomes) and alien genes from sexually-incongruent donor species to recipient species are described. The essential requirements of the microprotoplast fusion technique are the induction of micronuclei at high frequencies, as well as the isolation and enrichment of sub-diploid microprotoplasts in donor species, efficient fusion of the donor microprotoplasts with normal recipient protoplasts and stable regeneration of plants from fusion products. The results on the production of microprotoplast hybrid plants between the transformed donor lines of Solanum tuberosum and Nicotiana Plumbaginifolia carrying various genetic markers, and a recipient line of Lycopersicon peruvianum or Nicotiana tabacum, and on the transfer and expression of alien genes (kanamycin resistance, β-glucuronidase) are presented. The data obtained on microprotoplast hybrid plants between S. tuberosum and L. peruvianum showed that many of the hybrids contained one potato chromosome carrying nptII and GUS, and 24 or 48 L. peruvianum chromosomes (monosomic additions), and that they were male-and female-fertile. Various applications of chromosome transfer by this technique, especially for economically-important traits (e.g. disease or stress resistance) from sexually-incompatible wild species, for construction of chromosome-specific DNA libraries through microdissection and microcloning of chromosomes, or by flow-sorting of chromosomes for genome analysis, are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00023954
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  • 9
    Electronic Resource
    Electronic Resource
    Water Vapor Adsorption and Desorption Isotherms of Biologically Active Proteins (1991)
    Springer
    Pharmaceutical research 8 (1991), S. 191-195 
    Details
    ISSN: 1573-904X
    Keywords: urease ; lipase ; α-amylase ; β-glucuronidase ; isolated soy protein ; static/dynamic water adsorption and desorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Using a protein isolated from soy, a dynamic water adsorption method was developed and the data were compared with those obtained from a static gravimetric procedure. Both methods gave comparable results, showing that Type II isotherms with considerable hysteresis were obtained. However, the dynamic procedure was preferred since it provided data rapidly and used significantly less material. Using the dynamic method, water adsorption isotherms at 25°C were also determined for four biologically active proteins: α-amylase, (β-glucuronidase, lipase, and urease. BET (Brunauer, Emmet, and Teller) parameters were calculated and the specific surface areas for the native, biologically active proteins were found to be similar, 238.4 ± 20.2 m2/g. On the other hand, the specific surface area for the denatured soy protein isolate was 144.6 m2/g. Nevertheless, the heat of absorbance for all of the proteins examined was similar, suggesting that they have comparable degrees of hydrophilicity.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1015835902721
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