Characterization of 10 trinucleotide microsatellite loci in the Critically Endangered Pyrenean yam Borderea chouardii (Dioscoreaceae)

The low levels of allozymic variability found in the Critically Endangered Borderea chouardii prompted us to develop microsatellite markers to assess the genetic variability and population structure for the adequate conservation management of this species. A (CTT)_n‐enriched partial genomic library was constructed. Ten polymorphic microsatellite loci were isolated from it, rendering 51 alleles in 47 individuals analysed. The allelic pattern observed for all of the loci with more than two alleles suggests that B. chouardii is tetraploid.     

Dioscorea howardiana,a new species in Dioscorea section Trigonobasis (Dioscoreaceae)

Dioscorea howardiana is proposed as a new species from Mexico and Central America as far south as Panama, and is described and illustrated. It most closely resemblesD. liebmannii. Relationships, distribution, and ecological preferences of the new species are discussed.

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Resumen   Dioscorea howardianase propone como una especie nueva de México y Centroamérica, es descrita e ilustrada. Es semejante aD. liebmannii. Se discuten sus relaciones, su distribución geográfica y preferencias ecológicas.

     

Plastid phylogenetics of Oceania yams (Dioscorea spp., Dioscoreaceae) reveals natural interspecific hybridization of the greater yam (D. alata)

Phylogenetic relationships of Oceanian staple yams (species of Dioscorea section Enantiophyllum) were investigated using plastid trnL‐F and rpl32‐trnL^(UAG) sequences and nine nuclear co‐dominant microsatellites. Analysis of herbarium specimens, used as taxonomic references, allowed the comparison with samples collected in the field. It appears that D. alata, D. transversa and D. hastifolia are closely related species. This study does not support a direct ancestry from D. nummularia to D. alata as previously hypothesized. The dichotomy in D. nummularia previously described by farmers in semi‐perennial and annual types was reflected by molecular markers, but the genetic structure of D. nummularia appears more complex. Dioscorea nummularia displayed two haplotypes, each corresponding to a different genetic group. One, including a D. nummularia voucher from New Guinea, is closer to D. tranversa, D. alata and D. hastifolia and encompasses only semi‐perennial types. The second group is composed of semi‐perennial and annual yams. However, some of these annual yams also displayed D. alata haplotypes. Nuclear markers revealed that some annual yams shared alleles with D. alata and semi‐perennial D. nummularia, suggesting a hybrid origin, which may explain their intermediate morphotypes and the difficulty met in classifying them.     

Two new species of Dioscorea (Dioscoreaceae) from Peru

Two new species ofDioscorea,D. gentryi (subgen.Helmia sect.Monadelpha) andD. andromedusae (subgen.Helmia sect.Centrostemon) from Peru, are described and illustrated.     

Revision of ploidy status of Dioscorea alata L. (Dioscoreaceae) by cytogenetic and microsatellite segregation analysis

Dioscorea alata is a polyploid species with several ploidy levels and its basic chromosome number has been considered by most authors to be x = 10. Standard chromosome counting and flow cytometry analysis were used to determine the chromosome number of 110 D. alata accessions of the CIRAD germplasm collection. The results revealed that 76% of accessions have 2n = 40 chromosomes, 7% have 2n = 60 chromosomes and 17% have 2n = 80 chromosomes. Progenies were produced from 2n = 40 types of D. alata and the segregation patterns of six microsatellite markers in four different progenies were analysed. The Bayesian method was used to test for diploid versus tetraploid (allo- and autotetraploid) modes of inheritance. The results provided the genetic evidence to establish the diploidy of plants with 2n = 40 chromosomes and to support the hypothesis that plants with 2n = 40, 60 and 80 chromosomes are diploids, triploids and tetraploids, respectively, and that the basic chromosome number of D. alata is x = 20. The findings obtained in the present study are significant for effective breeding programs, genetic diversity analysis and elucidation of the phylogeny and the species origin of D. alata.     

Studies on the developmental anatomy of Dioscorea sansibarensis Pax (Dioscoreaceae)

Certain developmental and anatomical studies on D. sansibarensis Pax are briefly reported. (1) Stem diameter, internodal length, vessel diameter, distribution and number of vascular bundles in internodes (from base to apex) are determined. There is no correlation between vessel size and stem diameter. Cut stems left in water develop tyloses. (2) Roots are all polyarch. The development and structure of the vessels are discussed. Xylem and phloem differentiation in the root tip is determined. A central meristem at the root apex gives rise to the different tissues in the root. Raphide development in root cortex is traced. (3) Both alternate and opposite leaf arrangements occur, sometimes on the same plant, and there is a correlation between stem thickness and leaf arrangement. During leaf development the lamina length exceeds the width, but the two dimensions reach their maximum size at the same time. The acuminate leaf-tip matures before the rest of the leaf. Stomatal development is traced.     

Pollen morphology of Dioscorea (Dioscoreaceae) and its relation to systematics

This paper adds new data on the pollen and orbicule morphology of 61 Dioscorea L. (Dioscoreaceae) species to the survey of Schols et al . (2001). The results indicate that pollen characters may be significant in infrageneric systematics in Dioscorea . Pollen and orbicule characters are described based on observations with light microscopy, and scanning and transmission electron microscopy, and are critically evaluated and discussed in the context of existing hypotheses of systematic relationships within the genus. Pollen is mostly disulculate (sometimes monosulcate) with a perforate, microreticulate, striate, gemmate, rugulate, or cerebroid perforate sexine. The basal section Stenophora is one of the few sections with monosulcate pollen. Brachyandra , Cardiocapsa , and Seriflorae , three Malagasy sections, are characterized by striate pollen. Pollen morphology strongly supports section Enantiophyllum as a monophyletic group. The correlation between pollen size and tuber type, as suggested previously by P. Su (1987), is confirmed by our data. As found in our earlier survey, orbicules in Dioscorea are mostly spherical with a smooth or spinulose surface.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 143 , 375–390.     

Polyploidy and sterility in relation to sex in Dioscorea alata L. (Dioscoreaceae)

Chromosome numbers and fertility studies of 73 male and 30 female flowering germplasm accessions of Dioscorea alata L. were carried out. All males were tetraploids showing the same chromosome number (n=20 or 2n=40) and were pollen fertile (10.9–96.2%), most of them being highly fertile. Among the female the majority were higher ploids (hexa-and octoploids; 2n=60 and 80) and sexually completely sterile. There were only two tetraploid female accessions which were sexually fertile. Pollination studies revealed that seed sterility in D. alata was due to female sterility associated with the occurrence of higher levels of ploidy. The female sex-limited occurrence of higher polyploidy and sterility observed in D. alata is a curious situation among dioecious higher plants.     

Improved production of transgenic Dioscorea zingiberensis (Dioscoreaceae) by Agrobacterium tumefaciens-mediated transformation

The establishment of high-efficiency Agrobacterium-mediated transformation techniques could improve the production of Dioscorea zingiberensis, a medicinal species with a high diosgenin content. We co-cultivated embryogenic calli induced from mature seeds with A. tumefaciens strain EHA105. A binary vector, pCAMBIA1381, which contains the gfp and hpt genes under the control of the ubiquitin promoter and the CaMV 35S promoter, respectively, was used for transformation. Pre-culture, basic medium, acetosyringone, and bacterial density were evaluated to establish the most efficient protocol. The optimal conditions consisted of MS medium without CaCl(2) for pre- and co-cultivation, three days for pre-culture, addition of 200 μM AS, and an OD(600) of 0.5. The transgenic plants grown under selection were confirmed by PCR analysis and Southern blot analysis. This protocol produced transgenic D. zingiberensis plants in seven months, with a transformation efficiency of 6%.     

Isolation and characterization of nuclear microsatellite loci in Lupinus group Microcarpi (Leguminosae)

Microsatellite loci are described for Lupinus group Microcarpi (Leguminosae), an assemblage of annual plants primarily inhabiting western North America. In all, 188 unique microsatellite loci were isolated, and seven loci (averaging 9.6 alleles each) have been developed as polymorphic markers for estimating population structure and outcrossing rates in Lupinus microcarpus var. densiflorus and var. horizontalis. Departures from Hardy–Weinberg expected frequencies of heterozygotes are consistent with structured populations and autogamous or mixed mating systems. Several loci were amplified successfully in other species of Lupinus, demonstrating the potential utility of these markers beyond group Microcarpi.     

New microsatellite loci for pomegranate, Punica granatum (Lythraceae)

? Premise of the study: A new set of pomegranate microsatellites was selected and characterized to assess the level of genetic diversity among cultivars and wild genotypes. ? Methods and Results: Nine Simple Sequence Repeat (SSR) markers were obtained using the Microsatellite-AFLP technique and were successfully amplified in 34 genotypes belonging to Italian, Spanish, and Turkish germplasm collections. The number of alleles per locus ranged from 1 to 5, and the total number of alleles was 22. ? Conclusions: Because only a few codominant markers are available for this species, the newly identified SSRs will facilitate genetic diversity studies, fingerprinting, and mapping. In addition, the 9 loci successfully amplified in P. granatum var. nana. No cross transferability was observed for Cuphea micropetala and Lagerstroemia indica (Lythraceae).     

Sixteen microsatellite loci for the Bufo boreas group

The four species that comprise the Bufo boreas group of toads are critically imperiled in all or portions of their geographical ranges. We present data from 16 microsatellite loci isolated from B. boreas that cross‐amplify in these four species. These markers have proven useful in the analyses of population structure and conservation genetics, and provide a powerful tool for future researchers who seek to understand the conservation genetics of these rare toad species. Polymorphism was assessed for 339 individuals from seven populations representing the four species. All loci were polymorphic (X? = 8, range of four to 19 alleles). Three loci were not in Hardy–Weinberg equilibrium (HWE) in one population, and one of these loci was out of HWE in a second population (P < 0.003 after Bonferroni correction for multiple tests). However, there were no systematic deviations from HWE across all study populations. Small populations in fragmented habitat appear to explain the existing HWE deviations.     

Expansion of trait of gibberellin-induced dormancy in the Tertiary relict species of the genus Dioscorea (Dioscoreaceae)

Effects of the application of gibberellin A₃ (GA₃) on sprouting of rhizomes and germination of seeds were tested in five species of the genus Dioscorea which are considered to be Tertiary relicts growing disjunctively in the Appalachians, the Balkans, and the Caucasus. Applied GA₃ at higher than 1 micro;M markedly inhibited sprouting of rhizomes of all species. However, inhibition of seed germination of these species by GA₃ application was not observed. It is hypothesized that before disjunction in the Tertiary period GA-induced dormancy probably prevailed in rhizomes of all members of the genus Dioscorea.     

Characterization of seven (CTT)(n) microsatellite loci in the Pyrenean endemic Borderea pyrenaica (Dioscoreaceae): remarks on ploidy level and hybrid origin assessed through allozymes and microsatellite analyses

We present the identification and characterization of microsatellite loci in the Pyrenean endemic Borderea pyrenaica Miégeville (Dioscoreaceae). Seven microsatellite loci were isolated from a (CTT)(n)-enriched partial genomic library. Electropherograms patterns suggest that B. pyrenaica is a tetraploid species, as is its congener B. chouardii. One microsatellite locus was monomorphic, whereas the remaining ones presented from 2 to 10 alleles when analyzed in a sample of 60 individuals. Microsatellites have revealed higher levels of genetic variability than those in previous studies based on allozymes. Levels of genetic diversity are discussed in terms of tetrasomic (autotetraploidy) or duplicated disomic (allotetraploidy) modes of allele segregation. According to the first hypothesis, mean levels of genetic variability (H(min)-H(max)) range between 0.36 and 0.41, whereas, according to the second hypothesis, the 7 primer pairs amplified 11 chromosomal loci, and mean levels of observed and expected heterozygosities were 0.217 and 0.229, respectively, and did not differ significantly from HW expectations. These results suggest a hybrid allopolyploid origin for the Borderea taxa.