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  • 1
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Physiologia plantarum 70 (1987), S. 0 
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A 5% increase of Ca2+ content of the incubation medium for cultured peanut (Arachis hypogaea L.) cells caused a rise of peroxidase (EC 1.11.1.7) activity in the medium, which could be abolished by the addition of the chelator EGTA [eth-yleneglycol-bis-(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid]. However, the determination of in vivo peroxidase synthesis in these cells showed that Ca2+ had a direct effect on the biosynthesis rather than on transport alone. This concept was re-enforced by the lack of effect by the ionophore A23187 on the transport. The Ca2+ content was 2 and 5 mol (mol protein−1) for the cationic and anionic peanut peroxidase, respectively. The latter is different from the value reported for the anionic peroxidase from horseradish.
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  • 2
    ISSN: 1432-203X
    Keywords: particle bombardment ; Hordeum vulgare L. ; promoter screening ; transient expression ; gus ; cereal transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The cauliflower mosaic virus 35S (35S) and the enhanced 35S (E35S) promoters fused with maize alcohol dehydrogenase (Adh1) intron1 or maize shrunken locus (sh1) intronl along with maize Adh1 and rice actin (Act1) promoters fused to their respective first introns were tested for transient expression of the E.coli β-glucuronidase (gus) reporter gene in cultured barley (Hordeum vulgare L) cells. The plasmids, carrying the respective promoterintron combinations to drive the gus fused to nopaline synthase (nos) terminator, were introduced into cultured barley cells using a particle gun. The rice Act1 promoter with its first intron gave the highest expression of all promoter intron combinations studied. This was followed by the E35S promoter and no significant differences were observed between the other two promoters tested. The rice actin promoter is now being used to drive selectable marker genes to obtain stably transformed cereal cells.
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  • 3
    ISSN: 1432-203X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An efficient genetic transformation protocol has been developed for strawberry cv. Redcoat using Agrobacterium tumefadens. The protocol relies on a high frequency (84%) shoot regeneration system from leaf disks. The leaf disks were inoculated with a non-oncogenic Agrobacterium tumefadens strain MP90 carrying a binary vector plasmid pBI121 which contains a chimeric nopaline synthase (NOS) promoter driven neomycin phosphotransferase (NPT II) gene and a cauliflower mosaic virus 35S (CaMV35S) promoter driven, ß-glucuronidase (GUS) marker gene. The inoculated leaf disks, pre-cultured for 10 days on non-selective shoot regeneration medium, formed light green meristematic regions on selection medium containing 50 μg/ml kanamycin. These meristematic regions developed into transformed shoots at a frequency of 6.5% on a second selection medium containing 25 μg/ml kanamycin. The selected shoots were multiplied on shoot proliferation medium in the presence of kanamycin. All such shoots were resistant to kanamycin and expressed varying levels of NPT II and GUS enzyme activity. Histochemical assays for GUS activity indicated that the 35S promoter was highly active in meristematic cells of shoot and root apices. Molecular analysis of each transgenic clone confirmed the integration of both marker genes into the strawberry genome. Leaf disks prepared from transformed plants, when put through the second selection cycle on kanamycin, formed callus and exhibited GUS activity. The rooted transformed plants were grown in a greenhouse for further characterization. The protocol may be useful for improvement of strawberry through gene manipulations.
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  • 4
    ISSN: 1432-203X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transformed calli and shoots of strawberry (Fragaria × ananassa Duch.) cv. Redcoat were obtained using Agrobacterium tumefaciens carrying plasmid pB1121. Inoculated leaf explants produced transgenic calli at a frequency of 3% on selection medium containing 50 μg/ml kanamycin. Twenty per cent of selected caili regenerated, giving rise to transgenic shoots. All transgenic calli and shoots expressed substantial amounts of GUS and NPT-II activity. The Southern blot analysis confirmed the insertion of both marker genes into the strawberry genome as single and multiple copy inserts. The transgenic shoots elongated on rooting medium in the presence of 25 μg/ml kanamycin, but exhibited reduced rooting ability.
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  • 5
    ISSN: 1573-5028
    Keywords: alternative splicing ; starch biosynthesis ; starch-branching enzyme ; transit peptide ; wheat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A wheat gene, denoted Sbe1, encoding a type I starch-branching enzyme (SBEI) was isolated from a genomic library and shown to comprise 14 exons distributed over a 5.7 kb DNA region. Analyses of kernel RNA by 5′ rapid amplification of cDNA ends (5′-RACE) and reverse transcription-polymerase chain reaction (RT-PCR) demonstrated a considerable sequence variation at the 5′ ends of SBEI gene transcripts. DNA sequence alignments between the 5′-RACE products and the Sbe1 genomic DNA indicated that the first two exons and first intron were differentially processed to generate three classes of the mature transcript. One form of the SBEI gene transcript in 12-day old kernels contained the exon I+II+III combination at the 5′ end, whereas other forms differed by inclusion of intron 1 or exclusion of exon II sequences. RT-PCR analysis of Sbe1-uidA::nptII chimeric mRNA produced in transgenic wheat cultured cells confirmed that the isolated Sbe1 was able to produce all three forms of SBEI gene transcripts by alternative splicing of the primary mRNA. The variants of processed Sbe1 mRNA were potentially translated into N-terminal variants of the SBEI precursor with different transit peptide sequences.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell, tissue and organ culture 60 (2000), S. 69-73 
    ISSN: 1573-5044
    Keywords: cereals ; tissue culture ; Triticum durum ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Fertile, green plants were regenerated from immature inflorescence explants from each of four Canadian wheat cultivars. The cultivars were representative of four classes of Canadian wheat. Explants from immature inflorescences of three size ranges were cultured on two types of media: MSI/MSR, which contains 1650 mg l-1 NH4NO3and sucrose as a carbon source, and BII/BIR, which contains 250 mg l-1 NH4NO3and maltose as a carbon source. Regeneration from all cultivars was significantly better on BII/BIR media than on MSI/MSR media. On BII/BIR media, `AC Karma', `Plenty', and `Fielder' gave the highest number of shoots per 10 explants, where the explants were derived from immature inflorescences 5.1 to 10.0 mm in length. 'Columbus' did not regenerate on MSI/MSR medium, and regenerated poorly on BII/BIR medium. Differences were found between cultivars with regard to the number of regenerant plants produced with the best treatments: `Plenty' produced 16.1 shoots per 10 explants, `AC Karma' 12.4, `Fielder' 6.4, and `Columbus' 2.2.
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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 29 (1995), S. 647-662 
    ISSN: 1573-5028
    Keywords: gene expression ; peroxidase ; powdery mildew ; splicing ; Triticum aestivum L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A PCR-based screening approach was used to isolate genomic clones from wheat encoding peroxidase isozymes. Three complete genes (pox1, pox2 and pox4) and one truncated gene (pox3) were characterized. The nucleotide sequences predicted mature proteins of 31 kDa, in which all the highly conserved motifs of secreted plant peroxidases were preserved. The coding regions showed 73–83% DNA sequence identity, with the highest level of similarity noted for the tandemly oriented pox2 and pox3. Expression of respective pox genes in various tissues of wheat was assessed by the RT-PCR technique, which showed that all four genes are active. The primary pox1 mRNA was spliced to remove three introns, whereas processing of the other pox transcripts involved only two intervening sequences. Splicing occurred at consensus GU/AG splice sites except for the first introns of pox1, pox2 and pox4 transcripts, where processing took place at unusual GC donor sites. The RNA analysis suggested that the pox1, pox2 and pox4 genes are predominantly expressed in roots. Lower levels of expression were found for pox4 and pox3 in leaves. Infection of wheat by the powdery mildew fungus selectively induced expression of pox2 in leaves.
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  • 8
    ISSN: 1573-9368
    Keywords: β-GUS ; embryogenesis reinduction ; kanamycin ; NPTII ; Piceaglauca ; selection ; somatic embryos ; somaticseedlings ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transformed white spruce cultures containing immature Stage I somatic embryos, were developed after particle bombardment of somatic embryos with pBI 426, carrying an expression cassette with a gus::nptII fusion gene. These Stage I cultures did not show tolerance to kanamycin concentrations above 3 to 5 mg l−1, although assays for GUS and NPTII showed that functional enzymes were present in the transgenic tissue. Embryonic liquid suspension cultures were initiated from this transformed tissue. After treatment on agar-solidified maturation medium with 48 μm (±)-ABA high numbers of Stage III (cotyledonary) somatic embryos were produced. When subjected to an embryogenesis re-induction process with 2,4-D and BA, these Stage III embryos produced a new generation of Stage I embryogenic tissues which could tolerate 5--10 mg l−1 kanamycin. Stage III somatic embryos could alternatively be placed onto germination medium for the development of somatic seedlings. When germinated in the presence of 20 mg l−1 kanamycin, 77% of inoculants were resistant. The stability of integration of the gus::nptII fusion gene in the genome of white spruce Stage III somatic embryos and somatic seedlings was confirmed through Southern blot analysis
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  • 9
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Electrophoresis 9 (1988), S. 213-216 
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: By inserting nitrocellulose strips into agarose gels alongside the electrophoresed lanes and passing an electric current perpendicularly in the direction of the strips, highly efficient transfer of DNA bands onto the membrane in the form of concentrated dots is achieved. DNA detection limits by this technique are enhanced, at least three times as visualized by ethidium bromide fluorescence and at least twice more by radiolabeling.
    Additional Material: 2 Ill.
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  • 10
    Publication Date: 2010-04-14
    Print ISSN: 0021-8561
    Electronic ISSN: 1520-5118
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition , Process Engineering, Biotechnology, Nutrition Technology
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