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  • 1
    Electronic Resource
    Electronic Resource
    Molecular-cytogenetic characterization of a higher plant centromere/kinetochore complex (1996)
    Springer
    Theoretical and applied genetics 93 (1996), S. 477-484 
    Details
    ISSN: 1432-2242
    Keywords: Key words Microdissection of plant centromeres ; Subtractive hybridization ; Fluorescence in situ hybridization ; Indirect immunofluorescence ; Immunoadsorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The centromeric region of a telocentric field bean chromosome that resulted from centric fission of the metacentric satellite chromosome was microdissected. The DNA of this region was amplified and biotinylated by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR)/linker-adapter PCR. After fluorescence in situ hybridization (FISH) the entire chromosome complement of Vicia faba was labelled by these probes except for the nucleolus organizing region (NOR) and the interstitial heterochromatin, the chromosomes of V. sativa and V. narbonensis were only slightly labelled by the same probes. Dense uniform labelling was also observed when a probe amplified from a clearly delimited microdissected centromeric region of a mutant of Tradescantia paludosa was hybridized to T. paludosa chromosomes. Even after six cycles of subtractive hybridization between DNA fragments amplified from centromeric and acentric regions no sequences specifically located at the field bean centromeres were found among the remaining DNA. A mouse antiserum was produced which detected nuclear proteins of 33 kDa and 68 kDa; these were predominantly located at V. faba kinetochores during mitotic metaphase. DNA amplified from the chromatin fraction adsorbed by this serum out of the sonicated total mitotic chromatin also did not cause specific labelling of primary constrictions. From these results we conclude: (1) either centromere-specific DNA sequences are not very conserved among higher plants and are – at least in species with large genomes – intermingled with complex dispersed repetitive sequences that prevent the purification of the former, or (2) (some of) the dispersed repeats themselves specify the primary constrictions by stereophysical parameters rather than by their base sequence.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001220050305
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  • 2
    Electronic Resource
    Electronic Resource
    Molecular-cytogenetic characterization of a higher plant centromere/kinetochore complex (1996)
    Springer
    Theoretical and applied genetics 93 (1996), S. 477-484 
    Details
    ISSN: 1432-2242
    Keywords: Microdissection of plant centromeres ; Subtractive hybridization ; Fluorescence in situ hybridization ; Indirect immunofluorescence ; Immunoadsorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The centromeric region of a telocentric field bean chromosome that resulted from centric fission of the metacentric satellite chromosome was microdissected. The DNA of this region was amplified and biotinylated by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR)/linker-adapter PCR. After fluorescence in situ hybridization (FISH) the entire chromosome complement of Vicia faba was labelled by these probes except for the nucleolus organizing region (NOR) and the interstitial heterochromatin, the chromosomes of V. sativa and V. narbonensis were only slightly labelled by the same probes. Dense uniform labelling was also observed when a probe amplified from a clearly delimited microdissected centromeric region of a mutant of Tradescantia paludosa was hybridized to T. paludosa chromosomes. Even after six cycles of subtractive hybridization between DNA fragments amplified from centromeric and acentric regions no sequences specifically located at the field bean centromeres were found among the remaining DNA. A mouse antiserum was produced which detected nuclear proteins of 33 kDa and 68 kDa; these were predominantly located at V. faba kinetochores during mitotic metaphase. DNA amplified from the chromatin fraction adsorbed by this serum out of the sonicated total mitotic chromatin also did not cause specific labelling of primary constrictions. From these results we conclude: (1) either centromere-specific DNA sequences are not very conserved among higher plants and are — at least in species with large genomes — intermingled with complex dispersed repetitive sequences that prevent the purification of the former, or (2) (some of) the dispersed repeats themselves specify the primary constrictions by stereophysical parameters rather than by their base sequence.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00417938
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  • 3
    Electronic Resource
    Electronic Resource
    The acetylation patterns of histones H3 and H4 along Vicia faba chromosomes are different (1998)
    Springer
    Chromosome research 6 (1998), S. 59-63 
    Details
    ISSN: 1573-6849
    Keywords: deacetylase inhibition ; field bean ; heterochromatin ; histone acetylation patterns ; indirect immunofluorescence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The acetylation pattern of H3 was studied on field bean chromosomes by means of indirect immunofluorescence using polyclonal antibodies recognizing H3 isoforms acetylated at lysine positions 9/18, 14 and 23. H3 was found to be hypoacetylated at lysine residues 9/18 and 14 within the heterochromatic regions composed of tandem repetitive Fok-I elements. Hyperacetylation of these residues was observed at the nucleolar organizing region (NOR) and in heterochromatic regions composed of repeats other than Fok-I elements. In contrast, H4 was underacetylated (H4.Ac5, 8, 12) or uniformly acetylated (H4.Ac16) at all heterochromatic regions, and acetylated above the average at all four lysines only within the NOR. Acetylation of lysine-23 of H3 was uniform, except for the NOR that showed no fluorescence. Inhibition of deacetylase during and after replication of heterochromatin by trichostatin A had no influence on the acetylation status of H3 but mediated an increase in acetylation of lysines 5, 12 and 16 of H4 above the average in the field bean heterochromatin. Thus, the chromosomal acetylation patterns of H4 and H3 of this species revealed common and divergent features. Whereas the acetylation level of H4 correlates well with the potential transcriptional activity and inversely with the time of replication of defined chromatin domains of Vicia faba, this is not generally true for H3.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009222609581
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