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Brian W. Trader, Richard E. Veilleux, and Holly L. Scoggins

The genus Astilbe (Saxifragaceae) comprises about 13 species and is ranked consistently among the top 10 landscape perennials. Through extensive hybridization, selection and marketing, the lineage of many Astilbehas been lost. Subdioecious Astilbebiternatais the only species in the genus native to North America while other members of the genus are endemic to Asia and monoecious. Due to the unusual geographic distribution of the species and the variation in floral development among them, development of genetic markers using single nucleotide polymorphisms (SNPs) would confirm phylogenetic relationships and establish lineage within the genus. Astilbespecies, hybrids, and cultivars were obtained from plant nurseries and botanical gardens across the country. To elucidate relationships among the genus, we conducted phylogenetic analysis of DNA sequences of the chloroplast gene matKand the internal transcriber spacer (ITS) of ribosomal rDNA genes. DNA was extracted, and gene primers trnK3914 and trnK2R were used to amplify matK, and primers 1406F and ITS2 were used to amplify the ITS1 region between 18S and 5.8S ribosomal DNA units. Both matKand ITS were sequenced for each plant specimen and sequences were aligned to identify nucleotide diversity and detect SNPs. Variation in nucleotide sequence for either gene yielded similar dendrograms. Nucleotide variation among the Astilbeutilized in this study has allowed the development of SNP markers that may be useful for fingerprinting unknown hybrids or cultivars in the industry, and may be used for species alignment within the genus.

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H. Budak, R.C. Shearman, R.E. Gaussoin, and I. Dweikat

A simple marker technique called sequence-related amplified polymorphism (SRAP) provides a useful tool for estimation of genetic diversity and phenetic relationships in natural and domesticated populations. Previous studies and our initial screen showed SRAP is highly polymorphic and more informative when compared to AFLP, RAPD and SSR markers. In this study, applicability of the SRAP markers to obtain an overview of genetic diversity and phenetic relationships present among cool-season (C3) and warm-season (C4) turfgrass species and their relationship with other Gramineae species were tested. Phenetic trees based on genetic similarities (UPGMA, N-J) were consistent with known taxonomic relationships. In some cases, well-supported relationships as well as evidence by genetic reticulation could be inferred. There was widespread genetic variation among C3 and C4 turfgrass species. In Dice based cophenetic matrix, genetic similarities among all species studied ranged from 0.08 to 0.94, whereas in Jaccard based cophenetic matrix, genetic similarities ranged from 0.05 to 0.85. C3 and C4 species were clearly distinguishable and a close relationship between italian ryegrass and tall fescue were obtained based on SRAP. Genome structures of turfgrasses are comparable to other Gramineae species. This research indicates that the SRAP markers are useful for estimating genetic relationships in a wide range of turfgrass species. The SRAP markers identified in this study can provide a useful reference for future turfgrass breeding efforts.

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Sudheer Beedanagari* and Patrick Conner

Pecan, [Carya illinoinensis (Wangenh.) C. Koch], is a member of Juglandaceae family and is one of the most important nut crops produced in the United States. The objective of this study is to generate the first genetic linkage maps for pecan. Maps were constructed for the cultivars `Elliot' and `Pawnee' using the double pseudo-testcross mapping method whereby a separate linkage map is made for each parent using markers heterozygous in that parent. First generation maps consisted primarily of randomly amplified polymorphic DNA (RAPD) markers. We have now used fluorescently labeled amplified fragment length polymorphism (AFLP) markers to produce more complete maps. In the development of the AFLP markers, 64 primer combinations were originally screened to find the most informative combinations. Ten primer combinations were then chosen to produce markers for the maps. The maps currently consist of approximately 100 RAPD and 100 AFLP markers on each cultivar map. `Pawnee' is a high quality commercial pecan cultivar with a very early ripening date. `Elliot' possesses high levels of resistance to pecan scab, caused by the fungus Cladosporium caryigenum. The maps will be used to find markers linked to scab resistance genes and other traits of interest to the breeding program.

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N. Xiang, Y. Hong, and L.T. Lam-Chan

Intensive breeding activities of tropical orchids have given rise to many hybrids, among which genetic relationships are difficult to evaluate due to free interbreeding of different species in the same genus or even from different genera, the use of hybrids for further breeding, use of abbreviated or trade names and sometimes intentional non-disclosure of parentage for commercial considerations. We have subjected 43 popular commercial Dendrobium hybrids to fluorescence amplified length polymorphism (AFLP) analysis and their genetic relationship was estimated. The hybrids bearing flowers of similar shapes and colors were clustered into five groups. Each hybrid tested had a distinct AFLP fingerprint profile except the tissue culture mutants. Sibling hybrids were closely clustered (with genetic distance <0.09) followed by those sharing one parent. These results suggest that AFLP fingerprint profiling gives accurate and objective estimation of genetic relationship of the Dendrobium hybrids tested. Our study also found that the AFLP fingerprint profiles were uniform in different parts of tested plants, stable among individuals in vegetatively propagated populations throughout different growth periods. We conclude that AFLP fingerprint profiling has the potential to be an integral part of current new plant varieties protection sytems.

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Jernej Jakse, William Martin, John McCallum, and Michael J. Havey

The commercial production of onion (Allium cepa L.) inbreds, hybrids, and open-pollinated (OP) cultivars would benefit from a robust set of molecular markers that confidently distinguish among elite germplasms. Large-scale DNA sequencing has revealed that single nucleotide polymorphisms (SNPs), short insertion-deletion (indel) events, and simple sequence repeats (SSRs) are relatively abundant classes of codominant DNA markers. We identified 398 SNPs, indels, and SSRs among 35 elite onion ulations and observed that all populations could be distinguished. Phylogenetic analyses of simple-matching and Jaccard's coefficients for SSRs produced essentially identical trees and relationships were consistent with known pedigrees and previous marker evaluations. The SSRs revealed that elite germplasms from specific companies or breeding programs were often closely related. In contrast, phylogenetic analyses of SNPs and indels did not reveal clear relationships among elite onion populations and there was no agreement among trees generated using SNPs and indels vs. SSRs. This discrepancy was likely due to SNPs and indels occurring among amplicons from duplicated regions (paralogs) of the onion genome. Nevertheless, these PCR-based markers will be useful in the quality control of inbred, hybrid, and OP onion seed lots.

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Wichan Eiadthong, Keizo Yonemori, Shinya Kanzaki, Akira Sugiura, Naoki Utsunomiya, and Suranant Subhadrabandhu

The phylogenetic relationships among 14 Mangifera L. species including three economically important species, i.e., common mango (M. indica L.), horse mango (M. foetida Lour.) and kwini (M. odorata Griff.), were analyzed by comparing 217 amplified fragment length polymorphism (AFLP) markers. The unweighted pair grouping method using arithmetic averages (UPGMA) and neighbor-joining (NJ) method were used and two outgroup taxa, cashew nut (Anacardium occidentale L.) and gandaria (Bouea macrophylla Griff.), were added to both analyses. The common mango was closely related to banana mango (M. sylvatica Roxb.), M. laurina Bl., and M. oblongifolia Hook.f. Intraspecific variation among seven cultivars of common mango was much smaller than interspecific variation and these cultivars were classified into one M. indica group using both methods. Mangifera macrocarpa Bl., M. foetida, and M. odorata were also related to M. indica in both UPGMA and NJ trees, although these three species are classified into a different subgenus (subgenus Limus) from the subgenus Mangifera to which M. indica belongs. Also, in both UPGMA and NJ trees, M. gedebe Miq. and M. griffithii Hk.f. were placed in distant positions among the Mangifera species tested, indicating these two species are related distantly to M. indica. The AFLP technique was confirmed to be useful for phylogenetic analysis.

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Salih Kafkas, Hakan Ozkan, and Mehmet Sutyemez

Turkey has more than 4 million walnut trees (Juglans regia L.), most of which are derived from seedlings, and are nongrafted trees. This characteristic leads to a huge opportunity to select superior walnut genotypes from natural populations for cultivation and for breeding programs. Several selection studies have been performed in the last decades and few genotypes were selected. The goal of this study was to characterize and determine genetic relationships among 21 walnut genotypes with potential in walnut production using amplified fragment length polymorphism (AFLP) and selective amplification of microsatellite polymorphic loci (SAMPL) techniques. Eight primer combinations (six for AFLP and two for SAMPL) were applied to 21 walnut genotypes and a total of 230 bands of which 50.4% of them were polymorphic were obtained. The SAMPL technique was more effective than AFLP in the separation of very closely related genotypes. Genotypes of the pairs `Maras-18' with `Maras-46', `KSU-5' with `Sutyemez-1', `Maras-12' with `Sutyemez-2,' `Kaman-3' with `Kaman-4', and `KSU-11' with `Maras-10' were the most closely related.

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Jiahua Xie, Todd C. Wehner, and Mark A. Conkling

Combining the use of PCR and single-strand conformation polymorphisms (SSCP), nine sequences from the cucumber genome were successfully identified and cloned that encoded two well-conserved asparagine-proline-alanine (NPA) domain homologues to aquaporin genes. The sensitivity and detection efficiency of SSCP and restriction enzyme analysis for detecting DNA sequence variation were evaluated using similar-sized DNA fragments. The SSCP analysis was more sensitive and efficient for discriminating different clones than restriction enzyme analysis, although some sequence variation inside similar-sized DNA fragments could be identified by restriction analysis. Consideration of the results of SSCP analysis with DNA sequence information indicated that one or two base pair changes in the amplified regions could be detected. Moreover, the SSCP analysis results of genomic DNA PCR products that were amplified by degenerate primers can provide rough information about the number of member genes. If the SSCP bands of a cloned fragment (such as CRB7) did not have the corresponding bands from genomic DNA PCR products, that fragment might be a misamplified product. The PCR-based SSCP method with degenerate oligonucleotide primers should facilitate the cloning of member genes.

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He Li, Cheng-Jiang Ruan, Li Wang, Jian Ding, and Xing-Jun Tian

distinguish among sea buckthorn cultivars, e.g., amplified fragment length polymorphism, random amplified polymorphic DNA, intersimple sequence repeat, and sequence-related amplified polymorphism ( Bartish et al., 2001 ; Li et al., 2009 , 2010 ; Ruan, 2006

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Peter J. Leonard, Mark H. Brand, Bryan A. Connolly, and Samuel G. Obae

amplified fragment length polymorphism analysis. The intergeneric, F 1 hybrid species × Sorbaronia dippelii (Zabel) C. K. Schneid. ( A. melanocarpa × S. aria ), × S. alpina (Willd.) C. K. Schneid. ( A. arbutifolia × S. aria ), and × S. fallax ( A