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  • 1
    ISSN: 1432-2242
    Keywords: Key words Genomic in situ hybridization (GISH) ; Oryza sativa and O. punctata ; Somatic hybrids ; Cell fusion ; Chromosome reduction ; A ; B and C genomes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  A multi-color genomic in situ hybridization (McGISH) method has been developed. Three different rice genomes, A, B and C, involved in rice somatic hybrids were distinguished using three different fluorescent signals. All the rice chromosomes from the different genomes could be identified by different fluorescent colors, and the distribution of each genome in the nucleus was clearly visualized under a fluorescence microscope. The relationship between chromosomal constitution and morphological variations observed in the somatic hybrids, and the utility of McGISH, are discussed based on the results currently obtained.
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  • 2
    ISSN: 1432-2242
    Keywords: Key words Brassica campestris ; Multicolor ; FISH ; Self-incompatibility ; S-glycoprotein (SLG) gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The physical localization of the S-glycoprotein (SLG) locus in the chromosome of Brassica campestris L. ‘pekinensis’ cv ‘Kukai’ was visualized by multi-color fluorescent in situ hybridization (McFISH). ‘Kukai’, which is an F1 hybrid between two parental lines, T-17 and T-18, has two SLG genes from both T-17 and T-18. In this study, a 1.3-kb DNA fragment was amplified from the genomic DNA of T-17 by PCR using a set of primers specific to the class-I SLG. From the genomic DNA of T-18, no DNA fragment was amplified using these primers. In the genomic Southern hybridization, a cloned PCR product hybridized with the genomic DNA of T-17 or F1 but not with that of T-18. The PCR product had a sequence homology of approximately, 85% to another class-I SLG gene, SLG-9. Therefore, the PCR product from T-17 was named SLG-17, as it is thought to be a member of the class-I SLG. Using SLG-17 as the probe, FISH was carried out to visualize the position of the SLG locus. McFISH was also carried out simultaneously using the SLG-17 and SLG-9 genes as probes. The SLG-17 gene was detected as a doublet signal at the interstitial region close to the end of a small chromosome, with the signal site being identical to that of SLG-9. Therefore, it is concluded that the SLG-17 gene is localized at the interstitial region close to the end of the chromosome derived from T-17 in Brassica campestris L. ‘pekinensis’ cv ‘Kukai’.
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  • 3
    ISSN: 1432-2242
    Keywords: Key words Tomato and Solanum lycopersicoides intergeneric hybrid ; Chloroplast DNA ; Nuclear genome ; RFLP ; GISH
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  RFLP (restriction fragment length polymorphism) and GISH (genomic in situ hybridization) analyses were employed to identify the chloroplast and nuclear genomes of the somatic hybrids and progeny between tomato ‘Ohgata zuiko’ and Solanum lycopersicoides (‘LA 2386’). A random distribution of the chloroplast genotype was determined using a cloned 19.6-kb BamHI fragment (Ba1) of tobacco chloroplast DNA. Eight selected hybrids were analyzed for their chromosomal compositions; 4 were tetraploids (2n=48) with an equal number of chromosomes derived from each parent as accurately determined by GISH, and the other 4 were hexaploids, containing an average of two sets of tomato chromosomes and one set from the wild parent. RFLP analysis with six tomato nuclear probes of known chromosomal locations revealed no major variation among the 44 hybrid plants surveyed. However, it also showed the presence of both parent-specific alleles and the loss of some and the presence of a few non-parental alleles, indicating rearrangement and/or recombination of the nuclear DNA. The relevance of the molecular and cytological methods and the potential use of somatic hybrids for plant breeding are demonstrated.
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 99 (1999), S. 391-397 
    ISSN: 1432-2242
    Keywords: Key words Centromeric specific repetitive sequence ; FISH ; Petunia hybrida ; Self-incompatibility ; S-RNase gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  S-RNase has been identified to be an S-allele-specific stylar determinant contributing to the self-incompatibility response in Solanaceae. In order to examine the physical location of the S-RNase gene, multi-color fluorescence in situ hybridization (FISH) using the S B1 -RNase cDNA probe and ribosomal RNA gene (rDNA) probe was performed on an S B1 S B2 heterozygote of Petunia hybrida. The S B1 -RNase gene was detected as a doublet signal close to the centromere of chromosome III. Next, we performed FISH using a large genome probe prepared from a λSB1–311 clone (20 kb) which contains the S B1 -RNase gene and its 3´ flanking region. This probe hybridized to the centromeric regions of all P. hybrida chromosomes. Sequence analysis of the λSB1–311 clone revealed the presence of a repetitive sequence consisting of a novel 666 bp unit sequence. A subclone (pBS-SB1B5) containing this unit sequence also hybridized to all of the centromeric regions, confirming that this unit is the centromeric specific repetitive sequence. These data suggested that the S B1 -RNase gene is located very close to (within a distance of 12 kb from) the centromeric-specific repetitive sequence. Likewise, the pBS-SB1B5 probe hybridized to the centromeric regions of all chromosomes in P. littoralis, another Petunia species. However, the probe did not hybridize to the centromere of the chromosomes from other species in Solanaceae. These results suggested that this centromeric repetitive sequence might be a genus-specific one.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 91 (1995), S. 212-217 
    ISSN: 1432-2242
    Keywords: Oryza glaberrima ; Karyotype analysis ; Ribosomal RNA genes ; Image analysis ; Multicolor in situ hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract African cultivated rice, Oryza glaberrima Steud., was cytologically characterized by using both karyotype analysis and molecular cytology. The somatic chromosomes resemble those of Asian cultivated rice, Oryza sauva L., in general morphology, although some minor differences were noted. Multicolor fluorescence in situ hybridization (McFISH) with chromosomes detected one 45s (17s-5.8s-25s) ribosomal RNA gene locus (45s rDNA) and one 5s ribosomal RNA gene locus (5s rDNA) in the chromosome complement. The 45s rDNA and 5s rDNA loci were physically mapped to the distal end of the short arm of chromosome 9 and to the proximal region of the short arm of chromosome 11 respectively, as in O. sativa. Based on the cytological observations and the physical map of the rDNA loci, the chromosomal organization of O.glaberrima and O. sativa seems to be very similar.
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 95 (1997), S. 1239-1245 
    ISSN: 1432-2242
    Keywords: Key words Genomic in situ hybridisation (GISH) ; Genus Oryza ; O. latifolia (CCDD) ; O. minuta (BBCC) ; Rice D genome
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The 24 rice D-genome chromosomes were identified among the 48 chromosomes of O. latifolia, which comprise the C- and D-genomes, using genomic in situ hybridisation (GISH). The B-genome chromosomes were also discriminated from the C-genome chromosomes in O. minuta (BBCC) by GISH. A comparison of the differences in the fluorescence intensity between the C and D genomes within O. latifolia (CCDD), and between the B and C genomes within O. minuta, indicated that the overall nucleotide-sequence homology between the B and C genomes is less than that between the C and D genomes. The origin of the D genome and the phylogenetic relationship of the D genome among the rice genomes are discussed, based on the results obtained.
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  • 7
    ISSN: 1432-2242
    Keywords: Key words Brassica rapa ; Brassica nigra ; Brassica oleracea ; Quantitative chromosome map ; Idiogram ; FISH ; 45s rDNA ; Systematic mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Chromosomes of the three diploid Brassica species, B. rapa (AA), B. nigra (BB) and B. oleracea (CC), were identified based on their morphological characteristics, especially on the condensation pattern appearing at the somatic pro-metaphase stage. The morphological features of the pro-metaphase chromosomes of the three Brassica spp. were quantified by imaging methods using chromosome image analyzing system II (CHIAS 2). As a result, quantitative chromosome maps or idiograms of the three diploid Brassica spp. were developed. The fluorescence in situ hybridization (FISH) method revealed the location of 45s rDNA (the 26s-5.8s-18s ribosomal RNA gene cluster) on the chromosomes involved. The number of 45s rDNA loci in the B. rapa, B. nigra and B. oleracea are five, three and two, respectively. The loci detected were systematically mapped on the idiograms of the three Brassica spp.
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  • 8
    ISSN: 1432-2242
    Keywords: Key words Map-based cloning ; Positional cloning ; Fluorescence in situ hybridization ; Disease resistance ; Orseolia oryzae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Ten yeast artificial chromosomes (YACs) spanning the Gm2 locus have been isolated by screening high-density filters containing a total of approximately 7000 YAC (representing six genome equivalents) clones derived from a japonica rice, Nipponbare. The screening was done with five RFLP markers flanking a gall midge resistance gene, Gm2, which was previously mapped onto chromosome 4 of rice. This gene confers resistance to biotype 1 and 2 of gall midge (Orseolia oryzae), a major insect pest of rice in South and Southeast Asia. The RFLP markers RG214, RG329 and F8 hybridized with YAC Y2165. Two overlapping YAC clones (Y5212 and Y2165) were identified by Southern hybridization, with Gm2-flanking RFLP markers, and their inserts isolated. The purified YACs and RFLP markers flanking Gm2 were labeled and physically mapped by the fluorescence in situ hybridization (FISH) technique. All of them mapped to the long arm of chromosome 4 of the resistant variety of rice, ‘Phalguna’, confirming the previous RFLP mapping data.
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  • 9
    ISSN: 1617-4623
    Keywords: Key words Bacterial Artificial Chromosome (BAC) library  ;  Contig  ;  Centromere  ;  Rice (Oryza sativa)  ;  Rice blast resistance gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We constructed a rice Bacterial Artificial Chromosome (BAC) library from green leaf protoplasts of the cultivar Shimokita harboring the rice blast resistance gene Pi-ta. The average insert size of 155 kb and the library size of seven genome equivalents make it one of the most comprehensive BAC libraries available, and larger than many plant YAC libraries. The library clones were plated on seven high density membranes of microplate size, enabling efficient colony identification in colony hybridization experiments. Seven percent of clones carried chloroplast DNA. By probing with markers close to the blast resistance genes Pi-ta 2 (closely linked to Pi-ta) and Pi-b, respectively located in the centromeric region of chromosome 12 and near the telomeric end of chromosome 2, on average 2.2 ± 1.3 and 8.0 ± 2.6 BAC clones/marker were isolated. Differences in chromosomal structures may contribute to this wide variation in yield. A contig of about 800 kb, consisting of 19 clones, was constructed in the Pi-ta 2 region. This region had a high frequency of repetitive sequences. To circumvent this difficulty, we devised a “two-step walking” method. The contig spanned a 300 kb region between markers located at 0 cM and 0.3 cM from Pi-ta 2 . The ratio of physical to genetic distances (〉 1,000 kb/cM) was more than three times larger than the average of rice (300 kb/cM). The low recombination rate and high frequency of repetitive sequences may also be related to the near centromeric character of this region. Fluorescent in situ hybridization (FISH) with a BAC clone from the Pi-b region yielded very clear signals on the long arm of chromosome 2, while a clone from the Pi-ta 2 region showed various cross-hybridizing signals near the centromeric regions of all chromosomes.
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Rock mechanics and rock engineering 30 (1997), S. 113-118 
    ISSN: 1434-453X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Architecture, Civil Engineering, Surveying , Geosciences
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