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Eric Stafne, John Clark, and Kim Lewers

Molecular markers have been used previously to identify linkages to important traits of interest. In this study two marker types, randomly amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR), were used to find molecular markers linked to two morphological traits in blackberry (Rubus L. subgenus Rubus). Thorniness and floricane fruiting are both qualitative, recessive traits that are inherited tetrasomically. A cross of `Prime-Jim'® × `Arapaho' was made to create a population that segregated for the two traits. A random sample of 98 plants from a population of 200 were assayed to find molecular markers that co-segregate with the two traits. Three putative markers were identified for the floricane fruiting trait (two SSRs and one RAPD; χ2 = 4.09 to 9.99, P < 0.001 to 0.043). Five potential RAPD markers were found for the thorny trait (χ2 = 3.88 to 10.23, P < 0.001 to 0.048). Identification of markers linked to these traits could potentially be useful in marker-assisted selection.

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Zhi-Hong Gao, Zhi-Jun Shen, Zhen-Hai Han, Jing-Gui Fang, Yu-Ming Zhang, and Zhen Zhang

Sequencing amplification fragments produced using simple-sequence repeat (SSR) primer pairs pchgms2 and UDP96008 in `Dayezhugan' japanese apricot showed that SSRs obtained included a microsatellite locus originally identified in peach. The microsatellite sequence homogeneity between UDP96008 in japanese apricot in this study and UDP96008 in the peach in GenBank was 98%. Twenty-four japanese apricot genotypes originating in diverse geographic areas had been identified with 14 SSR primer pairs developed in different species of Prunus. In total, 129 alleles were obtained and per primer pairs detected 2.5 alleles on the average. The results from cluster analysis showed that the genetic distance between `Nanhong' and `Zhonghong' was the closest, and cultivars from China and from Japan could not be separated completely.

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Warren Lamboy, Christopher Alpha, Amy Szewc-McFadden, and Sharon Bleik

The cold-hardy Vitis (grape) collection at the USDA/ARS Plant Genetic Resources Unit in Geneva, N.Y., comprises ≈1300 accessions. While much of the collection has been evaluated for morphological and viticultural traits, little of it has been well-characterized genetically. Lack of genetic information hampers the identification of accessions, the determination of genetic relationships among them, the evaluation of potential new accessions, and the construction of a core subset of the collection. Because simple sequence repeat DNA polymorphisms (SSRs or microsatellites) have already been proven to be useful genetic markers in Vitis vinifera (non-cold-hardy wine, raisin, and table grapes), our research focuses on the use of the markers both for the identification (“fingerprinting”) of species, hybrids, subspecies, cultivars (varieties), and accessions of cold-hardy Vitis, and for the determination of genetic relationships between these taxa. Our latest research results in this area will be presented.

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Hongmei Ma and Margaret Pooler

Ornamental flowering cherry trees (Prunus species) are popular landscape plants that are used in residential and commercial landscapes throughout most temperate regions of the world. Most of the flowering cherry trees planted in the United States represent relatively few species. The U.S. National Arboretum has an ongoing breeding program aimed at broadening this base by developing new cultivars of ornamental cherry with disease and pest resistance, tolerance to environmental stresses, and superior ornamental characteristics. Knowledge of the genetic relationships among species would be useful in breeding and germplasm conservation efforts. However, the taxonomy of flowering cherry species and cultivars is complicated by differences in ploidy levels and intercrossing among species. We have used simple sequence repeat (SSR) markers developed for other Prunus species to screen a diverse collection of over 200 ornamental cherry genotypes representing 70 taxa in order to determine the genetic relationships among species, cultivars, and accessions. Data were generated from 9–12 primer pairs using an automated DNA genetic analyzer (ABI3770), and subjected to UPGMA cluster analysis. Extremely high levels of polymorphism were exhibited among the materials studied, thus indicating that ornamental flowering cherry germplasm has substantial inherent genetic diversity. This information, combined with traditional morphological characteristics, will be useful in determining genetic relationships among accessions in our collection and for predicting crossability of taxa.

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A. Levi, C.E. Thomas, T. Trebitsh, A. Salman, J. King, J. Karalius, M. Newman, O.U.K. Reddy, Y. Xu, and X. Zhang

Seventy-one amplified fragment length polymorphism (AFLP), 93 sequence related amplified polymorphism (SRAP), and 14 simple sequence repeat (SSR) markers were used to extend an initial genetic linkage map for watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai]. The initial map was based on 151 randomly amplified polymorphic DNA (RAPD) and 30 and inter-simple sequence repeat (ISSR) markers. A testcross population previously used for mapping of RAPD and ISSR markers was used in this study: {plant accession Griffin 14113 [C. lanatus var. citroide (L.H. Bailey) Mansf.] × the watermelon cultivar New Hampshire Midget (C. lanatus var. lanatus)} × PI 386015 [C. colocynthis (L.) Schrad.]. The linkage map contains 360 DNA markers distributed on 19 linkage groups, and covers a genetic distance of 1976 cM with an average distance of 5.8 cM between two markers. A genomic DNA clone representing 1-amino-cyclopropane-1-carboxylic acid (ACC-) synthase gene, involved in ethylene biosynthesis, was also mapped. As in previous mapping studies for watermelon, a large number of AFLP and SRAP markers were skewed away from the 1:1 segregation ratio, and had to be excluded from the final mapping analysis. The stringent mapping criteria (JoinMap 3.0 mapping program) produced linkage groups with marker order consistent with those reported in previous mapping study for watermelon.

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Kentaro Kitahara, Shogo Matsumoto, Toshiya Yamamoto, Junichi Soejima, Tetsuya Kimura, Hiromitsu Komatsu, and Kazuyuki Abe

We examined the genetic diversity and relatedness among apple (Malus ×domestica Borkh.) cultivars in Japan. The 42 apple cultivars, including major cultivars in Japan, were divided into five groups based on SSR genotypes. Most economically important cultivars belong in three groups: Fuji-Delicious, Golden Delicious, and Jonathan groups, and their genetic backgrounds seemed to be narrow. We also investigated the parent-offspring relationships of nine apple cultivars. `Jonathan', `Fuji', and `Rero 11' were identified as the respective paternal parents of three cultivars described as having unknown paternal parents (i.e., `Akagi', `Ambitious', and `Hokuto'). `Starking Delicious', `Senshu', and `Golden Delicious', rather than `Ralls Janet', `Hatsuaki', and `Indo', seemed to be the paternal parents of `Kinsei', `Kiou', and `Mellow', respectively. `Carolina Red June' was excluded as a paternal parent of `Ranzan'. Both attributed parents of `Scarlet' (`Akane' and `Starking Delicious') were excluded, and it was suggested that `Fuji' was used as either a maternal or a paternal parent of `Scarlet'. `Jonathan' rather than `McIntosh' seems to be a maternal parent of `Yukari'.

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Anna Hale and Mark W. Farnham

Private and public vegetable breeders are interested in using current and emerging PCR-based marker systems in their respective improvement programs. However, before new systems are employed to replace existing ones, the new systems must prove to be efficient and cost-effective alternatives. Sequence related amplified polymorphisms (SRAPs), amplified fragment length polymorphisms (AFLPs), and simple sequence repeats (SSRs) were compared for their ability to differentiate individuals of a diverse group of 24 elite broccoli (Brassica oleracea L. italica) inbreds. Genomic DNA was assayed using 24 AFLP, 24 SRAP, and 44 SSR primer pairs. In this assessment, SSRs produced an average of only two bands per primer, with 25% of these bands being monomorphic, and the remaining bands detecting very few differences among the inbreds. Although the AFLP method resulted in a lower rate (63%) of polymorphism than the SSRs, it produced about 20 bands per primer. SRAPs produced an average of 14 bands per primer, with 82% of these bands being polymorphic. Since AFLP and SRAP markers had a higher multiplex ratio and SSRs were frequently monomorphic, AFLP and SRAPs were more effective in differentiating the elite broccoli inbreds examined in this study. Similarity matrices were generated from the AFLP and SRAP data, and resulting dendographs were compared.

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Warren F. Lamboy and Christopher G. Alpha

Curators of plant genetic resources collections must preserve germplasm possessing known useful characteristics as well as material displaying general genetic diversity. In order to ensure that both types of germplasm are included in a collection, germplasm curators require three fundamental types of information about each accession: taxonomic identity, genetic identity, and genetic relationship. Because simple sequence repeat DNA fragments (SSRs) have been successfully used to determine the genetic identity of grape clones, we conducted a study to determine if SSRs would supply all three types of information for the accessions in the cold-hardy Vitis (grape) germplasm collection. SSR fragments were amplified at six different loci for 23 accessions of cold-hardy grape spanning the range of species diversity in the collection. The minimum number of different alleles found at a locus was 9; the maximum was 26. Heterozygosity values ranged between 0.565 and 0.783, while gene diversity values were in the range 0.785 to 0.944. Two hundred fifty-two pairs of plants out of a possible 253 could be distinguished by their SSR profiles. Nei's genetic identities were computed between all pairs of plants and used in a UPGMA cluster analysis. The relationships obtained did not correspond well to expected relationships based on geography and taxonomy. Four species of grapes were represented by two or more accessions in this study. No DNA fragments found at these six loci served to unambiguously distinguish one species from another. Thus, SSR fragments from the six loci studied were useful in determining genetic identity of accessions, but were not helpful in determining genetic relationships or taxonomic identities. We are searching for additional loci that are informative for these types of information. Meanwhile we highly recommend SSRs for determining genetic identity in germplasm resources collections.

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Minou Hemmat, Norman F. Weeden, Patrick J. Conner, and Susan K. Brown

The columnar mutation `Wijcik McIntosh' has attracted much attention because of its compact growth habit, which is compatible with high-density plantings. Using bulked segregant analysis, we identified several randomly amplified polymorphic DNA (RAPD) markers that displayed a close linkage with the columnar locus (Co). The RAPD marker that displayed the closest linkage was end sequenced to develop a sequence tagged site for rapidly screening segregating populations. A simple sequence repeat (SSR) of (GA)17 was identified within the DNA fragment. Four allelic forms, including an apparent null allele, could be distinguished among the cultivars tested. The null allele displayed close linkage with Co in two progenies, and we used this marker to identify the location of the gene on the apple linkage map.

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Juan J. Ruiz, Santiago García-Martínez, Belén Picó, Muquiang Gao, and Carlos F. Quiros

We studied the genetic variability of some traditional tomato (Lycopersicon esculentum L. Mill.) cultivars of Spain, and established their relationships using both simple sequence repeats (SSR) and sequence related amplified polymorphism (SRAP) markers. These included cultivars from different locations of three main types, Muchamiel, De la pera, and Moruno. Additionally we tested two other local cultivars, `Valenciano' and `Flor de Baladre', plus a small sample of commercial cultivars and a few wild species. Both types of markers resolved the cultivars from different groups, but SSR failed to distinguish some of those classified under the same group. All the De la pera cultivars clustered together by genetic similarity with the SRAP markers. The other traditional cultivars, which are grown in a wider geographic range, formed a more diffuse group, which included the commercial cultivar Roma. The Mexican cultivar Zapotec, a breeding line, and the virus-resistant commercial hybrid `Anastasia' were the most distant of all the cultivars. The latter hybrid had higher similarity to the wild species due to introgressed segments from them carrying the resistance genes. Similar results were observed for SSR markers but with a lower level of resolution. This information would be useful to facilitate tomato germplasm conservation and management efforts.