Isolation of polymorphic microsatellite loci in the genus Clusia (Clusiaceae)

Microsatellite flanking region sequences may provide phylogenetically useful information. We isolated 13 polymorphic microsatellite loci from two species, Clusia minor (five loci) and Clusia nemorosa (eight loci), to aid in the determination of phylogenetic relationships within the genus Clusia. Eleven loci amplified across all 17 Clusia species tested, while two loci amplified in 10 out of 17 species. The extensive cross‐species amplification suggests that these loci may be useful for an examination of phylogenetic relationships in this genus.     

Conservation of microsatellite loci within the genus Vitis

Eleven microsatellites isolated from grapevine (Vitis vinifera) were used to study the degree of conservation of these sequences across different Vitis species. Nine microsatellites were newly isolated, the remaining two (VVS2 and VVS5) came from the literature. A preliminary assay on the conservation of priming sites was carried out on 14 non-V. vinifera species, including relevant taxa for breeding. Parthenocissus quinquefolia was added as representative of a related genus. Cross-species amplification was obtained in 94% of the 176 genotype×locus tested combinations. Three microsatellite loci were then cloned and sequenced in ten species. The microsatellite repeat was found present in all cases. The repeat region was often longer in V. vinifera than in the other species. Furthermore the non-source species showed interruptions in the repeat. In spite of these constraints, which could reduce the polymorphism of microsatellites in non-source species, the results demonstrate the possibility of extending the use of microsatellite markers to wild germplasm and inter-specific hybrids. Point mutations have been found in microsatellite flanking regions and these variations have been used to investigate the genetic relationship among taxa. The Neighbor-joining tree that was obtained on the basis of ten nucleotide variations, showed that there is not a clear cut difference between American, Asian and European species and that the actual taxonomy which reflects the geographical distribution of species must most likely be revised. Moreover, in general, nucleotide variations which occur in microsatellite flanking regions provide new molecular tools for investigating the evolution of species. Received: 24 October 1999 / Accepted: 11 November 1999     

Identification and characterization of nuclear microsatellite loci in Abies alba Mill.

Eleven polymorphic nuclear microsatellite markers for Abies alba Mill. were developed from an enriched genomic library. An average of 5.2 alleles per locus and a mean expected heterozygosity of 0.532 were found in a sample of 24 Abies alba individuals from different populations within Europe. These loci can be used in studies of genetic diversity for parentage analysis and for estimation of gene flow in silver fir populations. Moreover, successful amplifications were obtained for eight other Mediterranean Abies species, suggesting that these loci may be useful for similar applications in other fir species.     

Characterization of highly polymorphic nuclear microsatellite loci in Juniperus communis L.

We developed five nuclear microsatellite markers in Juniperus communis L. using an enriched library method. Samples from 28 juniper individuals were collected in Spain, Germany and Slovakia and were analysed at the five loci. A high level of allelic diversity with values ranging from nine to 23 alleles was found. These highly polymorphic markers will be used in ongoing population genetic studies to evaluate the genetic resources and to contribute to the maintenance of genetic diversity of juniper in Middle Europe.     

PCR multiplexing of nuclear microsatellite loci inQuercus species

Simple sequence repeats have become the tool of choice in a wide range of studies of forest trees. Despite frequent use of multicolor fluorescent labeling DNA fragment analyzers, there are few procedures that reduce time and cost of the analyses by PCR multiplexing. Here we present an example of 2 multiplexes of 7 unlinked nuclear microsatellite loci to increase the efficiency of genotyping of large samples of oaks, which is extremely useful in population genetic studies.     

New microsatellite loci for the socially diverse paper wasp genus Ropalidia

The Polistine wasps include both independent‐founding species, with small, single‐queen colonies founded by one or a few potential queens, and swarm‐founding species, which have larger societies, many queens and initiate colonies as a swarm of queens and workers. Swarm‐founding evolved from independent‐founding and Ropalidia is the only genus with both types, making it an excellent model system for understanding this dramatic shift in colony organization. We have isolated 18 polymorphic microsatellite loci from three species of Ropalidia, including two independent‐founding and one swarm‐founding species. These loci will allow us to reconstruct colony social and genetic structures in this important genus.     

Thirty polymorphic nuclear microsatellite loci from black walnut

Black walnut (Juglans nigra L) is a large tree, native to the eastern United States, that is prized for its high-quality timber and edible nut. Thirty (GA/CT)n nuclear microsatellite markers were identified from black walnut for use in population genetic studies, genome mapping, DNA genotyping of important clones, studies of gene flow, and tree breeding. The markers were polymorphic based on a diversity panel of 10 black walnut individuals from eight Midwestern U.S. states.     

Characterization of nuclear microsatellite loci in South American Araucariaceae species

The Araucariaceae family has only two species in South America: Araucaria angustifolia and Araucaria araucana. Both species are mainly used for timber and have been overexploited in the past. Currently, they are found as fragmented populations and are classified under the International Union for Conservation of Nature and Natural Resources (IUCN) guidelines as vulnerable species. Population fragmentation may seriously affect the genetic diversity of these two species of Araucariaceae and can consequently lead to decreased survival. To better understand the genetic structure of these South American Araucaria species, eight nuclear microsatellites are reported: six new microsatellites loci developed based on a membrane enrichment procedure and two microsatellites loci transferred from the related species, Araucaria cunninghamii.     

Cross-amplification and characterization of microsatellite loci for the Neotropical orchid genus Epidendrum

In this study we tested the cross-amplification of 33 microsatellite loci previously developed for two closely related Neotropical orchid genera (Epidendrum and Laelia). A set of ten loci were polymorphic across five examined species (20 individuals each) with 2 to 15 alleles per locus. The mean expected and observed heterozygosity (average across species) ranged from 0.34 to 0.82 and from 0.27 to 0.85, respectively. In addition we tested all loci in 35 species representative of the genus Epidendrum. Of these, 26 loci showed successful amplification. Cross-application of these loci represent a potential source of co-dominant markers for evolutionary, ecological and conservation studies in this important orchid genus.     

Species‐diagnostic microsatellite loci for the fig wasp genus Pegoscapus

To obtain tools for the estimation of inbreeding and assignment of offspring to matrilines, we developed 13 microsatellite loci from the fig wasps that pollinate Ficus obtusifolia. Based on morphological studies, it was thought that a single species (Pegoscapus hoffmeyeri) pollinated this fig. However, our data revealed the presence of two coexisting cryptic species. Several diagnostic microsatellite markers may be used to distinguish these two cryptic species. The new microsatellites can be used across a wide range of fig‐pollinating wasp species for both evolutionary and population genetic studies.     

Isolation and characterization of nuclear microsatellite loci in Pinus pinaster Ait

We developed 20 polymorphic nuclear microsatellites from Pinus pinaster using genomic libraries enriched with repeated motifs. A total of 25 individuals from five different populations were used to estimate genetic diversity parameters.     

Analysis of microsatellite loci of the chloroplast genome in the genus Capsicum (Pepper)

Six plastome microsatellites were examined in 43 accessions of the genus Capsicum. In total, 33 allelic variants were detected. A specific haplotype of chloroplast DNA was identified for each Capsicum species. Species-specific allelic variants were found for most wild Capsicum species. The highest intraspecific variation was observed for the C. baccatum plastome. Low cpDNA polymorphism was characteristic of C. annuum: the cpSSRs were either monomorphic or dimorphic. The vast majority of C. annuum accessions each had alleles of one type. Another allele type was rare and occurred only in wild accessions. The results testified again to genetic conservation of C. annuum and especially its cultivated forms. The phylogenetic relationships established for the Capsicum species on the basis of plastome analysis were similar to those inferred from the morphological traits, isozyme patterns, and molecular analysis of the nuclear genome.     

Characterization of highly polymorphic nuclear microsatellite loci from the ascidian Ciona intestinalis

Eight nuclear polymorphic microsatellite markers were characterized from the ascidian Ciona intestinalis whole genome sequence. The behaviour of these loci was investigated against two geographically distinct populations: one from Plymouth, UK the other from the Fusaro Lagoon, Italy, both belonging to the type A Ciona cryptic species. The markers exhibited six to 29 alleles and average observed heterozygosity ranging from 0.06 to 0.73. These new microsatellite loci demonstrated to be valuable tools for both population genetic analysis at different scales and genetic identification of mutant phenotypes frequently encountered in Mediterranean populations of C. intestinalis.     

Isolation and characterization of polymorphic nuclear microsatellite loci in Pinus cembra L

We developed eight polymorphic nuclear microsatellite markers for the Swiss stone pine (Pinus cembra L.), of which seven may be amplified in a multiplex polymerase chain reaction. Allelic polymorphism across all loci and 40 individuals representing two populations in the Swiss Alps was high (mean = 7.6 alleles). No significant linkage disequlibrium was displayed between pairs of loci. Significant deviation from Hardy–Weinberg equilibrium was revealed at three loci in one population. Cross–amplification was achieved in two related species within the genus (P. sibirica and P. pumila). Thus, the markers may be useful for population genetic studies in these three pine species. They will be applied in ongoing projects on genetic diversity and patterns of gene flow in P. cembra.     

Isolation and characterization of nuclear microsatellite loci in the anadromous marine fish Morone saxatilis

Molecular genetic studies of population structure and gene flow are commonly employed in fish stock assessment and breeding population delineation. Genetically structured breeding populations may have different effective population sizes, distinct reproductive rates and variable susceptibility to harvesting or breeding habitat degradation. Nine microsatellite loci were isolated for Morone saxatilis (Moronidae), an anadromous fish inhabiting the mid‐Atlantic. Microsatellite loci were isolated with a subtractive hybridization method and will be used to estimate population structure. The loci averaged 8.5 alleles each. Seven loci in the Choptank population and two loci in the Potomac population deviated from Hardy–Weinberg expected frequencies of heterozygotes.     

Characterization of microsatellite loci in red wood ants Formica (s. str.) spp. and the related genus Polyergus

Ants are interesting subjects for studies of evolution of altruism. We developed 13 microsatellite loci in a red wood ant Formica (s. str.) yessensis from random amplified polymorphic DNA fragments to study genetic structure within populations and colonies. Five loci bore two to five alleles in both F. (s. str.) yessensis and F. (s. str.) truncorum and two were also polymorphic in a related species, Polyergus samurai. Results suggest that the loci will be useful in evolutionary studies on Formica and Polyergus species.     

Cross-species amplification of microsatellite loci within the dioecious, polyploid genus Actinidia (Actinidiaceae)

Microsatellite marker transfer across species in the dioecious genus Actinidia (kiwifruit) could offer an efficient and time-effective technique for use during trait transfer for vine and fruit improvement in breeding programmes. We evaluated the cross-species amplification of 20 EST-derived microsatellite markers that were fully informative in an Actinidia chinensis mapping family. We tested all 20 markers on 120 genotypes belonging to 21 species, 5 with varieties and/or chromosome races. These 26 taxa included 16 diploids, 7 tetraploids, 2 hexaploids and 1 octaploid, and represented all four taxonomic sections in the genus. All 20 markers showed some level of cross-species amplification. The most successful marker amplified in all genotypes from all species from all sections of the genus, the least successful amplified fragments only in A. chinensis and A. deliciosa. One species, A. glaucophylla, failed to amplify with all but 2 markers. PIC (Polymorphism information content) values were high, with 14 of 17 markers recording values of 0.90 and above. Sequence data demonstrated the presence of the microsatellite in all the amplified products. Sequence homology was less 5′ of the microsatellite and increased toward the start codon of the translated region of the EST from which the marker was derived. The data confirm that EST-derived microsatellite markers from Actinidia species show cross-species amplification with high levels of polymorphism which could make them useful markers in breeding programmes.