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Richard G. Novy and Nicholi Vorsa

Cranberry (Vaccinium macrocarpon Ait.) has few qualitative, morphological characteristics that can be used to reliably distinguish among cultivars. Fifty-two silver-stained random amplified polymorphic DNAs (ssRAPDs) were used to assess genetic heterogeneity and relatedness within accessions of four major cranberry cultivars (`Early Black', `Howes', `McFarlin', and `Searles'). Rather than being represented by one genotype, as might be expected in an asexually propagated crop, each cultivar was represented by multiple genotypes, which in many cases did not appear to be closely related to one another. The intracultivar heterogeneity was often so extreme that clonal representatives of a cultivar would group with representatives of other cultivars following cluster analysis. Of the total ssRAPD variation, 9.7% could be attributed to variation among the four cultivar groups and 90.3% to variation within the cultivars. `Howes' was the only cultivar in which a consensus DNA fingerprint among regional representatives could be identified.

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Samir Mhameed, Dror Sharon, Jossi Hillel, Emanuel Lahav, Daniel Kaufman, and Uri Lavi

To estimate heterozygosity level in the avocado (Persea americana Mill.) genome, two types of variable number of tandem repeat (VNTR) markers were used. Multilocus DNA fingerprints (DFPs) were analyzed on avocado progeny resulting from either crosses or selfing of cultivars. In five crosses, heterozygosity was 100%, while in two self-pollinated families, heterozygosity was 90% and 94%. Single locus, simple sequence repeat (SSR) DNA markers were analyzed by typing 59 loci on five avocado cultivars. Average heterozygosity varied from 0.50 to 0.66, while gene diversity varied from 0.42 to 0.66. Heterozygosity varied from 38% to 70%. The percentage of fragments that exhibited Mendelian inheritance was 62.5% to 85% (P < 0.05) for the DFP fragments and 85% for the SSR alleles.

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Myneni Aruna, Max E. Austin, and Peggy Ozias-Akins

Cultivars of the economically important rabbiteye blueberry (Vaccinium ashei Reade) were differentiated at the DNA level using the technique of randomly amplified polymorphic DNA. Single decanucleotide primers of arbitrary sequence were used to amplify genomic DNA by the polymerase chain reaction. All cultivars tested exhibited a unique set of collective amplified fragments of distinct molecular weight. A blind fingerprinting experiment resulted in identification of unknown samples without ambiguity. We also clarified the genetic identity of two wild selections of rabbiteye blueberry, `Ethel' and `Satilla', which have been maintained as two different selections, hut are considered by some blueberry breeders to be of the same genetic constitution. The technique also verified the probable identity of two cultivars in a commercial blueberry field by comparing their amplified DNA patterns with those of standard cultivars. No variation was observed between the amplification profiles of `Brightwell' and its presumed sport. A cultivar key based on 11 markers amplified by four primers is presented.

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Dror Sharon, Avital Adato, Samir Mhameed, Uri Lavi, Jossi Hillel, Maria Gomolka, Conny Epplen, and Jorg Thomas Epplen

Plant genomes contain polymorphic repetitive sequences that can be used as DNA markers. Minisatellites (16 to 64 bp per repeat) and simple-sequence repeats (2 to 6 bp per repeat) are the most polymorphic markers found in plant and animal genomes. In this study, the hybridizations between genomic DNA and variable number of tandem repeat probes were examined in Arabidopsis thaliana L. (Heynn), onion (Allium cepa L.), tomato (Lycopersicon esculentum L.), wheat (Triticum aestivum L.), avocado (Persea americana Mill.), litchi (Chinensis Sonn.), mango (Mangifera indica L.), and Carica species. Some of the probes detected polymorphic sequences in all the species, but others were useful only for one or two species. None of the probes gave clear band patterns in either onion or wheat. The in-gel hybridization method was similar to Southern blot hybridization using the simple-sequence repeat probes.

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Warren F. Lamboy and Christopher G. Alpha

The USDA-ARS Vitis genetic resources collections in Geneva, N.Y., and Davis, Calif., contain ≈3600 accessions of >35 species. Accurate and unambiguous identification of these grapes is essential for efficient and effective use of this germplasm. Previous workers have successfully used polymerase chain reaction (PCR)-generated SSRs to fingerprint cultivars of the wine and table grape species, V. vinifera. Building on this work, we conducted a test of five previously characterized SSR loci on 110 accessions of 25 grape taxa (21 Vitis species and 4 hybrids) to determine if they would satisfy our need for identifying cultivars within the USDA-ARS grape collections. Scorable SSR fragments were produced with all 550 primer-accession combinations, with no null loci observed. The loci were highly polymorphic, with 16 to 38 different alleles found at a locus. Heterozygosity values ranged from 0.464 to 0.818, while gene diversity values ranged from 0.875 to 0.955. Discrimination power at a locus varied from a low of 0.947 to a high of 0.987. Combined discrimination power of all loci was effectively 1.000, with 2 chances in 100,000,000 that two sexually, independently derived grape accessions would not be distinguishable using this set of five SSR loci. Two plants in the study that had previously been classified as belonging to different grape species were shown to have identical SSR fingerprints, showing that they almost certainly possessed the same genotype. Because SSR markers are codominant and highly polymorphic and SSR loci are generally conserved across a range of related species, we strongly recommend SSRs for fingerprinting not only grape, but other clonal genetic resources collections as well.

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Margaret R. Pooler and Ruth L. Dix

The historic Japanese flowering cherry trees planted around the Tidal Basin in Washington, D.C., were given to the United States in 1912 as a gift from Japan, and have become a popular tourist attraction. Unfortunately, only a small portion of the original trees remain, and these trees are in various states of decline due to old age and stress. In cooperation with the National Park Service, we have propagated from cuttings nine trees that are known to be original and 10 trees that are thought to be original. DNA from these and other P. × yedoensis were compared using RAPD markers. Twenty-one 10-nucleotide primers yielded 80 repeatable bands that were used to assess genetic distance among the accessions. Twenty of these bands were monomorphic across all 28 accessions tested, so were not informative. The frequency of the remaining 60 bands varied from 0.04 to 0.96, with an average frequency of 0.58. Thirteen of the accessions, including six of the nine that are known to be original germplasm, were identical at all loci tested. Other accessions that are thought to be original trees were similar, with similarity values of 0.93 to 0.99. The most genetically dissimilar trees were P. × yedoensis accessions from our collection that were collected as seed in Japan. Accessions obtained from commercial nurseries including `Afterglow', `Akebono” and Yoshino were also dissimilar to the Tidal Basin trees. This study indicates that most of the older trees planted around the Tidal Basin are genetically very similar, but that variability in P. × yedoensis exists, especially in accessions collected as seed from Japan.

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Noémi Makovics-Zsohár, Magdolna Tóth, Dezső Surányi, Szilvia Kovács, Attila Hegedűs, and Júlia Halász

The hexaploid European plum (Prunus domestica L.) is an economically important fruit species with limited information on its genetic structure. Our objective was to fingerprint 55 cultivars using seven simple sequence repeat (SSR) markers to estimate the polymorphism level and determine allelic variation and genetic relationships among local and international cultivars. The primer pairs amplified a total of 135 alleles ranging from six to 27 alleles per locus, displaying high polymorphism. All genotypes were clearly distinguished with the seven SSRs used in this study. In a neighbor-joining cluster analysis, cultivars belonging to the same species did not group together. Foreign modern cultivars clustered together, and Hungarian landraces positioned distantly from those. STRUCTURE analysis indicated three genetically distinct groups of the studied genotypes. Each cluster of Hungarian landrace cultivars received strong bootstrap support (89% to 100%). Most genotypes kept under identical name showed different DNA fingerprints. A principal component analysis (PCA) confirmed the information provided by the dendrogram and clarified the origin of ʻFehérszilva’. Our results confirmed the potential of the application of SSR markers in plum breeding.

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M. Hubbard, J. Kelly, S. Rajapakse, A. Abbott, and R. Ballard

We have identified cloned rose DNA fragments that detect restriction fragment length polymorphisms (RFLP) in rose (Rosa ×hybrida) cultivars. RFLP can be used as genetic markers for identification, certification, and patent protection. By comparing RFLP patterns for each of six probes, we have been able to characterize eight cultivars. These results confirm that RFLP analyses are useful for rose cultivar identification and may provide a means for protecting patent rights to new cultivars.

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Zhen-Xiang Lu, G.L. Reighard, W.V. Baird, A.G. Abbott, and S. Rajapakse

Eighteen peach rootstock cultivars, most of Prunus persica (L.) Batsch, were screened for diagnostic random amplified polymorphic DNA (RAPD) markers using synthetic decamer oligonucleotide primers. Twenty of the 80 primers were informative, and 40 amplified DNA bands from the informative primers were selected as RAPD markers. Based on combined banding patterns, all 18 rootstock cultivars were identified with only six of the 20 informative primers. Cluster analysis of the 18 peach rootstock cultivars using 40 RAPD markers produced a dendrogram of genetic relatedness in good agreement with their putative pedigrees. The first major bifurcation in the dendrogram divided these rootstock cultivars into two groups according to their resistance or susceptibility to root-knot nematodes [Meloidogyne incognita (Kofoid and White) Chitwood and M. javanica (Treub) Chitwood].

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Hilde Nybom, Susan Gardiner, and Charles J. Simon

Individual-specific DNA fragment patterns were obtained by hybridization of endonuclease-digested apple (Malus ×domestica Borkh.) DNA with a probe (pAR72) derived from the rDNA spacer region of the `White Angel' crab apple. Fragment detection was carried out with a nonradioactive method, using a horseradish peroxidase-catalyzed luminol oxidation. Paternity could be inferred by comparison of the fragment pattern generated by a seedling with those derived from putative parents.