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  • 1
    Electronic Resource
    Electronic Resource
    Preparation and flow cytometric analysis of metaphase chromosomes of tomato (1991)
    Springer
    Theoretical and applied genetics 82 (1991), S. 101-111 
    Details
    ISSN: 1432-2242
    Keywords: Tomato chromosome suspension ; Flow cytometry ; Univariate analysis ; Bivariate analysis ; Chromosome sorting
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A procedure for the preparation of tomato chromosome suspensions suitable for flow cytometric analysis is described. Rapidly growing cell suspension cultures of Lycopersicon esculentum cv VFNT cherry and L. pennellii LA716 were treated with colchicine to enrich for metaphase chromosomes. Metaphase indices between 20 and 35% were routinely obtained when cultures were exposed to 0.1% colchicine for 15–18 h after 2 days of subculture. Mitotic cells were isolated by brief treatment with cell wall digesting enzymes in a medium with low osmolarity (∼325 mOsm/kg of H52O). The low osmolarity medium was needed to avoid the chromosome clumping and decondensation seen in standard media. Suspensions of intact chromosomes were prepared by lysing swollen protoplasts in various buffers (MgSO4, polyamines, hexylene glycol, or KCl-propidium iodide) similar in contents to the buffers used to isolate mammalian chromosomes. For univariate flow cytometric analysis, chromosome suspensions were stained with a fluorescent DNA-binding stain (propidium iodide, Hoechst 33258, mithramycin, or chromomycin A3) and analyzed using an EPICS flow cytometer (Profile Analyzer or 753). Peaks for the chromosomes, chromatids, clumps of chromosomes, nuclei, and fluorescent debris were seen on a histogram of log of fluorescence intensity, and were confirmed by microscopic examination of the objects collected by flow-sorting. Chromosome suspensions prepared in MgSO4 buffer have the highest frequency of intact chromosomes and the least fluorescent cellular debris. Peaks similar to theoretical univariate flow karyotypes of tomato chromosomes were seen on the observed univariate flow karyotypes, but were not as well resolved. Bivariate flow analysis of tomato chromosome suspension using double-stain combination, Hoechst 33258 and chromomycin A3, and two laser beams showed better resolution of some chromosomes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00231283
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  • 2
    Electronic Resource
    Electronic Resource
    DNA content of wheat monosomics at interphase estimated by flow cytometry (1997)
    Springer
    Theoretical and applied genetics 95 (1997), S. 1300-1304 
    Details
    ISSN: 1432-2242
    Keywords: Key words Flow cytometry ; Triticum aestivum ; DNA analysis ; Monosomics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Two complete, independently maintained sets of 21 monosomic wheat lines derived from cv. ‘Chinese Spring’ were analyzed for their DNA content at the G1 stage with flow cytometry. The DNA content of individual chromosomes was estimated by subtracting the value of a monosomic line from that of euploid wheat. Our data show that the estimated 2C DNA of individual wheat chromosomes in 21 monosomics at the G1 stage ranges from about 0.58 pg in chromosome 1D to approximately 1.12 pg in chromosome 3A. The A genome (2C=6.15 pg) seems to contain more DNA than the B (2C=6.09 pg) and D (2C=5.05 pg) genomes. Analysis of variance showed significant differences (α=0.01) in DNA content both among homoeologous groups and among genomes. Our estimates of interphase DNA content of wheat chromosomes from monosomic lines were poorly correlated to the chromosome sizes at metaphase (r=0.622, P≤0.01). This poor correlation might be due to differential coiling among chromosomes during cell division, possible bias of fluorochrome binding to heterochromatin, or heterogeneity among monosomic lines. Finally, flow cytometry may aid but cannot replace cytological checks in aneuploid maintenance.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001220050696
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  • 3
    Electronic Resource
    Electronic Resource
    Relationships between nuclear DNA content and seed and leaf size in soybean (1998)
    Springer
    Theoretical and applied genetics 96 (1998), S. 1064-1068 
    Details
    ISSN: 1432-2242
    Keywords: Key words Glycine max ; Genome size ; Seed size ; Leaf size ; Flow cytometry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  A correlation between genome size and agronomically important traits has been observed in many plant species. The goal of the present research was to determine the relationship between genome size, seed size, and leaf width and length in soybean [Glycine max (L.) Merr.] Twelve soybean strains, representing three distinct seed size groups, were analyzed. Flow cytometry was used to estimate their 2C nuclear DNA contents. Data on seed size and leaf size of the 12 strains were obtained from 1994 and 1995 field experiments. Variation of 2C nuclear DNA among the 12 soybean strains was 4.6%, ranging from 2.37 pg for a small-seed strain to 2.48 pg for a large-seed strain. Strain seed size was positively associated with leaf width (r=0.92) and leaf length (r=0.93). Genome size was highly correlated with seed size (r=0.97), leaf width (r=0.90) , and leaf length (r=0.93). The results of our study indicate that there is a significant correlation between genome size and leaf and seed size in soybean. It is possible that selection for greater seed size either leads to, or results from, greater genome size. If so, this relationship might be worth exploring at a more fundamental level.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001220050840
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  • 4
    Electronic Resource
    Electronic Resource
    Isolating individual wheat (Triticum aestivum) chromosome arms by flow cytometric analysis of ditelosomic lines (1999)
    Springer
    Theoretical and applied genetics 98 (1999), S. 1248-1252 
    Details
    ISSN: 1432-2242
    Keywords: Key words Wheat ; Chromosome sorting ; Chromosome isolation ; Aneuploids ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  We are reporting the successful isolation of wheat chromosome arm 1DS by flow cytometry. A chromosome suspension was prepared for the 1DS ditelosomic line and the normal ‘Chinese Spring’ (CS) by chopping 2-day-old root tip meristems, synchronized by hydroxyurea, in HEPES-magnesium sulfate buffer containing propidium iodide. Chromosomes were analyzed and sorted with a FACS Vantage flow cytometer and cell sorter. An extra peak was observed in the flow karyotype of the ditelosomic line that was absent in ‘CS’. The estimated size of chromosomes from the extra peak matched with the expected size of chromosome 1DS. Chromosomes from the putative 1DS peak were analyzed by both fluorescent microscopy and N-banding analysis. A total of 571 chromosomes from two separate experiments were analyzed, and all were observed to be telosomics except for 2 which were broken. About 82% of these telosomics showed the diagnostic N-band of 1DS, the remaining were unbanded and are probably also 1DS. This strategy can also be used to sort other wheat arms.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001220051190
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  • 5
    Electronic Resource
    Electronic Resource
    Chromosome 2-specific DNA clones from flow-sorted chromosomes of tomato (1994)
    Springer
    Molecular genetics and genomics 242 (1994), S. 551-558 
    Details
    ISSN: 1617-4623
    Keywords: Lycopersicon pennellii ; Flow cytometry ; Degenerate oligonucleotide primer (DOP) ; Chromosome-specific library ; RFLP mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We obtained DNA clones specific to tomato chromosome 2 from a small number of chromosomes collected by flow sorting. Suspensions of metaphase chromosomes were prepared from 3-month-old tomato cell cultures of Lycopersicon pennellii. Isolated chromosomes stained with chromomycin A3 and Hoechst 33258 were analyzed on an EPICS 753 flow cytometer using a UV laser to excite Hoechst fluorescence and a 458 nm laser to excite chromomycin A3 fluorescence. Chromosomes from well-resolved peaks on a bivariate flow karyotype were sorted directly onto membrane filters for spot-blot analysis. The filters were processed and hybridized with chromosome-specific repetitive DNA probes. In this way tomato chromosome 1 and chromosome 2 were assigned to peaks in the bivariate flow karyotypes. One thousand copies of the putative chromosome 2 were flow-sorted directly into microfuge tubes. DNA specific to chromosome 2 was amplified by a polymerase chain reaction (PCR) technique using universal 22mer degenerate oligonucleotide primers (DOP) sequences. DOP-PCR yields a smear of fragments of various sizes from 250 to 1600 bp. Amplified products were cloned into the Bluescript plasmid vector. Approximately 11 of the clones contained sequences with highly repetitive elements, and 85% contained only low-copy-number sequences. Eleven clones containing low-copy-number sequences that detect restriction fragment length polymorphisms were placed on the molecular linkage map of tomato. All showed linkage to chromosome 2.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00285278
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  • 6
    Electronic Resource
    Electronic Resource
    Ploidy Level Determinations in Buffalograss Clones and Populations (1998)
    Springer
    Crop science 38 (1998), S. 478-482 
    Details
    ISSN: 1435-0653
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Buchloë dactyloides (Nutt.) Engelm.] is the subject of breeding programs for turfgrass use because of its drought and heat tolerance, low growing habit, and low management input. Three ploidy levels are known to exist (x=n=10), but plants at each level are nearly morphologically indistinguishable. In this paper, flow cytometry was evaluated as a means of determining ploidy levels of 31 buffalograss clones and 34 seeded populations. Six clones were analyzed for nuclear DNA base-pair composition by flow cytometry. Based on DNA contents, buffalograss clones can be grouped into four sets corresponding to chromosome number or ploidy level. Mean DNA contents were 0.93, 1.80, 2.15, and 2.63 pg DNA/nucleus for diploid, tetraploid, pentaploid, and hexaploids, respectively. Chromosome numbers explained most of the DNA content variation as shown by the strong linear relationship between DNA content and chromosome number. Cultivar 315 is a pentaploid and is the first report of a pentaploid among buffalograss. Seeded populations analyzed include those exclusively tetraploid, mixture of tetraploid through hexaploid, and those exclusively hexaploid. Aneuploids between pentaploid and hexaploid levels were noted. No differences were observed between ploidy levels or genders with regard to base-pair composition.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/
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  • 7
    Electronic Resource
    Electronic Resource
    Nuclear DNA Content of Perennial Grasses of the Triticeae (1999)
    Springer
    Crop science 39 (1999), S. 661-667 
    Details
    ISSN: 1435-0653
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Agropyron (PP) 13.9 pg, Pseudoroegneria (StSt) 8.8 pg, Hordeum (HH) 9.5 pg, Psathyrostachys (NsNs) 16.7 pg, and Thinopyrum genomes (EbEb) 14.9 pg and (EeEe) 12.0 pg. The YY genome in Elymus was determined by difference to be 9.3 pg. The unknown or XmXm genome or genomes in Leymus could have DNA contents that range from 2.7 to 7.7 pg/2C. There were significant differences in DNA content of species with similar diploid genomes. There were also significant differences in nuclear DNA content of the polyploid species of the Triticeae were similar to the expected DNA contents on the basis of previous genomic classifications. However, in some allopolyploid genera such as Thinopyrum and Pascopyrum, the nuclear DNA content of some species was less than expected on the basis of summation of the DNA of constituent genomes. The results indicate that gain or loss of nuclear DNA has occurred during the evolution of the perennial Triticeae and was probably a part of speciation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/
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  • 8
    Electronic Resource
    Electronic Resource
    Nuclear DNA Content of Thirteen Turfgrass Species by Flow Cytometry (1999)
    Springer
    Crop science 39 (1999), S. 1518-1521 
    Details
    ISSN: 1435-0653
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/
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