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R.L. Jarret and K.V. Bhat

Fifty-seven accessions of Musa including cultivated clones Of 6 genomic groups (AA, AB, AAA, AAB, ABB, ABBB), M. balbisiana (BB), M. acuminata ssp. banksii (AA), M. acuminata ssp. malaccensis (AA) and M. velutina were examined for random amplified polymorphic DNA (RAPD) genetic markers using PCR with sixty 10-mer random primers. Forty-nine of 60 tested primers gave reproducible DNA amplification patterns. The number of bands resolved per amplification was primer dependent and varied from 1 to a maximum of 24. The size range of the amolification products also differed with the select& primer sequence/genotype and ranged from 0.29 to 3.0 kb. RAPD data were used to generate Jaccard's similarity coefficients which were analyzed phenetically. Phenetic analysis separated clones into distinct groupings that were in agreement with clusterings revealed when data were subsequently analyzed by principal coordinate analysis (PCO). In both the phenetic and the PCO analyses, previously unclassified cultivars grouped with cultivars previously classified for their genomic group based on morphological keys. The implications of RAPD analysis for Musa germplasm classification, clonal identification, and management are discussed.

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R.L. Jarret, N. Gawel, and A. Whittemore

Twenty-four accessions of Ipomoea, representing 13 species of section Batatas and the outgroup species I. gracilis and I. pes-caprae were analyzed for restriction fragment length polymorphisms. Polymorphisms were detected by probing Southern blots of restriction enzyme-digested genomic DNA with 20 low or moderate copy number sequences isolated from an I. batatas cv. Georgia Red genomic library. Data were analyzed cladistically and phenetically. Ipomoea trifida, I. tabascana, and collection K233 are, of the materials examined, the most closely related to sweetpotato (I. batatas). Ipomoea littoralis, the only Old World species in the section, is a sister species to I. tiliacea. Ipomoea littoralis, I. umbraticola, I. peruviana, I. cynanchifolia, and I. gracilis are shown to be diploid (2n = 2x = 30). In contrast, I. tabascana is tetraploid (2n = 4x = 60). The intrasectional relationships of section Batatas species and the role of tetraploid related species in the evolution of the cultivated I. batatas are discussed.

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R.L. Jarret, G. Lovell, and M. Spinks

The S-9 Plant Germplasm Collection maintains and distributes germplasm of various horticultural crops, including pepper (Capsicum spp.), watermelon (Citrullus lanatus), okra (Abelmoschus spp.), eggplant (Solanum melongena), miscellaneous Solanum spp., sweetpotato (Ipomoea batatas spp.), luffa (Luffa spp.), gourds (Lagenaria and Momordica spp.), squash (Curcurbita moschata), pumpkin (Curcurbita maxima), marigold (Tagetes spp.), Stokes' aster (Stokesia laevis), hibiscus (Hibiscus spp.), Engelman daisy (Engelmannia pinnatifolia), pampasgrass (Cortaderia selloana), ornamental bamboo (Bambusa spp.), and other ornamental grasses. Seed or other propagules of these plant materials are available for research purposes. Detailed information on individual collections and general information on the USDA National Plant Germplasm System will be presented.

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Diego Fajardo, Don R. La Bonte, and Robert L. Jarret

The USDA gene bank currently maintains 668 accessions of cultivated sweetpotato and 219 accessions of related Ipomoea species. Information on the genetic diversity of the collection does not exist due to funding constraints. The development of a core collection would provide a subset of accessions that represent the genetic diversity of the main collection with a minimum of repetitiveness. The small size of the core collection would facilitate the evaluation of the accessions for economically important traits. The objective of this research is to develop a core collection of Papua New Guinea sweetpotato germplasm using the Amplified Fragment Length Polymorphisms (AFLPs) marker system. This approach to quantifying genetic diversity would later serve as a model for the development of a USDA sweetpotato germplasm core collection. The germplasm choosen for this study was collected from this crop's secondary center of genetic diversity based on its potential as a source of new traits. All genotypes were fingerprinted using four primer combinations that generated 224 markers. The molecular data was then analyzed using NTSYSpc 2.0 program to determine the relatedness of the genotypes. The molecular analysis showed a homogeneous genetic constitution. The extent of diversity among accessions was correlated with the geographic origin of the plant material.

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Diego Fajardo, Don R. La Bonte, and Robert L. Jarret

The USDA gene bank currently maintains 668 accessions of cultivated sweetpotato and 219 accessions of related Ipomoea species. Information on the genetic diversity of the collection does not exist due to funding constraints. The development of a core collection would provide a subset of accessions that represent the genetic diversity of the main collection with a minimum of repetitiveness. The small size of the core collection would facilitate the evaluation of the accessions for economically important traits. The objective of this research is to develop a core collection of Papua New Guinea sweetpotato germplasm using the Amplified Fragment Length Polymorphisms (AFLPs) marker system. This approach to quantifying genetic diversity would later serve as a model for the development of a USDA sweetpotato germplasm core collection. The germplasm choosen for this study was collected from this crop's secondary center of genetic diversity based on its potential as a source of new traits. All genotypes were fingerprinted using four primer combinations that generated 224 markers. The molecular data was then analyzed using NTSYSpc 2.0 program to determine the relatedness of the genotypes. The molecular analysis showed a homogeneous genetic constitution. The extent of diversity among accessions was correlated with the geographic origin of the plant material.

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Mario I. Buteler, Don R. LaBonte, and Robert L. Jarret

Microsatellites or simple sequence repeats (SSRs) were used to characterize 20 sweetpotato genotypes and to assign paternity for offspring from crosses among them. The PCR amplifications were performed with each of the sweetpotato genotypes and primers flanking a SSR loci previously characterized with the varieties Beauregard and Excel and 20 offspring from a cross among them. The PCR reaction products were separated in nondenaturing 12% acrylamide gels run at 25 V·cm–1 for 5 hours, and DNA fragments were visualized with silver staining. Gels were scanned on a flat bed scanner and analyzed using the Pro-RFLP software package. Three primer pairs were sufficient to produce an allelic profile capable of differentiating the 20 genotypes from each other. More than seven alleles/loci were found using each of the three primer pairs assayed. Occasionally primers produced allelic products clearly localized in two or three regions of the gel. These multiple loci segregated independently in a diploid fashion. This evidence suggests that there is not total homology among the three sweetpotato genomes.

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R.L. Jarret, N. Bowen, S. Kresovich, and Z. Liu

Simple sequence repeats (SSRs) were isolated from a size-fractionated genomic DNA library of sweetpotato [Ipomoea batatas (L.) Lam.]. Screening of the library with five oligonucleotide probes, including; (GT)11, (AT)11, (CT)11, (GC)11, and (TAA)8, detected the occurrence of 142 positive colonies among ≈12,000 recombinants. Automated DNA sequencing revealed the presence of simple, compound, perfect, and imperfect SSRs. Five homologous PCR primer pairs were synthesized commercially and used to screen 30 sweetpotato clones for the occurrence of SSR polymorphisms. All primer pairs produced an amplification product of the expected size and detected polymorphisms among the genotypes examined. The potential for the use of SSRs as genetic markers for sweetpotato germplasm characterization is discussed.

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A.Q. Villordon, C.A. Clark, R.A. Valverde, R.L. Jarret, and D.R. LaBonte

Previous work by our group has detected the presence of a heterogeneous population of Ty1-copia-like reverse transcriptase retrotransposon sequences in the sweetpotato genome. Recently, we detected the presence of putatively active Ty1-copia-like reverse transcriptase sequences from a virus-infected `Beauregard' sweetpotato clone. In the current study, we report the differential detection of putatively stress-activated sequences in clones from seedling 91-189. The clones were infected with different combinations of virus isolates followed by extraction of leaf RNA samples at three sampling dates (weeks 2, 4, and 6) after inoculation. After repeated DNAse treatments to eliminate contaminating DNA, the RNA samples were subjected to first strand cDNA synthesis using random decamer primers followed by PCR analysis utilizing Ty1-copia reverse transcriptase-specific primers. Through this approach, we detected amplified fragments within the expected size range (280-300 bp) from clones infected with isolates of sweetpotato leaf curl (SPLC) and feathery mottle viruses (FMV) (week 2 and 6) and FMV (week 4). We were unable to detect PCR products from the noninfected clones or the other infected samples. The data suggests that specific viruses may be involved in the expression of these Ty1-copia-related reverse transcriptase sequences. It also appears that sampling at various dates is necessary to detect putative activity over time. This preliminary information is essential before proceeding to the construction and screening of cDNA libraries to isolate and fully characterize the putatively active sweetpotato Ty1-copia-like retrotransposon sequences. Through the partial or complete characterization of sweetpotato Ty1-copia elements, sequences that correspond to cis-regulatory element(s) can be identified and further studied for their roles in responding to specific stress factors.

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M.I. Buteler, D.R. LaBonte, R.L. Jarret, and R.E. Macchiavelli

Using codominant molecular markers (microsatellites) for paternity identification was investigated in hexaploid sweetpotato [Ipomoea batatas (L.) Lam.]. Two experimental populations (CIP and LAES), each consisting of progeny of known parentage, were scored for the presence or absence of alleles segregating at IB-316 and IB-318 microsatellite loci. Paternity was assessed using paternity exclusion and the most-likely parent methods. In the former, paternity is assigned based on the identification of incompatible parent-progeny marker data. In contrast, the latter method incorporates paternity exclusion and a log-likelihood or LOD score that weighs progeny allelic patterns as to the likelihood that they could have come from a given paternal parent. The number of correctly allocated progeny differed for the methods. Paternity exclusion correctly allocated 7% and 25% of the progeny in the LAES and CIP populations, respectively. The most-likely parent method correctly allocated 23% and 88% of the progeny in the LAES and CIP populations, respectively. The greater misassignments in the LAES population were attributed to low allelic diversity at the LAES IB-318 locus and a larger parental population. This study demonstrates the feasibility of identifying paternity in sweetpotato using a minimal number of loci.

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R.L. Jarret, S. Kresovich, T. Holms, Janelle Evans, and Z. Liu

Simple sequence repeats (SSRs) were isolated from a size-fractionated genomic DNA library of watermelon (Citrullus lanatus L. cv. New Hampshire Midget). Screening of the library with five oligonucleotide probes, including (GT)11, (AT)11, (CT)11, (GC)11, and (TAA)8, detected the occurrence of 96 positive colonies among ≈8000 recombinants. Automated DNA sequencing revealed the presence of SSRs. PCR primer pairs homologous to the regions flanking the SSR loci were synthesized commercially and used to screen 56 watermelon genotypes for the occurrence of SSR polymorphisms. Amplification products were separated using nondenaturing PAGE. Eighty percent of the primer pairs produced amplification products of the expected size and detected polymorphisms among the genotypes examined. The use of SSRs for watermelon germplasm characterization is discussed.