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  • 1998  (67)
  • 1
    ISSN: 1432-1890
    Keywords: Key words DNA polymorphism ; Ectomycorrhizal fungi ; Genetic diversity ; Pisolithus tinctorius ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Twenty Pisolithus tinctorius isolates from different geographic locations and different hosts were characterized by the random amplified polymorphic DNA technique. Thirteen arbitrary primers generated 87 DNA fragments, all of them polymorphic. These data were used to calculate genetic distances among the isolates. The pairwise genetic distances ranged from 1 to 100%, with an average of 58.7%. Cluster analysis based on the amplified fragments grouped the isolates according to their host and geographical origins. Group I contained isolates collected in Brazil and group II those collected in the Northern Hemisphere. In addition to the diversity seen at the molecular level, the isolates also showed host specificity. Greenhouse experiments demonstrated that isolates from the Northern Hemisphere colonized mainly Pinus whereas isolates from Brazil colonized only Eucalyptus. The molecular data suggest that the Pisolithus tinctorius isolates analyzed belong to two distinct groups. The data also suggest new guidelines for future investigations on the taxonomy and systematic of this important fungus species. Furthermore, these results support future experiments aimed at the selection and development of improved isolates of P. tinctorius.
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology reporter 16 (1998), S. 139-139 
    ISSN: 1572-9818
    Keywords: competition ; DNA mixture ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three amplification protocols were analyzed for error rate and generation of polymorphisms during RAPD analysis. Using a set of 240 primers, the protocols detected similar frequencies of polymorphisms in two inbred sugar beet lines. The error rate was investigated by including a 1:1 mixture of DNA from the two lines in all analyses. Similar error rates, approximately 18%, were detected by the three protocols. Thus, altered amplification conditions did not substantially affect the error rate during RAPD analysis. For each of the three possible pairs of protocols, a positive correlation was obtained for primer and number of polymorphisms. Thus, a set of highly polymorphic RAPD primers can be used effectively, without prior screening, to detect polymorphisms for each protocol.
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  • 3
    ISSN: 1572-9818
    Keywords: amplified fragment length polymorphism ; cocoa ; RAPD ; woody plant
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Accurate identification of parental plants and their hybrids is essential for an effective breeding programme. Traditional classification of cocoa varieties relies on the characterisation of agricultural traits at plant maturity. A rapid and reliable method is described, based on genotypic analysis. An efficient DNA isolation procedure was developed, yielding unsheared DNA of high purity. Two genetic fingerprinting techniques, RAPD and AFLP™, were evaluated for their suitability in distinguishing cocoa varieties. RAPD analysis was unsatisfactory due to the low frequency of polymorphisms and poor reproducibility. AFLP™ was reliable in distinguishing phenotypically identical, known varieties of cocoa. Importantly, AFLP™ also revealed intra- and inter-varietal variation. Abbreviations: AFLP™, amplified fragment length polymorphism; APS, ammonium persulphate; CTAB, hexadecyltrimethylammonium bromide; DEB, DNA extraction buffer; f.wt., fresh weight; NEB, nuclei extraction buffer; PMSF, phenylmethanesulphonyl fluoride; RAPD, random amplified polymorphic DNA; T4 PNK, Bacteriophage T4 polynucleotide kinase; Taq, Thermus aquaticus; TBE, tris-borate-EDTA; TEMED, NNN′N′ tetramethylethylenediamine.
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  • 4
    Electronic Resource
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    Springer
    Plant molecular biology reporter 16 (1998), S. 91-91 
    ISSN: 1572-9818
    Keywords: Amaranthus ; DNA fingerprinting ; PCR ; polysaccharides ; RAPD ; total DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A simple, efficient and reliable method is described for isolation of total DNA from young leaves of Amaranthus species. This procedure yields a high amount (600–800 µg DNA/g fresh leaf tissue) of good quality DNA free from contaminating proteins, polysaccharides, and coloured pigments. The DNA is suitable for digestion with several restriction endonucleases, preparation of Southern blots, and PCR amplification. The DNA has been successfully used for generating DNA fingerprint profiles and RAPD banding patterns in two species of Amaranthus. The procedure is suitable for processing of a large number of samples simultaneously.
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  • 5
    ISSN: 1573-0832
    Keywords: armadillo ; Paracoccidioides brasiliensis ; PCR ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Sixty-three Paracoccidioides brasiliensis isolates obtained from three nine-banded armadillos ( Dasypus novemcinctus), one Amazonian armadillo's and 19 clinical isolates were compared by random amplified polymorphic DNA analysis with the primer OPG-19. The isolates were divided into three major clusters, I, II and III. Coincidences between human and armadillo isolates were observed in clusters I and II. Cluster III consisted only of armadillos' isolates. The results suggested that (I) humans may acquire P. brasiliensis infection by contact with armadillo's environment, (II) there may be P. brasiliensis genotypes peculiar to the animal, and (III) individual armadillos may be infected with P. brasiliensis cells with different genotypes.
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  • 6
    ISSN: 1573-0832
    Keywords: Candida ; identification ; PCR ; phylogeny ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Fast and reliable identification of different species of the genus Candida is important to define adequate therapeutic decisions, because the different species have highly variable susceptibilities to antifungal drugs; azoles and amphothericin B. Accurate statistical records on case history and epidemiological studies also depend on effective identification. To address this problem we established a RAPD method that enabled direct identification of five very common species of Candida. Initially, reference band patterns were established for C. albicans, C. tropicalis, C. parapsilosis, C. glabrata and C. krusei. One of the primers, M2, showed remarkably conserved intra-specific patterns of approximately 10 bands each, ranging in size from 2.0 to 0.1 kb. These patterns were significantly different and species-specific. Few bands were conserved between different species of Candida, which was assumed to be consistent with their phylogenetic relatedness. In addition, band patterns were constant and reproducible and DNA isolated from single colonies yielded sufficient DNA for identification. The reference band patterns were then used, in blind experiments, to identify species of Candida in 50 randomly chosen samples, including clinical isolates and ATCC strains. RAPD results were 100% consistent with results obtained by conventional diagnostic methods and were achieved in one day instead of several days taken by conventional methods. Because ideal identification methods should be consistent with phylogeny and taxonomy we tested whether RAPD could be used to calculate genetic distances. Comparison of RAPD phylogenetic trees with 18S rRNA trees showed significant differences in tree topologies which indicated that RAPD data could not accurately measure the relative distances between different species. Also, computer simulations of RAPD random patterns were used to test whether the observed degree of RAPD band pattern similarities could occur at random. These simulations suggested that the level of inter-specific band pattern similarities observed in our data could be obtained at random, while intra-specific pattern similarities could not. RAPD would be helpful to discriminate between isolates but not to quantitate the differences. We suggest that the inaccurate estimate of genetic distances from RAPD is a general limitation of the technique and not a specific problem of our identification method. Because of the repetitive character of the target sequences, genetic distances calculated from RAPD could be affected by paralogy, namely, recombination and duplication events not parallel with speciation events.
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  • 7
    ISSN: 1432-203X
    Keywords: Key words Somatic hybridization ; RAPD ; Citrus huanglongbin ; Sexual and graft incompatibility ; Aurantioideae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts isolated from `Page' tangelo (Minneola tangelo × clementine) cell suspension cultures were electrically fused with mesophyll protoplasts of orange jessamine [Murraya paniculata (L.) Jack]. Shoots were regenerated after 6 – 10 months of culture, but they were extremely recalcitrant to producing roots in root-induction medium. Complete plantlets were formed via micrografting. Chromosome counting of shoot tips revealed they were tetraploids (2n = 4x = 36). Glutamateoxaloacetate transaminase isozyme and randomly amplified polymorphic DNA analysis confirmed their hybridity. Orange jessamine is immune to citrus huanglongbin, a severe disease of citrus, but sexual incompatibility and limited graft compatibility exist between Citrus and orange jessamine. The cell fusion technique may make it possible to transfer the huanglongbin resistance trait from orange jessamine to Citrus.
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  • 8
    ISSN: 1432-203X
    Keywords: Key words Genetic stability ; Micropropagation ; Pinewood-nematode ; Pinus thunbergii ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Random amplified polymorphic DNA (RAPD) markers were used to determine the genetic stability of long-term (more than 10 years) micropropagated shoots of Japanese black pine (Pinus thunbergii Parl.). Thirty-six shoots consisting of three morphotypes (short, medium, and long needles) were randomly chosen from about 4,000 micropropagated shoots regenerated from the explants of a single nematode-resistant mother plant. Out of 126 primers screened, 30 gave 134 clear reproducible bands. A total of 4,824 bands obtained from these studies exhibited no aberration in RAPD banding patterns among the tested shoots. Our results show that regenerants from our plant micropropagation system are genetically stable.
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  • 9
    ISSN: 1432-203X
    Keywords: Key wordsPopulus alba L. ; Protoplast ; Plant regeneration ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We developed an efficient plant regeneration system from protoplasts for poplar (Populus alba L.). Protoplasts were isolated from 4-day-old suspension cultures derived from seed-induced calli with a yield of 6.96× 106 cells/g fresh weight cells and then cultured at a concentration of 2.5×105 cells/ml in NH4NO3-free Murashige and Skoog (MS) medium supplemented with 5 µM 2,4-dichlorophenoxyacetic acid (2,4-D), 0.05 µM thidiazuron (TDZ) and 0.5 M glucose as a osmoticum. The plating efficiency of the cultured protoplasts was calculated at 26.5% at day 7 and 31.7% at day 14. Cell colonies were observed after culturing for 4 weeks. Regenerated colonies were propagated through subculture in liquid MS medium supplemented with 5 µM 2,4-D. Buds were induced from regenerated calli on MS medium containing 10 µM kinetin or 1 µM TDZ. Regenerated shoots were rooted on half-strength MS medium, and the plantlets were transplanted in soil. Randomly amplified polymorphic DNA analysis did not detect any DNA polymorphism among the regenerated plants.
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  • 10
    ISSN: 1432-203X
    Keywords: Key words DNA content ; Morphology ; Protoplast fusion ; RAPD ; Somatic hybrid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Solanum acaule Bitt. is a disomic tetraploid (4x) wild potato species which is resistant to several potato diseases. Introgression of disease resistance and abiotic stress tolerance to the tetrasomic tetraploid (4x) cultivated potato (S. tuberosum L.) gene pool via crossing has been limited due to the difference in the endosperm balance number. In the present study, protoplast fusion was applied to produce hexaploid (6x) somatic hybrids between the parental lines, tetraploid (4x) S. acaule and two anther-derived dihaploid (2x) lines of S. tuberosum cv. White Lady. One callus (0.4%) of a total of 229 calli obtained regenerated into shoots in the fusion combination S. acaule (+) White Lady 15.dh.8.2.2. All the regenerated shoots were confirmed to be interspecific somatic hybrids using species-specific RAPD markers. In another fusion combination, S. acaule (+) White Lady 7.dh.23.1.1, fifteen calli (5%) regenerated into a total of sixteen shoots from 289 calli. All the analysed somatic hybrids between S. acaule and S. tuberosum were hexaploid. The mean DNA content (2C value) of the combination S. acaule (+) White Lady 15.dh.8.2.2 somatic hybrids (4.55 pg), was approximately the sum (4.69 pg) of the DNA contents of the parental lines, S. acaule (2.95 pg) and S. tuberosum (1.74 pg). In the greenhouse, the two somatic hybrids analysed were normal in their morphological characteristics and more vigorous than their parental lines. Most of the morphological characteristics were closer to the tetraploid S. acaule than to the dihaploid S. tuberosum. The interspecific somatic hybrids are currently being tested for frost tolerance and glycoalkaloid composition.
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  • 11
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    Theoretical and applied genetics 96 (1998), S. 101-111 
    ISSN: 1432-2242
    Keywords: Key words Hordeum ; Barley ; RAPD ; Variability ; Phylogeny ; DNA analyses
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The genetic variation of 102 natural populations of wild barley growing in Spain was assessed using RAPDs (random amplified polymorphic DNA). The plant material included the annual species H. marinum subsp. marinum (22 populations) and subsp. gussoneanum (14), H. murinum subsp. murinum (7) and subsp. leporinum (35), and the perennial species H. bulbosum (17) and H. secalinum (7). Ten of the tested 64 arbitrary 10-mer primers amplified polymorphic DNA in all taxonomic units. Analyses was performed within and between populations, species and subspecies. The primers gave a total of 250 RAPD products. The level of polymorphism varied between taxonomic units depending on the primers employed and the plant reproductive system. In general, the most variable were the allogamous species H. secalinum and H. bulbosum and the autogamous H. marinum subsp. marinum. Among the amplified bands, 69 (27%) were shared by at least two different taxonomic units. The remaining bands were specific. The results demonstrate differences in the degree of similarity between taxonomic units. Jaccard’s similarity coefficients for interval measure within and between populations were used to produce a cluster diagram using the unweighted pair-group method (UPGMA). The different populations of the species and subspecies of Hordeum fell into three groups. The first group contained the populations belonging to both subspecies of H. marinum, plus those of H. secalinum. The populations of H. marinum subsp. gussoneanum were very closely associated. Those of H. marinum subsp. marinum were grouped in a broad cluster. The second group, occupying the innermost position of the tree, was very closely associated with the populations of both subspecies of H. murinum. The third branch segregated H. bulbosum. A series of RAPD markers were investigated by cleaving the amplified products of the same size with restriction endonucleases that recognize targets of 4- or 6-bp. The production of equivalent fragments following cleavage by the same enzyme would seem to demonstrate their homology in samples from different individuals, populations or taxonomic units.
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  • 12
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    Theoretical and applied genetics 96 (1998), S. 683-687 
    ISSN: 1432-2242
    Keywords: Key words Essential oils ; Mint ; Mentha piperita ; M. spicata ; Somatic hybridization ; Plant regeneration ; RAPD ; Southern hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Twenty eight somatic hybrid plants were identified following protoplast fusions between peppermint (Mentha piperita L. cv Black Mitcham), producing high-quality oil, and spearmint (Mentha spicata L. cv Native Spearmint), likewise producing high-quality oil and also possessing resistance to verticillium wilt. Prior to fusion, peppermint protoplasts were subjected to iodoacetic acid to inhibit cell division. Protoplasts of peppermint and spearmint were fused using polyethylene glycol plus DMSO. Fusion products were cultured according to an efficient protoplast-to-plant-cycle protocol developed for peppermint. Using this protocol, iodoacetic acid-treated peppermint protoplasts were not able to divide, whereas untreated spearmint protoplasts had the ability to produce callus but not shoots. Therefore, selection of somatic hybrid calli was based on the presumed capability of hybrid cells to form calli and shoots. Shoots in vitro were initially identified as hybrids using RAPD profiles. Subsequently, observations on morphology, chromosome counts, and Southern-hybridization patterns confirmed their hybrid status. The results of verticillium tests revealed that 18 somatic hybrids were more susceptible than Native Spearmint, while hybrid II-14 had a level of susceptibility intermediate between that of the fusion parents. Oil-analysis of hybrid plants indicated that they all have a GC-profile typical of spearmint oil.
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  • 13
    ISSN: 1432-2242
    Keywords: Key words Medicago sativa ; RAPD ; Cultivars ; Genetic distance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Alfalfa (Medicago sativa L.) is a forage legume of world-wide importance whose both allogamous and autotetraploid nature maximizes the genetic diversity within natural and cultivated populations. This genetic diversity makes difficult the discrimination between two related populations. We analyzed this genetic diversity by screening DNA from individual plants of eight cultivated and natural populations of M. sativa and M.  falcata using the RAPD method. A high level of genetic variation was found within and between populations. Using five primers, 64 intense bands were scored as present or absent across all populations. Most of the loci were revealed to be highly polymorphic whereas very few population-specific polymorphisms were identified. From these observations, we adopted a method based on the Roger’s genetic distance between populations using the observed frequency of bands to discriminate populations pairwise. Except for one case, the between-population distances were all significantly different from zero. We have also determined the minimal number of bands and individuals required to test for the significance of between-population distances.
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  • 14
    ISSN: 1432-2242
    Keywords: Key words Mangroves ; Genome relationship ; Phylogeny ; RAPD ; RFLP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  DNA from pooled leaf samples of 11 true major mangrove, three true minor mangrove, two mangrove associate, two mangrove parasite, three terrestrial and one cultivated species were isolated for the present study. In total, 198 random amplified polymorphic DNAs (RAPDs) and 180 restriction fragment length polymorphism (RFLP) loci were scored by using ten primers and 14 enzyme-probe combinations respectively. The polymorphism observed for these markers revealed a high degree of genetic diversity in mangroves at both inter-specific or inter-generic levels. A dendrogram, constructed after pooling both RAPD and RFLP data, using a similarity index was analysed for genome relationships among these species. The dendrogram showed clustering of all the major mangroves, except for Nypa fruticans (Arecaceae), into one group. All species under the tribe Rhizophorae formed a sub-cluster, to which Xylocarpus granatum was found to be the most closesly related species. The clustering pattern implied that Excoecaria agallocha and Acanthus ilicifolius should be considered as true minor mangroves. The present study also provided molecular data favouring the separation of Avicennia spp. from the Verbenaceae to create a monotypic family the Avicenniaceae. The separation of Viscum orientale into the Viscaceae was also favoured.
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  • 15
    ISSN: 1432-2242
    Keywords: Key words RFLP ; RAPD ; Genetic map ; Null loci ; Gene family
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  We have constructed a sex-averaged genetic linkage map in coastal Douglas-fir (Pseudotsuga menziesii [Mirb.] Franco var ‘menziesii’) using a three-generation outcrossed pedigree and molecular markers. Our research objectives are to learn about genome organization and to identify markers associated with adaptive traits. The map reported here is comprised of 141 markers organized into 17 linkage groups and covers 1,062 centiMorgans (cM). Of the markers positioned on the map, 94 were derived from a Douglas-fir complimentary-DNA (cDNA) library that was constructed from new-growth needle tissue. Other markers include 11 Douglas-fir genomic-DNAs, 20 loblolly pine (Pinus taeda L.) cDNAs, 15 random amplified polymorphic DNAs (RAPDs) and a PCR-amplified phytochrome probe. A high degree of variation was detected in each of the two parents of our mapping population, and many of the restriction fragment length polymorphism (RFLP) and RAPD phenotypes were complex. Marker data were analyzed for linkage using mapping software JOINMAP version 2.0.
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  • 16
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    Theoretical and applied genetics 96 (1998), S. 791-796 
    ISSN: 1432-2242
    Keywords: Key words Sorghum ; RAPD ; CMS ; Fertility restoration ; MtDNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Seven sorghum restorer lines that differentially restore (or maintain) the A1 and A2 cytoplasmic male-sterile (CMS) cytoplasms were studied by RFLP analyses of their mtDNAs and RAPD analyses of their mitochondrial DNA (mtDNA) and total DNA to understand nuclear mitochondrial combinations that are present in these lines. RFLP data from 11 mitochondrial gene probes were inadequate to classify these seven lines. However, the analysis of RAPD profiles of total DNA could distinguish these lines on the basis of their ability to restore completely or partially the fertility in the A1/A2 CMS cytoplasms. Interestingly, RAPD profiles of mtDNAs of these lines also followed the same pattern as that of the total DNA. These results indicate that the different restorer lines possess specific nuclear-cytoplasm combinations. Further, the results also show that the RAPD technique can be used to identify markers for different cytoplasms used in CMS.
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  • 17
    ISSN: 1432-2242
    Keywords: Key words Brassica napus ; Raphanus sativus ; Restorer gene ; Introgression ; RFLP ; RAPD ; Genetic mapping
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Bulked segregant analysis and comparative mapping were applied to identify molecular markers linked to the Rfo restorer gene used for the Ogu-INRA cytoplasmic male-sterility system in rapeseed. These markers were then used to localise the radish introgression on the B. napus genetic map constructed from the cross ‘Darmor.bzh’ x ’Yudal’. The introgression mapped on the DY15 linkage group. From the comparison of this latter group to the linkage group constructed on a F2 progeny segregating for the radish introgression, it was concluded that the introgression had occurred through homoeologous recombination, that it was not distal and that it had replaced a B. napus region of around 50 cM. A QTL involved in aliphatic seed glucosinolate content was located on the DY15 linkage group at a position corresponding to one end of the introgression. The DNA markers identified in this study are being used in map-based cloning of the Rfo gene and in marker-assisted selection.
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  • 18
    ISSN: 1432-2242
    Keywords: Key words CAP ; py-1 ; RAPD ; RFLP ; Breeding
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  We report the molecular mapping of the py-1 gene for resistance to corky root rot [Pyrenochaeta lycopersici (Schneider and Gerlach)] in tomato using RAPD and RFLP marker analysis. DNA from near-isogenic lines (NILs) of tomato differing in corky root rot resistance was screened with 575 random oligonucleotide primers to detect polymorphic DNAs linked to py-1. Three primers (OPW-04, OPC-02, OPG-19) revealed polymorphisms between the NILs. Twelve resistant and eight susceptible DNA pools derived from segregating F3 families were used to confirm that the RAPD markers were linked to the py-1 gene. Two of the linked amplified fragments, corresponding to OPW-04 and OPC-02, were subsequently cloned and mapped on the tomato molecular linkage map as RFLPs. These clones were located between TG40 and CT31 on the short arm of chromosome 3. Further analysis with selected RFLP markers showed that 7% (8.8 cM) of chromosome 3 of the resistant line ‘Moboglan’ was introgressed from the L. peruvianum donor parent. Three RFLP markers (TG40, TG324, and TG479) from the introgressed part of chromosome 3 were converted to cleaved amplified polymorphism (CAP) markers for use in a polymerase chain reaction (PCR) assay. These PCR markers will allow rapid large-scale screening of tomato populations for corky root rot resistance.
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  • 19
    ISSN: 1432-2242
    Keywords: Key words Apical dominance ; Bulk segregant analysis ; Map ; Pea ; RAPD ; SCAR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Random amplified polymorphic DNA (RAPD) markers linked to two morphological markers ( fa and det), three ramosus genes (rms2, rms3 and rms4) and two genes conferring flowering response to photoperiod in pea (sn, dne) were selected by bulk segregant analysis on F2 populations. Two RAPD fragments were cloned and sequenced to generate the two SCAR markers V20 and S2 which are linked to rms3 and dne, respectively. All these genes, except rms2, were previously located on the pea classical linkage map. Rms2 mapped to linkage group IB which contains the afila gene. Precise genetic maps of the regions containing the genes were obtained and compared to the RAPD map generated from the recombinant inbred-lines population of the cross Térèse×K586. This cross was chosen because several mutants were obtained from cultivars Térèse and Torsdag (K586 was derived from Torsdag). This collection of isogenic lines was used for the construction of F2 mapping populations in which polymorphic RAPD markers were already known and mapped. Moreover, the well-known problem in pea of variability in the linkage associations between crosses was avoided. This work contributes to the precise integration between the classical map and the molecular maps existing in pea.
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  • 20
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    Theoretical and applied genetics 96 (1998), S. 37-45 
    ISSN: 1432-2242
    Keywords: Key words Microsatellite ; RAPD ; PCR ; Linkage map ; Wheats
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The potential of PCR-based markers for construction of a genetic linkage map in Einkorn wheat was investigated. From a comparison of polymorphisms between two Einkorn wheats, Triticum monococcum (Mn) and T. boeoticum (Bt), we obtained 49 polymorphic bands produced by 33 primers for inter-simple sequence repeat (ISSR) and 36 polymorphic bands shown by 25 combinations of random amplified polymorphic DNA (RAPD) primers for mapping in 66 individuals in the F2 population. Although 44 ISSR fragments and 29 RAPD fragments statistically showed a 3 : 1 segregation ratio in the F2 population, only 9 markers each of the ISSR and RAPD bands were able to be mapped on the RFLP linkage map of Einkorn wheat. ISSR markers were distributed throughout the chromosomes. The mapped positions of the ISSR markers seemed to be similar to those obtained by the RFLP markers. On the other hand, 4 of the 9 RAPD markers could map the RFLP marker-poor region on the short arm of 3Am, suggesting a potential to map novel regions containing repetitive sequences. Comparisons of the genetic linkage map of Einkorn wheat to the linkage map and cytological map of common wheat revealed that the marker orders between the two maps of Einkorn wheat and common wheat coincided except for 4A, which harbors chromosome rearrangements specific for polyploid wheats, indicating a conservatism between the two genomes. Recombinations in Einkorn wheat chromosomes took place more frequently around the centromere and less at the distal part of chromosomes in comparison to those in common wheat. Nevertheless, recombinations even in Einkorn wheat chromosomes were strongly suppressed around the centromere. In fact, the markers located within 1 cM of the centromere were located almost in the central part of the chromosome arm.
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  • 21
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    Theoretical and applied genetics 97 (1998), S. 950-959 
    ISSN: 1432-2242
    Keywords: Key words Vitis vinifera. L ; Seedlessness ; RAPD ; SCAR ; BSA ; Marker-assisted selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The variety Vitis vinifera cv Sultanine presents a type of seedlessness in which fertilization occurs but seeds subsequently fail to develop. It has been suggested that this trait might be controlled by three complementary recessive genes regulated by a dominant gene named I. Bulk segregant analysis was used to search for random amplified polymorphic DNA (RAPD) markers linked to the I gene in progeny obtained by crossing two partially seedless genotypes. One hundred and forty decamer primers were screened using bulks obtained by pooling the DNA of extreme individuals from the phenotypic distribution. We identified two RAPD markers which appeared tightly linked to I (at 0.7 and 3.5 cM respectively). The closest marker was used to develop a codominant SCAR (sequence characterized amplified region), named SCC8. This latter marker appeared of great value either to exclude from the progeny potentially seeded individuals or to select for seedless individuals. Indeed, all the seeded individuals of the progeny were found to be homozygous scc8 -/scc8 -, and all the individuals homozygous SCC8 +/SCC8 + were seedless. Moreover, this marker was successfully applied to other natural seedless varieties where codominance persisted. SCC8 was also used to dissect more precisely the genetics of seedlessness. ANOVA analysis indicated that this SCAR marker accounted for at least 64.9% of the phenotypic variation of the seed’s fresh weight and for at least 78.7% of the phenotypic variation of the seed’s dry matter. These results confirmed the presence of a major gene, and also the existence of other complementary recessive genes, controlling the expression of seedlessness.
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  • 22
    ISSN: 1432-2242
    Keywords: Key words Chromosome addition line ; Synteny group ; Brassica campestris ; Brassica oxyrrhina ; Monosomic ; Alloplasmic ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Monosomic chromosome addition lines of Brassica oxyrrhina in the background of alloplasmic B. campestris carrying B. oxyrrhina cytoplasm were generated and characterised through morphology, cytology and molecular (RAPD) analysis. Four successive backcrosses of the synthetic alloploid B. oxycamp with B. campestris yielded 24 monosomic addition plants that were grouped into seven different synteny groups based on morphological similarity and RAPD patterns. Each synteny group exhibited morphological features diagnostic for the presence of individual B. oxyrrhina chromosomes including some novel phenotypes. Meiotic studies of the addition lines revealed the homoeology of four B. oxyrrhina chromosomes (synteny groups 1, 3, 5 and 6 ) with B. campestris chromosomes as indicated by trivalent associations, with the highest homoeology (44.23%) in synteny group 1 and the lowest (6.1%) in synteny group 3. Seed fertility of the addition lines ranged from 94.85% (synteny group 1) to 56.98% (synteny group 5). All of the addition lines were male-sterile except synteny group 6 which had 12–16% stainable pollen. Ovule transmission of the B. oxyrrhina chromosomes added to the progenies of addition lines ranged from 23.52% (synteny group 6) to 14% (synteny group 7). RAPD analysis confirmed the validity of synteny grouping based on morphological observations. Approximately 45% of the primers studied were informative, giving B. oxyrrhina-specific RAPD bands unique for each synteny group, except group 6.
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  • 23
    ISSN: 1432-2242
    Keywords: Key words NESTUR ; Stem growth efficiency ; RAPD ; QTL ; Haploid megagametophyte
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    Topics: Biology
    Notes: Abstract  NESTUR (needle-to-stem unit rate) is a stem growth index of conifer seedlings that measures the efficiency of stemwood production per unit of needle growth, and is related to other seedling traits such as height, stem diameter, stem volume and needle volume. Quantitative trait loci (QTLs) affecting the expression of stem growth efficiency in radiata pine seedlings were investigated using a RAPD linkage map constructed from markers scored on haploid, megagametophytic DNA. Four putative QTLs were detected which accounted for 8.5–36.4% of the population variance. A search for evidence of epistasis, using both complete pairwise and conditional interactions, did not yield any statistically significant result. Over a 3-year period, seedlings with high-NESTUR marker alleles showed a superior growth performance of 17–40% for height, diameter and volume over those with low-NESTUR marker alleles.
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  • 24
    ISSN: 1432-2242
    Keywords: Key words Quercus robur L ; Linkage map ; RAPD ; SCAR ; Microsatellite ; Minisatellite ; 5S rDNA ; Isozymes
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    Topics: Biology
    Notes: Abstract  A genetic map of Pedunculate oak (Quercus robur) was constructed based on one 5S rDNA, 271 RAPD, ten SCAR, 18 microsatellite, one minisatellite, and six isozyme markers. A total of 94 individuals from a full-sib family was genotyped. Two maps, including 307 markers, were constructed according to the “two-way pseudo-testcross” mapping strategy. Testcross markers segregating in the 1 : 1 ratio were first used to establish separate maternal (893.2 cM, 12 linkage groups) and paternal (921.7 cM, 12 linkage groups) maps. Both maps provided 85–90% genome coverage. Homologies between the male and female linkage groups were then identified based on 74 intercross markers segregating in the 3 : 1, 1 : 2 : 1 and 1 : 1 : 1 : 1 ratios (RAPDs, SCARs, SSRs, 5S rDNA and isozymes) in the hybrid progeny. In each map, approximately 18% of the studied markers showed segregation distortion. More than 60% of the skewed markers were due to an excess of heterozygote genotypes. This map will be used for: (1) studying the molecular organisation of genomic regions involved in inter- and intraspecific differentiation in oaks and (2) identification of QTLs for adaptive traits.
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  • 25
    ISSN: 1432-2242
    Keywords: Key words Prunus persica ; Linkage map ; RFLP ; RAPD ; AFLP
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    Topics: Biology
    Notes: Abstract  A genetic linkage map of peach [Prunus persica (L.) Batch] was constructed in order to identify molecular markers linked to economically important agronomic traits that would be particularly useful for long-lived perennial species. An intraspecific F2 population was generated from self-pollinating a single F1 plant from a cross between a flat non-acid peach, ‘Ferjalou Jalousia®’ and an acid round nectarine ‘Fantasia’. Mendelian segregations were observed for 270 markers including four agronomic characters (peach/nectarine, flat/round fruit, acid/non-acid fruit, and pollen sterility) and 1 isoenzyme, 50 RFLP, 92 RAPD, 8 inter-microsatellite amplification (IMA), and 115 amplified fragment length polymorphism (AFLP) markers. Two hundred and forty-nine markers were mapped to 11 linkage groups covering 712 centiMorgans (cM). The average density between pairs of markers is 4.5 cM. For the four agronomic characters studied, molecular markers were identified. This map will be used for the detection of QTL controlling fruit quality in peach and, particularly, the acid and sugar content.
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  • 26
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    Theoretical and applied genetics 96 (1998), S. 621-627 
    ISSN: 1432-2242
    Keywords: Key words Cacao ; Theobroma cacao ; Genetic diversity ; Crop evolution ; RAPD
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    Topics: Biology
    Notes: Abstract  Neotropical tree crops are affected by a combination of biological and human factors that complicate the study of genetic diversity and crop evolution. Genetic diversity and relationships among southern Mexican populations and horticultural collections of Theobroma cacao (chocolate, cocoa, cacao) are examined in light of the agricultural practices of the Maya. Collections of cacao were obtained from the extremes of its geographic range including archeological sites in southern Mexico where cacao was first domesticated. Genetic diversity was assayed by 57 informative random amplified polymorphic DNA (RAPD) marker loci. A unique sample of the total diversity found in this study exists in the southern Mexican populations. These populations are significantly different from all other cacao with regards to their profile of RAPD bands, including the ‘criollo’ variety, their morphological and geographical group. A population of cacao found in a sinkhole (cenote) in northern Yucatan with genetic affinities to populations in Chiapas suggests the Maya maintained plants far away from their native habitat. This finding concurs with known agroforestry practices of the Maya. Modern efforts to increase germplasm of tropical tree crops such as cacao should carefully examine archeological sites where genetic diversity, either deliberately or by chance, was collected and maintained by ancient cultures.
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  • 27
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    Theoretical and applied genetics 96 (1998), S. 823-831 
    ISSN: 1432-2242
    Keywords: Key words Mangifera indica L. ; Anthracnose ; Somatic embryogenesis ; Dual culture ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Embryogenic nucellar cultures of two polyembryonic mango cultivars, ‘Hindi’ and ‘Carabao’, were selected for resistance to the culture filtrate and phytotoxin of a virulent strain of Colletotrichum gloeosporioides Penz. that was isolated from mango leaves. The cultures were recurrently selected either with progressively increasing concentrations of culture filtrate or by continuous challenge with the same concentration of either culture filtrate phytotoxin. Mycelium growth was inhibited when the pathogen was cocultured with the selected, resistant embryogenic cultures. Conditioned plant growth medium containing macerated resistant embryogenic cultures did not inhibit mycelium growth, confirming that extracellular antifungal compounds were involved in the defense response. Enhanced secretion of chitinase and glucanase was observed in the plant growth medium in which resistant embryogenic cultures and regenerated somatic embryos were grown in comparison with the controls.
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  • 28
    ISSN: 1432-2242
    Keywords: Key words Citrus ; RFLP ; RAPD ; Phylogeny ; Taxonomy
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    Topics: Biology
    Notes: Abstract  Relationships among 88 accessions representing 45 Citrus species, three man-made hybrids, and six related genera were examined for restriction fragment length polymorphisms (RFLP). Thirty-two Citrus and three Microcitrus accessions were also examined by random amplified polymorphic DNA (RAPD) analysis. A measure of relative heterozygosity was estimated based on the mean of the number of fragments per individual per probe-enzyme combination (PEC) divided by total number of fragments per PEC for all non-hybrid Citrus individuals. The presence in a Citrus species of a rare band found also in a related genus was taken as an indication of possible introgression, while the presence of several fragments unique to 1 species was used to indicate non-involvement of that species in hybridization events. Most species that have been described in the literature as hybrids had high heterozygosity indices and no unique fragments. Distance matrices and dendrograms were generated using simple matching coefficient and neighbor-joining cluster analysis. RFLP and RAPD data gave approximately the same results. These data showed C. maxima was affiliated with the papedas C. hongheensis and C. latipes. C. medica clustered with C. indica when only non-hybrid taxa were examined, or among limes, lemons, and relatives when all species were considered. Mandarins did not show strongly supported groupings among themselves, nor with other species. These data showed that several accessions were probably assigned to the wrong species.
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  • 29
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    Theoretical and applied genetics 96 (1998), S. 844-851 
    ISSN: 1432-2242
    Keywords: Key words Common bean ; Anthracnose resistance ; RAPD ; Genetic structure ; Centre of origin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The genetic structure of wild common bean populations was studied in the South-Andean centre of origin of the species. Plants were collected from 21 populations in Argentina and genetic variability was assessed for molecular and resistance markers. Polymorphism was weak for phaseolin, the major seed-storage protein, and for RAPD markers, while a high level of polymorphism was observed for resistance to anthracnose, one of the most important diseases of common bean. For the three traits, within-population variability was important and represented between 43.6% and 67.5% of the total variation. Although among-population differentiation was significant for all the traits, no correlation was found between the population distances calculated from RAPDs and resistance. These results indicate that pathogen selection pressure may be an important factor influencing the distribution of variability within and among host plant populations.
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  • 30
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    Theoretical and applied genetics 96 (1998), S. 1162-1169 
    ISSN: 1432-2242
    Keywords: Key words Wheat ; Russian wheat aphid ; Dn2 resistance gene ; RAPD ; SCAR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  RAPD (random amplified polymorphic DNA) analysis was used to identify molecular markers linked to the Dn2 gene conferring resistance to the Russian wheat aphid (Diuraphis noxia Mordvilko). A set of near-isogenic lines (NILs) was screened with 300 RAPD primers for polymorphisms linked to the Dn2 gene. A total of 2700 RAPD loci were screened for linkage to the resistance locus. Four polymorphic RAPD fragments, two in coupling phase and two in repulsion phase, were identified as putative RAPD markers for the Dn2 gene. Segregation analysis of these markers in an F2 population segregating for the resistance gene revealed that all four markers were closely linked to the Dn2 locus. Linkage distances ranged from 3.3 cM to 4.4 cM. Southern analysis of the RAPD products using the cloned RAPD markers as probes confirmed the homology of the RAPD amplification products. The coupling-phase marker OPB10880c and the repulsion-phase marker OPN1400r were converted to sequence characterized amplified region (SCAR) markers. SCAR analysis of the F2 population and other resistant and susceptible South African wheat cultivars corroborated the observed linkage of the RAPD markers to the Dn2 resistance locus. These markers will be useful for marker-assisted selection of the Dn2 gene for resistance breeding and gene pyramiding.
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  • 31
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    Theoretical and applied genetics 97 (1998), S. 83-89 
    ISSN: 1432-2242
    Keywords: Key words Lens culinaris subsp. orientalis ; Recombinant inbred lines ; AFLP ; RAPD ; Genetic map
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    Topics: Biology
    Notes: Abstract  A genetic linkage map of Lens sp. was constructed with 177 markers (89 RAPD, 79 AFLP, six RFLP and three morphological markers) using 86 recombinant inbred lines (F6:8) obtained from a partially interspecific cross. The map covered 1073 cM of the lentil genome with an average distance of 6.0 cM between adjacent markers. Previously mapped RFLP markers were used as anchor probes. The morphological markers, pod indehiscence, seed-coat pattern and flower-color loci were mapped. Out of the total linked loci, 8.4% showed segregation distortion. More than one-fourth of the distorted loci were clustered in one linkage group. AFLP markers showed more segregation distortion than the RAPD markers. The AFLP and RAPD markers were intermingled and clustering of AFLPs was seldom observed. This is the most extensive genetic linkage map of lentil to-date. The marker density of this map could be used for the identification of markers linked to quantitative trait loci in this population.
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  • 32
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    Theoretical and applied genetics 97 (1998), S. 99-102 
    ISSN: 1432-2242
    Keywords: Key words Genetic mapping ; CMS ; Fertility restoration ; RAPD ; RFLP ; Rye ; Secale cereale L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  A gene determining the restoration of cytoplasmic genic male sterility (CMS) caused by the Gülzow (G)-type cytoplasm was mapped by analyzing an F2 and F3 population comprising 140 and 133 individual plants, respectively. The target gene, designated Rfg1, was mapped on chromosome 4RL distally to three RFLP (Xpsr119, Xpsr167, Xpsr899) and four RAPD (XP01, XAP05, XR11, XS10) loci. Xpsr167 and Xpsr899 are known to be located on the segment of chromosome 4RL which was ancestrally translocated and is homoeologous to the distal end of other Triticeae 6S chromosomes. It is suggested that Rfg1 may be allelic to the gene determining the restoration of rye CMS caused by the Pampa (P) cytoplasm (chromosome 4RL) and to Rfc4 that on rye addition lines of chromosome 4RL restores male fertility of hexaploid wheat with T. timopheevi cytoplasm. Homoeoallelism to two loci for cytoplasmic-male-sterility restoration on chromosomes 6AS and 6BS in hexaploid wheat is also suggested.
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  • 33
    ISSN: 1432-2242
    Keywords: Key words Targeted mapping ; RFLP ; RAPD ; Brassica napus ; Polima CMS ; Nearly isogenic line ; Bulked segregant analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  We have used two targeting approaches [pairs of nearly isogenic lines (NILs) and bulked segregant analysis] to identify DNA markers linked to the Rfp1 restorer gene for the pol CMS of canola (Brassica napus L.). We were able to target the Rfp1 locus as efficiently by comparing NILs as by bulked segregant analysis, and it was demonstrated in this instance that double-screening strategies could significantly improve the overall targeting efficiency. The chance occurrence of shared homozygosity at specific unlinked chromosomal regions in the bulks was found to limit the efficiency of bulked segregant analysis, while the efficiency of NIL comparison was limited by residual DNA from the donor cultivar at scattered sites throughout the genome of the NILs.
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  • 34
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    Theoretical and applied genetics 97 (1998), S. 422-430 
    ISSN: 1432-2242
    Keywords: Key words Sunflower ; Helianthus ; Polyploidy ; Genomes ; RAPD
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    Topics: Biology
    Notes: Abstract  Forty taxa belonging to 36 species and four unclassified accessions of Helianthus were studied using RAPD technology. Single ten-mer primers were screened for those amplifying fragments common to several species. We found that when several species shared a common fragment, they belong to the same section of the genus. Moreover, we also found that some fragments are common to all species of the Helianthus. Most of the fragments were found to be of the same size in these species and to share the homology indicated by molecular hybridization. Out of 118 retained fragments, 33 were common to all Helianthus species, 56 were unique to perennial species of sects. Atrorubentes and Ciliares, 24 were unique to sect. Atrorubentes, 29 were unique to sect. Helianthus, whereas 0 were unique to sect. Ciliares. Each set of common or specific fragments was assumed to belong to a genome: (1) the C genome carrying the fragments common to all species of the three sections, (2) the H genome unique to sect. Helianthus, (3) the P genome common to perennial species (sects. Atrorubentes and Ciliares), and (4) the A genome unique to sect. Atrorubentes. The genomic structure was therefore HC for sect. Helianthus, CPA for sect. Atrorubentes, and CP? for sect. Ciliares. Molecular hybridizations with amplification products revealed homologies between Helianthus genomes and several other genera in the Helianthinae sub-tribe. The simple method used to characterize these fragments led to powerful tools for recognizing genomes which reconcile the section organization of the genus and the degree of difficulty in crossing perennial and annual forms.
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  • 35
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    Theoretical and applied genetics 97 (1998), S. 439-445 
    ISSN: 1432-2242
    Keywords: Key words Actinidia ; RAPD ; SCAR ; Sex-linked markers ; Marker-assisted selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  Two sex-linked random amplified polymorphic DNA (RAPD) markers identified from Actinidia chinensis were converted into sequence-characterised amplified regions (SCARs) for the large-scale screening of Actinidia breeding populations. Initial SCAR primers converted one RAPD (SmX) into a dominant marker, but the other (SmY), which was potentially more useful because of its linkage to the male determining ‘Y’ locus, failed to retain polymorphism. This difficulty was overcome by cloning and sequencing the alternate ‘allele’ from female plants, and then designing ‘allele’-specific primers that utilised nucleotide differences between the sexes. Using a quick squash-blot method of DNA extraction, the SCAR primers were tested in 120 A. chinensis plants to determine their gender. The system is now in use for large-scale screening of seedling populations in the Actinidia breeding programme. The sex-linked SCAR primers also functioned with plants from some other geographically separate accessions of A. chinensis and with plants in the closely related polyploid species A. deliciosa, but did not amplify a sex-linked band in more distantly related species of Actinidia.
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  • 36
    ISSN: 1618-2545
    Keywords: intraspecific group ; isozymes ; morphology ; Pythium ultimum var.ultimum ; RAPD
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    Topics: Biology
    Notes: Abstract Comparisons were made between two morphological groups ofPythium ultimum var.ultimum strains isolated in a vegetable field in Japan. The groups were distinguished as having smaller or larger sexual organs by the sizes of their antheridia and oogonia. Morphological study indicated that the two groups comprised a single taxon,P. ultimum var.ultimum, by the current taxonomical keys. The smaller group grew faster in the lower temperature range of 4–15°C, whereas the larger group grew faster in the higher temperature range of 25–37°C. Random amplified polymorphic DNA (RAPD) and isozyme analyses revealed genetic dissimilarity between the two groups. Cluster analysis of the isozyme banding patterns with four otherPythium spp. demonstrated that the genetic dissimilarity between the two groups was equivalent to species level. In the field survey, the smaller group was frequently detected in February, May and September but not in July, while the larger group was detected mainly in July and September. The two groups were not distinguishable by their pathogenicity to spinach seedlings.
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  • 37
    ISSN: 1618-2545
    Keywords: Coprinus ; RAPD ; strain identification
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    Topics: Biology
    Notes: Abstract All five examined strains ofCoprinus cinereus could be clearly discriminated from the strains of five otherCoprinus species by RAPD patterns with 12 of 13 primers. Also one specimen of unknownCoprinus strain was identified to beC. cinereus by this method. The RAPD patterns were similar among the strains in the same species; many common DNA fragments were recognized as well as some strain-specific DNA fragments. Thus all seven strains ofC. cinereus and all four strains ofC. angulatus examined could be distinguished individually. Diakryotic strains showed the combined RAPD patterns of the two monokaryotic strains constituting the dikaryon. The combined RAPD markers observed in the dikaryons were segregated in their basidiospore progeny. All 18 randomly picked progeny showed different combinations of RAPD markers from the parental strains.
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  • 38
    ISSN: 1572-9788
    Keywords: transgenic rice ; particle bombardment ; cell electroporation ; RAPD ; AFLP ; AFRP ; RAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract In the present work we utilised some of the most discriminative molecular tools, such as RAPD, AFLP, AFRP and RAMP, to analyse the genome of independently derived transgenic plants from three elite Italian cultivars (cv. Lido, Carnaroli and Thaibonnet) and found that two methods for direct gene transfer, namely particle bombardment and intact cell electroporation (the latter being a procedure set up in this work), result in transgenic rice (Oryza sativa L.) plants that exhibit negligible genomic changes. This is in contrast with recently published results showing relevant changes in the DNA of transgenic rice plants generated through protoplasts electroporation and of transgenic poplar plants engineered through Agrobacterium tumefaciens infection. Implications of these findings are discussed in the context of selecting appropriate gene transfer methodologies to produce transgenic plants expressing genes of interest while retaining their genomic integrity and, thus, their superior agronomic and/or industrial traits.
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  • 39
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    Molecular breeding 4 (1998), S. 227-234 
    ISSN: 1572-9788
    Keywords: Helianthus annuus ; PCR ; Puccinia helianthi ; RAPD ; rust resistance ; sunflower
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract In this study we report on the identification of molecular markers, OX20600 and OO04950, linked to the geneR Adv in the proprietary inbred line P2. This gene confers resistance to most of the pathotypes of Puccinia helianthi identified in Australia. Analysis indicates these RAPD markers are linked to the resistance locus at 0.0 cM and 11 cM respectively. SCAR markers SCX20600 and SCO04950 derived from these two RAPD markers, and SCT06950 derived from a previously reported RAPD marker linked at 4.5 cM from the R 1 rust resistance gene were developed. SCX20600 and SCO04950 were linked at similar distances from their resistance locus as the RAPD markers. SCTO6950 co-segregated completely with rust resistance. The robustness of the R 1 SCAR marker was demonstrated through the amplification of the marker in a diverse range of sunflower germplasm considered to possess the R 1 gene. The SCAR markers forR Adv were not amplified in the sunflower rust differential set thereby supporting the contention that this is a novel resistance gene. They did amplify in a number of proprietary lines closely related to the line P2. This locus is under further investigation as it will be useful in our attempts to use molecular-assisted breeding to produce durable resistance in sunflower to P. helianthi.
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  • 40
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    European journal of plant pathology 104 (1998), S. 611-617 
    ISSN: 1573-8469
    Keywords: Exserohilum turcicum ; Johnson grass ; maize ; northern corn leaf blight ; population genetic structure ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Setosphaeria turcica is the causal agent of northern corn leaf blight, a foliar maize disease of worldwide economic importance. In Europe, its severity increases. To investigate the pathogen's population-genetic structure in central Europe, a total of 80 isolates was sampled in Germany, Switzerland, France, Austria, and Hungary and investigated with 52 random amplified polymorphic DNA (RAPD) markers. The mating type of the isolates was determined in testcrosses. Among the 73 isolates from maize there were 26 different RAPD haplotypes. All isolates with identical haplotype are considered clonemates. The haplotype shared by most members was represented by 22 isolates from Germany, Switzerland, and France, indicating high fitness and substantial migration. Only a single clone had members in both southeastern Austria and southwestern Switzerland, suggesting that the Alps constitute a major barrier for this pathogen. Several haplotypes differed by only one or two RAPD bands from the predominant haplotype and may have arisen by mutation. Few other clonal lineages were detected. The evolution of some haplotypes could not be explained by mutation alone. Sexual recombination may rarely occur. In population samples from Germany, Switzerland, and France, mating type MAT2 was predominating, while most isolates from Austria and Hungary had MAT1. Seven isolates from Johnson grass (Sorghum halepense), an alternative host of S. turcica, were clonemates and very different in RAPD haplotypes from all isolates collected from maize.
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  • 41
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    Plant systematics and evolution 212 (1998), S. 53-77 
    ISSN: 1615-6110
    Keywords: Leguminosae ; Leucaena leucocephala ; L. diversifolia ; L. ×spontanea ; Hybridization ; hybrid detection ; spontaneous hybrid ; RAPD ; RFLP ; chloroplast DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The detection of hybridity inLeucaena is discussed in relation to: (i) traditional criteria, (ii) molecular criteria and (iii) models to predict hybrid leaf morphology. Morphological, geographical and molecular evidence for the occurrence of interspecific hybrids betweenL. leucocephala andL. diversifolia in south-central Mexico, northern Guatemala, Jamaica, Dominican Republic, the Philippines and Papua New Guinea is presented. Predicted mean hybrid leaf trait values calculated from parent material are compared with data from putative hybrids and shown to be similar. The origin of these hybrids is discussed and shown to be the result of artificial sympatry resulting from indigenous, and recent exotic, domestication of the parent species. The hybrid is described asL. ×spontanea.
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  • 42
    ISSN: 1573-5060
    Keywords: sweet potato ; Ipomoea batatas ; RAPD ; genetic variation ; Chile
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    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract RAPD (randomly amplified polymorphic DNA) technology was applied to analyze the genetic variability of sweet potato germplasm existing in Chile and elsewhere. Analysis of 28 cultivars from all over the world showed polymorphic bands with all 18 primers tested. A total of 124 RAPD bands were scored with an average of 6.9 polymorphic bands per primer. These results confirm that sweet potato exhibits high genetic variation. Two groups were distinguished: one containing Peruvian cultivars, and another containing cultivars from the rest of the world. Analysis of 14 accessions from Central Chile and one from Northern Chile showed polymorphic bands with 24 of 26 primers tested, but almost all of the 140 polymorphic bands merely showed the distinctness of the Northern accession. The almost complete uniformity of the other 14 accessions shows that sweet potato germplasm collected in Central Chile has very little genetic variability and may be derived from a single cultivar. Based on these results and on historical records, some hypotheses are proposed to explain the origin of sweet potatoes cultivated in Chile.
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  • 43
    ISSN: 1573-5060
    Keywords: cultivar identification ; fingerprinting ; genetic similarity ; Juglans regia ; RAPD ; walnut
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The potential use of the Randomly Amplified Polymorphic DNA (RAPD) technique for characterization and assessment of genetic relationships was investigated in nineteen walnut (Juglans regia L.) genotypes used as parents or released as cultivars from the breeding program of the University of California at Davis. Most of the 72 decamer primers used yielded scorable amplification patterns based on discernable bands. The results obtained produced a unique fingerprint for each of the walnut genotypes studied. Cluster analysis separated the 19 walnut genotypes into two main groups whose differences were related to their pedigree. Genotypes sharing common parents tend to group together and with at least one of the parents. Thus, RAPD markers can detect enough polymorphism to differentiate among walnut genotypes, even among closely related genotypes, and the genetic similarity based on RAPDs appears to reflect the known pedigree information. RAPD technology can be useful in current walnut breeding programs, allowing the identification of new cultivars as well as the assessment of the genetic similarity among genotypes which will help in selecting the best parents to obtain new genetic combinations.
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    Euphytica 101 (1998), S. 249-255 
    ISSN: 1573-5060
    Keywords: avocado ; classification ; ecological races ; Persea americana ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract RAPD (randomly amplified polymorphic DNA) analysis was carried out on 16 accessions representing the three ecological races of avocado (Persea americana Mill.), and one accession of P. schiedeana Nees. Twenty two preselected primers produced 133 polymorphic DNA fragments in the RAPD assay of the avocado accessions. One primer was identified which could differentiate each of the avocado accessions. Potentially race-specific markers for each of the Mexican, Guatemalan, and West Indian races, have been detected. A Jaccard's similarity coefficient matrix was generated and a dendrogram constructed using UPGMA (unweighted pair-group method of arithmetic averages) cluster analysis. Percentage similarity between avocado accessions ranged from 46% to 85%. The lowest similarity (between 22% and 29%) was revealed between P. schiedeana and any P. americana accession. Average similarity within races of avocado was 75% for the Mexican race, 71% for the West Indian race and 73% for the Guatemalan race. Average similarity between races ranged from 53% to 58%. The dendrogram identified three groups, representing the races of avocado. These results are in concordance with the present classification of avocado into three subspecies (varieties) of P. americana, namely drymifolia, americana, and guatemalensis, corresponding to the Mexican, West Indian and Guatemalan races, respectively, and confirm the separate species status of P. schiedeana. We conclude that RAPD markers may be useful for the classification of avocado and for the assessment of genetic diversity of avocado germplasm.
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  • 45
    ISSN: 1573-5060
    Keywords: Brassica napus ; RAPD ; bulked DNA ; DNA fingerprinting
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Since DNA-based markers are unaffected by environmental or physiological factors, they have potential utility in the description of plant cultivars required for award of proprietary rights (i.e. Plant Breeders' Rights). The high discriminating power of this class of markers, however, can also make demonstration of uniformity and stability of such a marker within a cultivar difficult, especially for genetically-complex cultivars. This report examines the usefulness of bulking equal quantities of DNA from 14 to 20 individuals of a cultivar to identification of RAPD DNA markers that distinguish between Brassica napus cultivars of varying genetic complexity. For the four cultivars assessed (Quantum, OAC Springfield, Innovator and AC Excel), it is shown that consistent presence/absence scores are obtained from bulked DNA samples for three different RAPD markers despite a significant degree of variation among samples from individuals. Use of bulked DNA samples thus may enable identification of a distinguishing profile of RAPD markers whose presence/absence is uniform and stable even in complex cultivars. Nevertheless, RAPD markers remain limited in that they are not strictly quantitative in nature. This limitation is discussed with respect to cultivar description for plant breeders' rights applications.
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  • 46
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    Euphytica 104 (1998), S. 181-189 
    ISSN: 1573-5060
    Keywords: Genetic diversity ; macha wheat ; spelta wheat ; RAPD ; accession duplication
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Genetic diversity in a crop species is basic to improvement of the species and can be estimated at the molecular level. The objective of this study was to estimate genetic diversity within and between spelta and macha wheats. Random amplified polymorphic DNA (RAPD) analysis was conducted on 69 spelta and 32 macha wheat accessions. The classification of spelta and macha accessions, based on Jaccard genetic similarity coefficients for RAPD markers, was consistent with their geographic origin. The results indicated that the germplasm of macha wheat was more diverse than that of spelta wheat. In the dendrogram of macha wheat, four spelta-like accessions grouped together, separate from the remaining macha accessions, suggesting that these accessions were misclassified. In addition, accessions with identical RAPD patterns were found, indicating that these accessions were probably duplicated. Thus RAPD analysis can be used to estimate genetic diversity and identify duplicate accessions in wheat germplasm collections.
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  • 47
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    Euphytica 99 (1998), S. 167-173 
    ISSN: 1573-5060
    Keywords: AFLP ; Capsicum annuum ; genetic distances ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Genetic relationships were examined among thirty-four pepper (Capsicum annuum) cultivars of different types. Two types of PCR-based markers were used, RAPD and AFLP, and their relative effectiveness was compared. A dendrogram based on RAPD markers separated the large-fruited sweet cultivars from the small-fruited pungent peppers, and the former group showed less divergence than the latter. The percentage of polymorphic markers was lower for AFLP than for RAPD markers (13 and 22% respectively). However, AFLP primers amplified on average six times more products than RAPD markers. The average numbers of polymorphic products per primer were 1.6 and 6.5 for RAPD and AFLP primers, respectively, i.e., AFLP primers were four times more efficient than RAPD primers in their ability to detect polymorphism in pepper. While four blocky type cultivars were indistinguishable by RAPD, two AFLP primer pairs were sufficient to distinguish the four cultivars from each other.
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  • 48
    ISSN: 1573-5060
    Keywords: embryo sac ; RAPD ; interspecific hybrid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract In cassava, apomixis fixes heterosis and avoids transmission of systemic pathogens which complicate vegetative propagation of the crop. A combination of evidence from maternal inheritance of RAPD markers and the structure of the embryonic sac in large progeny sets of two distinct genotypes have further confirmed the occurrence of apomixis in cassava. We could advance further on earlier reports of the detection of apomixis in four ways: (1) we could arrive at an estimate of the rate of facultative apomixis in the range of 2%; (2) we detected the occurrence of apomixis in a second genotype, derived from a different interspecific cross; (3) apomictic behavior was demonstrated in an F1 individual and (4) parallel embryonic evidence was generated that corroborate the potential occurrence of apomixis by apospory. The fact that apomixis was detected in an F1 interspecific hybrid hints to the possibility of directly transferring genes for apomixis from a wild relative to cultivated cassava.
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  • 49
    ISSN: 1573-5060
    Keywords: bottleneck ; isozyme ; landrace ; lentil ; RAPD ; South Asia
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Lentil landraces from South Asia exhibit a low diversity and discordance with landraces from other countries according to a combination of qualitative and quantitative agromorphological characters. They exhibit specific phenological adaptation to the South Asian environment which precludes the direct use of alien germplasm in breeding programs in South Asia. An understanding of the genetic relationships and diversity of South Asian lentil landraces, in relation to landraces from other countries, is important in attempting to widen the genetic base of germplasm in the region. The objectives of this study were to investigate the genetic relationships between lentil landraces from 3 South Asian countries (India, Nepal and Pakistan) and those from 13 other countries and to determine their relative genetic diversities, using both isozyme electrophoresis and random amplified polymorphic DNA (RAPD) analysis. Polymorphisms were observed for 7 isozyme loci (16 alleles) and 22 RAPD loci. According to Nei's genetic distance, germplasm from Afghanistan clustered with that from the South Asian countries. The germplasm from these countries was striking different to that from the other countries studied. Based on genetic distance estimates from RAPD analysis, the countries with the lowest diversity were Pakistan, Afghanistan and Nepal. These data support evidence at the morphological level of a genetic bottleneck in lentil landraces from South Asia. Genetic relationships between countries outside the South Asian group are discussed. Classification into macrosperma and microsperma types did not reflect overall country relationships.
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  • 50
    ISSN: 1573-5060
    Keywords: dihaploid Solanum tuberosum ; Phytophthora infestans (Mont.) de Bary ; polygenically inherited resistances ; somatic hybrids ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Five dihaploid Solanum tuberosum genotypes, encompassing different levels of polygenically inherited resistance to potato late blight disease (Phytophthora infestans (Mont.) de Bary) in foliage and tubers, were used in four intraspecific protoplast fusion combinations. Vigorous growing putative hybrid calli were selected four weeks after electrofusion. Intact plants were regenerated from 6% of the selected calli. Verification of hybridity was accomplished by use of RAPD analysis which revealed that 53% of the regenerated plants were true somatic hybrids. The score of true somatic hybrids in the different fusion combinations ranged within 21% – 100%. The hybrid plants were analysed for resistance to foliage blight in a field trial and assessed for tuber blight resistance by use of a laboratory test. Resistance to late blight in foliage and tubers varied between the hybrids. Very high levels of resistance to both foliage and tuber blight were obtained in some hybrids. However, loss of resistance in some hybrids as compared to the parental plants were also observed. Possible reasons for the phenotypic disappearance of resistance to either foliage or tuber blight are suggested.
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  • 51
    ISSN: 1573-5060
    Keywords: taro ; Colocasia esculenta ; RAPD ; DNA fingerprinting ; phylogenetic tree
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Forty-four taro (Colocasia esculenta), two tanier (Xanthosoma species) and one Colocasia gigantea accessions were evaluated for genetic diversity using random amplified polymorphic DNA (RAPD) primers. Seventy-three of 112 primers amplified PCR DNA products used to fingerprint the accessions. Thirty-two primers were considered highly informative because they amplified more than 5 bands or amplified one or more polymorphic bands that distinguished between accessions. RAPDs showed high genetic diversity in taro accessions from Indonesia, were capable in distinguishing between Hawaiian accessions, and could separate triploid from diploid accessions. UPGMA cluster analysis of genetic similarity estimates (Jaccard's coefficient), separated the accessions into 3 main groups with C. esculenta divided into 5 subgroups. These primers will be useful for future genetic analysis and provide taro breeders with a genetic basis for selection of parents for crop improvement. Polymorphic markers identified in the DNA fingerprinting study will be useful to screen a segregating population which is being generated in our laboratory aimed at developing a taro genetic linkage map.
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  • 52
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    Genetic resources and crop evolution 45 (1998), S. 511-523 
    ISSN: 1573-5109
    Keywords: Allium ; crop evolution ; GISH ; hybridogenic cultivars ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Three vegetative crops of sect. Cepa in genus Allium (Top onion, French grey shallot and viviparous triploid onion) of suspected hybridogenic origin were studied with genomic in situ hybridization (GISH) and random amplified polymorphic DNA (RAPD) markers. Related wild and cultivated species were included in the analysis in order to assess their contributions to the genomes of the investigated species. In A. × proliferum, the parental chromosomes derived from A. fistulosum and A. cepa were unequivocally identified by GISH, proving the hybrid status of this crop. The French grey shallot proved to belong to A. oschaninii according to the RAPD analysis and the GISH results, it is clearly separate from the normal shallots of A. cepa var. aggregatum. Thus the grey shallots are a new crop species and can be considered as an aggregatum form of the wild progenitor species A. oschaninii. The triploid viviparous onion comprises mostly A. cepa derived DNA in its genome. A non-cepa component could not be attributed to any of the sect. Cepa species included in this study and is most likely derived from one of the species of this section not yet identified by molecular means.
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  • 53
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    Genetic resources and crop evolution 45 (1998), S. 243-251 
    ISSN: 1573-5109
    Keywords: Phaseolus ; genetic variability ; amino acids ; isozymes ; proteins ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The genetic variation within and between Spanish landraces or varieties of Phaseolus vulgaris L. (common bean) and P. coccineus L. (runner bean) has been estimated by means of isozymes and random amplified polymorphic DNA (RAPD) analyses. Likewise, storage protein and amino acid content in dry seeds have been estimated. Fifteen landraces (60 accessions) of P. vulgaris and six of P. coccineus (six accessions) have been studied. Of the seven isozymatic systems analyzed only three systems and three loci showed variability in each species. Isozyme analyses revealed that genetic variability within and between landraces exist in both species. Even variability within accession was detected in some P. vulgaris landraces. Comparison of isozyme data indicated that Spanish landraces have a lower level of genetic variability than wild American materials and probably also lower than American landraces. RAPD analysis allowed for the uniquely distinguishing of all landraces. Genetic similarity among landraces, estimated by both isozymes and RAPDs, were not related with the seed morphological characters (color, size and shape) which define each variety or landrace. Variation in protein and amino acid content among landraces was also detected. The average protein content in common bean (20.48%) was similar to values previously reported in this species and higher than the average in the runner bean landraces (16.33%). In relation to the amino acid content methionine and cysteine showed the lowest values in all samples, although the content of these two amino acids varied widely among landraces.
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  • 54
    ISSN: 1573-5109
    Keywords: DNA fingerprints ; genetic diversity ; Ipomoea batatas ; polymerase chain reaction ; RAPD ; sweetpotato
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The island of New Guinea is considered a secondary center on diversity for sweetpotato, because of its range of isolated ecological niches and large number of cultivars found within a small area. Information of genetic diversity in Papua New Guinea (PNG) sweetpotato is essential for rationalizing the global sweetpotato germplasm collection. Using random amplified polymorphic DNA (RAPD), we compared the genetic variation and genetic diversity in 18 PNG cultivars versus 18 cultivars from South America. The analysis of molecular variance revealed large genetic diversity in both groups of cultivars. The within-group (among individuals) variation accounted for 90.6% of the total molecular variance. However, the difference between PNG and South American groups is statistically significant, although it explained only 9.4% of the total molecular variance. The PNG cultivars are also less divergent than their South American ancestors as the mean genetic distance in PNG group is significantly smaller than that of South American group. The lower level of genetic diversity in PNG cultivars was also reflected by multidimensional scaling. This study shows that PNG cultivars, after many years of isolated evolution in an unique agro-ecological environment are substantially divergent from their ancestors in South America. The genetic diversity level in PNG cultivars is significantly lower than that in South American cultivars. It thus provides a baseline for continuing studies of genetic diversity in different sweetpotato gene pools.
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  • 55
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    Genetic resources and crop evolution 45 (1998), S. 337-342 
    ISSN: 1573-5109
    Keywords: burr medic ; genetic variability ; Medicago ; outcrossing rate ; RAPD ; selfing
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Although burr medic (Medicago polymorpha L.) is commonly considered a self-pollinating species, intrapopulational variation for morphological, biochemical and molecular markers is relatively high. To investigate whether part of this variation is the result of outcrossing, we designed RAPD analysis experiments to reveal both inter- and intra-accession crossing. No cases of inter-accession hybrids were documented, but an intra-accessional crossing rate of 0 to 4% was estimated for one of the four accessions studied. Therefore, a rare outcrossing event between local individuals and migrating genotypes may contribute to high genetic variability observed in natural M. polymorpha. A better understanding of the factors which influence outcrossing in M. polymorpha is pertinent both for medic breeding programmes and for assessing risks associated with releasing transgenic herbicide-resistant crops.
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  • 56
    ISSN: 1573-5109
    Keywords: AMOVA ; Lens ; population genetic structure ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract An understanding of the genetic structure of populations is vital for the formation of optimum collection, conservation and utilization strategies for plant genetic resources. This is of particular importance in the case of in-situ conservation, a strategy gaining in popularity. The population genetic structures of five wild lentil taxa, Lens culinaris subsp. orientalis, L. odemensis, L. ervoides, L. nigricans and L. lamottei were investigated using isozyme electrophoresis and random amplified polymorphic DNA (RAPD). Approximately 20 plants from each of 5 populations per taxon were screened for variation at 11 isozyme loci and using three RAPD primers. Levels of variation were generally low, although considerable variation existed in the levels of diversity found within populations of L. culinaris subsp. orientalis and L. lamottei. Comparison of the results obtained in this study with the results obtained in a previous study indicate that this is a trend occurring across all species. It implies that levels of diversity within populations must be measured and considered prior to targeting of specific populations for in-situ conservation. Analysis of molecular variance of both isozyme and RAPD data revealed that between 78% and 99% of the variation was attributable to between-population differences. Isozyme results from L. lamottei populations were, however, contradictory. Possible explanations for this difference are discussed.
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  • 57
    ISSN: 1573-5117
    Keywords: RAPD ; Littorina saxatilis ; L. neglecta ; phylogeny
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A genetic analysis of ‘morphotypes’ of Littorina saxatilis from two locations on the north-east coast of England (Filey and Ravenscar), using randomly amplified DNA polymorphisms (RAPD) generated with a single primer, revealed quite different patterns of variation. Thin shelled, wide-apertured (H-form) animals from Ravenscar tended to cluster separately from thick shelled (M) forms, indicating genetic differentiation of these morphs. Animals of similar morphology (H and M) from Filey (about 30 km distant) did not display such an obvious pattern, and although there was still evidence of differentiation from discriminant analysis of RAPD data, levels of correct classification were reduced at Filey. This suggests that the utility of a single RAPD primer for separation of such forms varies over a relatively small distance. L. arcana from Ravenscar, included as an outgroup, were generally well differentiated from L. saxatilis and were noted to exhibit less variation, a phenomenon that has been noted previously in some allozyme and RAPD analyses. A similar RAPD analysis undertaken on small, barnacle dwelling, brooding forms from Peak Steel, Ravenscar revealed that animals appeared to have as great a tendency to cluster together on a microgeographic scale (by collection patch) as by ‘species’ ( L. neglecta or L. saxatilis b) although predominance of certain species in individual patches largely explains this. Discriminant analysis of RAPD presence/absence data did correctly place over 90% of barnacle dwelling animals to their respective species, and we consider this as evidence of separate gene pools. RAPD is taken to be a useful tool for screening genetic variation in this complex of animals on a local scale when either a pre-selected informative primer is utilised or a battery of primers is used, but its efficacy may be reduced when a single primer is employed for screening animals from different shores.
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  • 58
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    Hydrobiologia 378 (1998), S. 33-42 
    ISSN: 1573-5117
    Keywords: protocols ; RAPD ; Littorina saxatilis ; L. obtusata ; L. fabalis ; L. littorea ; extracting DNA ; amplifying DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Randomly amplified polymorphic DNA (RAPD) is a fast and useful method of genome marking that is useful for studies of, for example, parentage, mating patterns, taxonomy of sibling species and intra-specific population genetic structures. Here we compare three different procedures for extracting high molecular weight genomic DNA; phenol-chloroform, hexadecyltrimethyl ammonium-bromide (CTAB) and Chelex 100. Double phenol-chloroform and CTAB extractions both generated high amounts of high quality DNA while Chelex 100 failed to do so. We also compared PCR-amplification with different concentrations of template DNA and found that 1–2 ng per 25 μl of amplification cocktail gave the best results. Amplifying DNA prepared by the three extraction methods revealed that DNA extracted with double phenol-chloroform gave the clearest bands. The double phenol-chloroform extraction seems thus the most suitable extraction method for RAPD in Littorina, however Chelex may be the only method useful for extracting DNA from very small individuals, for example, pre-hatching stages.
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  • 59
    ISSN: 1573-6857
    Keywords: genus Pellia ; liverworts ; RAPD ; taxonomy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The random amplified polymorphic DNA (RAPD) technique was used to study the genomic relationships of three liverworts from the genus Pellia: P. epiphylla, P. borealis and P. neesiana. Altogether 150 characters (150 DNA fragments obtained using PCR) were scored. These characters were used to create a matrix of pairwise distances between all the pairs of taxa. Both distance (UPGMA, Fitch–Margoliash and Neighbor–Joining) and binary character‐state (Wagner and Camin‐Sokal parsimony and compatibility) methods were applied for trees' construction. Our results strongly support distinction of the recently discovered sibling species of P. epiphylla – species N and P. epiphylla – species S, which have an allopatric distribution in Poland (N – North, S – South Poland). Moreover, our data also supports the hypothesis of a hybrid origin (alloploid) of the polyploid P. borealis from P.epiphylla−N×P.epiphylla−S. P. neesiana was excluded as a donor of either of the genomes of P. borealis.
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  • 60
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    World journal of microbiology and biotechnology 14 (1998), S. 415-420 
    ISSN: 1573-0972
    Keywords: Arbitrary primers ; Brucella abortus ; Brucella melitensis ; polymerase chain reaction ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Randomly amplified polymorphic DNA (RAPD) profiles of Brucella and non-Brucella DNA were established after polymerase chain reaction (PCR) amplification. Five arbitrary oligonucleotide primers were screened to generate Brucella-specific DNA fingerprints. The arbitrary primer OPB-01 (5′-GTTTCGCTCC-3′) produced DNA bands specific to Brucella. Amplification conditions must be optimized for reproductibility. Accordingly, we optimized and established the conditions, which included Mg2+, enzyme (DNA polymerase), primer, template and deoxyribonucleoside triphosphate (dNTP) concentrations as well as the optimum number of thermal cycles to produce OPB-01 directed Brucella DNA fingerprints. The optimized RAPD method can produce a 1.3 kb DNA fragment specific to Brucella. This DNA fragment was common to eight biovars of B. abortus and one biovar of B. melitensis. The fragment was not detected in genetically related species such as Ochrobactrum anthropi and other non-Brucella organisms associated with farm animals. We anticipate the use of this fragment as a possible probe for the detection of Brucella organisms.
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  • 61
    ISSN: 1572-9699
    Keywords: Sinorhizobium meliloti ; RAPD ; AMOVA ; population genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We analysed the genetic diversity of 270 Sinorhizobium meliloti strains isolated from nodules of three different Medicago sativa varieties, planted in three different Italian soils, combining the Analysis of Molecular Variance (AMOVA) with the Random Amplified Polymorphic DNA (RAPD) technique to estimate variance among RAPD patterns with the aim to draw an objective description of the population genetic structure. Results indicated that a general intraspecific genetic diversity was globally distributed among all the population, however a very high level of diversity was found among strains nodulating different Medicago sativa varieties. Moreover the distribution of the RAPD haplotypes among the plant varieties also showed to be non-random. The overall data indicated that the plant genotype is a major factor in shaping the genetic structure of this natural Rhizobium population.
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  • 62
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    Plant cell, tissue and organ culture 54 (1998), S. 85-91 
    ISSN: 1573-5044
    Keywords: cpDNA ; Ebenaceae ; electrofusion ; flow cytometry ; polyploid ; RAPD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Interspecific somatic hybrids between Diospyros glandulosa (2n=2x=30) and D. kaki cv. Jiro (2n=6x=90) were produced by electrofusion of protoplasts. Protoplasts were isolated from calli derived from leaf primordia, fused electrically, and cultured by agarose-bead culture using a modified KM8p medium. Flow cytometry revealed that the nuclear DNA content was the sum of those of D. glandulosa and D. kaki cv. Jiro in 149 of the 166 calli obtained. RAPD analysis showed that the 149 callus lines yielded specific bands for both D. glandulosa and D. kaki cv. Jiro and further confirmed that they were interspecific somatic hybrid calluses. Shoots were regenerated from 63 of the 149 interspecific hybrid calluses. Chloroplast DNA analysis by PCR-RFLP, flow cytometric determination of nuclear DNA content, and RAPD analysis revealed that the 63 interspecific hybrid shoot lines contained the nuclear genomes from both parents but only the chloroplast genome from D. glandulosa. Microscopic observation of root tip cells demonstrated that somatic chromosome number of the interspecific hybrids was 2n=8x=120.
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  • 63
    ISSN: 1573-5060
    Keywords: TMV ; RAPD ; gene-tagging ; tomato
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A polymerase chain reaction (PCR)-based co-dominant marker was developed which is tightly linked to Tm22. This dominant locus confers resistance to ToMV in tomato. Random-amplified-polymorphic DNA (RAPD) screening was carried out with DNA from ToMV-susceptible and resistant tomato near-isogenic lines. A polymorphic band linked to ToMV resistance was observed. The polymorphic fragment was cloned and the DNA sequences of both ends determined. Specific PCR primers were designed from these sequences. PCR amplification with the specific primers resulted in an amplified band (SCAR) in both susceptible and resistant tomato lines. The amplified band from the susceptible lines could, however, be discerned from that of the resistant ones after cleavage with the restriction enzyme Hind III. In an F2 population of 90, the polymorphic markers co-segregated with susceptibility or resistance, as determined by biological assays for ToMV resistance. The reported SCAR marker is linked to ToMV resistance not only in cultivars derived from American lineage, but also from European lineage. This method enables the distinction of homozygous and heterozygous individual plants in segregating populations, and provides a convenient and rapid assay for both selection and quality control during breeding programs and hybrid seed production, respectively.
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  • 64
    ISSN: 1573-5060
    Keywords: flow sorting ; RAPD ; somatic hybrid ; S. tuberosum ; S. commersonii
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A method for the selection of potato heterokaryons after the electrofusion of dihaploid lines of potato (Solanum tuberosum L.) or dihaploid potato lines and Solanum commersonii has been developed. The selection of heterokaryons was based on the staining of mesophyll protoplasts with fluorescein diacetate or scopoletin. In several experiments, up to 100 000 objects were sorted in a dual laser equipped FACStar Plus, using a 160 μm nozzle and culture medium as sheath fluid. A large number of plants was regenerated from 1 intraspecific and 5 interspecific fusion combinations. RAPD analysis of 244 tetraploid plants from 5 different fusion combinations showed that 240 of these were hybrids. The potential of this selection method for the efficient integration of protoplast fusion technology into potato breeding is discussed.
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  • 65
    ISSN: 1573-3017
    Keywords: mosquito fish ; genotype ; genetic ecotoxicology ; radionuclide contamination ; DNA strand breaks ; RAPD ; allozymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract DNA polymorphism in mosquito fish (Gambusia affinis), as revealed by RAPD (randomly amplified polymorphic DNA) and allozyme analysis, was compared to relative amounts of DNA strand breakage in blood and liver tissues. Mosquito fish were exposed to radionuclide contamination in situ and to X-rays in the laboratory. The types of RAPD metrics used were the number of RAPD bands per individual and the frequency of certain RAPD bands. In a previous study, it was noted that in some instances the number of RAPD bands and the frequency of certain RAPD bands were elevated in radionuclide-contaminated sites relative to reference sites. In the present study, it was found that the median molecular length (MML) of the DNA (which is inversely proportional to the amount of DNA strand breakage) was correlated in several cases to the number of RAPD bands per individual. In addition, for those RAPD bands that occurred at a higher frequency in mosquito fish from radionuclide-contaminated sites, DNA strand breakage was often lower for those fish with than without these RAPD bands. RAPD data obtained on mosquito fish exposed to X-rays in the laboratory paralleled those from the field. Furthermore, analysis showed that heterozygotes for the allozyme locus nucleoside phosphorylase were more prevalent in radionuclide-contaminated sites and had fewer DNA strand breaks than did homozygotes. These results provide additional evidence that changes in population genetic structure of mosquito fish exposed to a genotoxicant (radiation) can be detected at the DNA level.
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  • 66
    ISSN: 1573-0972
    Keywords: V. parahaemolyticus ; plasmid ; RAPD ; cockles (Anadara granosa)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A total of 35 Kanagawa-negative strains of Vibrio parahaemolyticus isolated from cockles (Anadara granosa) were investigated by randomly amplified polymorphic DNA fingerprinting with three primers and their plasmid profiles. Eighteen strains carried small plasmid(s) of 2.4 to 7.3kb that enabled the V. parahaemolyticus to be grouped into eight plasmid patterns. The three primers generated polymorphisms in all 35 strains of V. parahaemolyticus tested, producing bands ranging from 0.25 to 3.9kb. The RAPD profiles revealed a high level of DNA sequence diversity within the Vibrio parahaemolyticus strains tested, and that cockles in the study area are populated by genetically polymorphic strains of V. parahaemolyticus.
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  • 67
    ISSN: 1573-8469
    Keywords: barley ; resistance elicitor ; cultivar mixtures ; complexity ; diversity ; RAPD ; virulence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Powdery mildew populations were analysed to determine the effects of a resistance elicitor and cultivar mixtures on genetic complexity and diversity. Isolations were made from a range of spring barley monocultures and mixtures in a field trial, and characterised for virulence and RAPD profile. In a second trial, isolates were taken from a single mixture from untreated and resistance elicitor-treated areas and from the components of the mixture in monoculture. The mildew population was not only highly heterogeneous for virulence characteristics, but also proved heterogeneous within pathotypes for molecular markers, indicating the major impact of sexual recombination on population structure and the lack of clonal dominance. Various diversity measurements were compared and the value of dissimilarity measurement for revealing genetic distance within a population was highlighted. There was a trend towards increasing complexity as the season progressed, but there was no consistent relationship between cultivar or mixture, disease control treatment, fertiliser treatment, replicate or position in trial, and pathogen genotype. Whilst the resistance elicitor did reduce mildew by 78% in the first trial, and there was no interaction with fertiliser level in its expression, control was substantially less in the second trial. There were no differences between mildew isolates from elicitor and control treatments. It was felt that more effective and consistent resistance elicitors need to be developed before it can be stated that they are unlikely to be eroded by selecting resistant or adapted mildew genotypes.
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