Isolation and characterization of microsatellite loci in the Pacific pleasure oyster,Crassostrea corteziensis,and their cross‐species amplification in four other oyster species

Microsatellite loci were isolated in Crassostrea corteziensis using (GT)_n, (CT)_n and (CTGT)_n‐enriched genomic libraries. Within each of 45 sequenced clones, an average of three microsatellite regions (156 total) were observed. Thirty‐three primers were designed, from which 11 microsatellite loci amplified. Ten of those were polymorphic, with a range of two to 30 alleles. Three loci were not in Hardy–Weinberg equilibrium, and linkage disequilibrium was found for six pairs of loci. These microsatellite loci will be further tested for segregation distortions and null alleles to establish a set for population genetic studies of the species in the Northwest coasts of Mexico, and for optimization of aquaculture development. Seven of the microsatellite loci cross‐amplified in Crassostrea palmula, a sympatric species, and will be useful in further genetic studies.     

Detection of genetically unstable loci in parthenogenic families of lizards of theLacerta genus by DNA fingerprinting

Two parthenogenic families of unisexual species of Caucasian rock lizards of genusLacerta, L. armeniaca andL. unisexualis, were analyzed by DNA fingerprinting. Inheritance of M13 minisatellite and of (GACA) _n , (GATA) _n , and (TCC) _n microsatellite loci in the first generation of the lizards was studied. M13, (GACA) _n , and (TCC) _n loci in the families ofL. armeniaca were strictly inherited, as well as M13 and (GACA) _n loci in the families ofL. unisexualis: each DNA fragment in the fingerprint patterns of progeny could be detected in the maternal pattern. However, when a (TCC)₅₀ microsatellite probe was applied in the study ofL. unisexualis families, specific DNA fragments with altered mobility were revealed in the progeny patterns, and the frequency of such events was rather high. It might be hypothesized that some of the (TCC) _n loci inL. unisexualis genome are highly mutable. Hence, the family analysis allowed us to demonstrate experimentally the presence of genetically unstable loci in genomes of parthenogenic species of vertebrates. The nature and mechanism of the instability of these loci in parthenogenesis remain obscure.     

Identification of microsatellite loci in lake sturgeon,Acipenser fulvescens,and their variability in green sturgeon,A. medirostris

From (CATC)_n, (GATA)_n, (AAAC)_n, and (CA)_n–enriched libraries for the lake sturgeon Acipenser fulvescens, 254 primer pairs were developed. These primer pairs resulted in the identification of 128 microsatellite loci in either A. fulvescens or A. medirostris. Polymorphic loci were identified in both sturgeon species for 48 of the primer pairs and 14 of the primer pairs amplified polymorphic loci only in A. medirostris. Most of the identified loci appear to be tetrasomic (79.1% in A. fulvescens and 64.5% in A. medirostris). These results offer estimates of the degree of diploidization in each of these species.     

Characterization of microsatellite loci for five members of the minnow family Cyprinidae found in the Sacramento–San Joaquin Delta and its tributaries

Two microsatellite‐enriched libraries [(CAGA)_n, (TAGA)_n] were constructed using pooled DNA from three cyprinid species native to the Sacramento–San Joaquin Delta of California: Sacramento splittail (Pogonichthys macrolepidotus); Sacramento pikeminnow (Ptychocheilus grandis); and tui chub (Siphateles bicolor). Primers were designed for 105 loci and tested for levels of polymorphism in five cyprinid species found in the Delta: Sacramento splittail, Sacramento pikeminnow, tui chub, hitch (Lavinia exilicauda), and Sacramento blackfish (Orthodon microlepidotus). Fifty‐one loci were polymorphic for at least one species and 31 loci were polymorphic for multiple species. The number of polymorphic loci per species ranged from 16 to 26.     

Microsatellite loci from Russula brevipes,a common ectomycorrhizal associate of several tree species in North America

Six microsatellite loci were isolated and characterized from genomic libraries enriched for (CA)_n (GA)_n (ATG)_n, and (CAG)_n, microsatellite motifs from Russula brevipes, a common ectomycorrhizal fungus that forms mutualisms with several species of trees in North America. The polymerase chain reaction primers were tested on 27 sporocarps of R. brevipes sampled in Douglas fir (Pseudotsuga menziesii), grey pine (Pinus sabiniana), and Sitka spruce (Picea sitchensis) stands. The number of alleles per locus ranged from two to 14 with expected heterozygosity values from 0.00 to 0.92 within populations. These are the first six microsatellite loci characterized from Russula brevipes that can be used for estimating genotypic diversity and population structure.     

Tightly linked di- and tri-nucleotide microsatellites do not evolve in complete independence: evidence from linked (TA)n and (TAA)n microsatellites of chickpea (Cicer arietinum L.)

In order to understand the dynamics of microsatellite evolution, we have studied allelic variation at a closely linked (TA) _n and (TAA) _n microsatellite loci in 114 land races of chickpea (Cicer arietinum L.), sampled worldwide. These two loci are separated by 27 bp. The two loci showed a very high degree of polymorphism and hence the combined length with the genetic diversity of 0.93, 0.90 and 0.98 for (TAA) _n , (TA) _n and the combined length, respectively. Using the variation data at the linked loci, a standardized index of linkage disequilibrium was also computed (I ^S _A =0.092), which tests the null hypothesis of no linkage and was significant, indicating the presence of linkage disequilibrium. Furthermore, the dynamics of allelic variation showed that there is a threshold combined length, below which both (TAA) _n and (TA) _n loci evolve independently, and above which, if one locus increase in size, the other closely linked locus has a tendency to decrease its size and vice versa, without change in the overall ratio of (TAA) _n and (TA) _n allele sizes at the region. This result indicates that there are processes in the cell, which read the combined size of the two loci both for proportion and length and determine the direction of tightly linked di- and tri-nucleotide repeat evolution.Communicated by R. Hagemann     

Isolation and characterization of polymorphic microsatellite markers in the gray,short‐tailed opossum (Monodelphis domestica)

Twenty‐six polymorphic microsatellite markers were isolated from (AC)_n and (AG)_n microsatellite‐enhanced genomic libraries of the gray, short‐tailed opossum Monodelphis domestica. All 26 loci showed high allelic diversity, with allele numbers ranging from five to 11 in a subset of 35 animals. Normal Mendelian inheritance was confirmed for 24 loci by analysing allelic segregation in 10, two‐generation, families. Non‐amplifying (null) alleles were detected at two loci, which we recommend be used only if pedigree data are available. We conclude that all of these microsatellite markers would be useful for quantitative trait locus mapping and population genetic studies.     

Isolation of microsatellite loci from the common buzzard,Buteo buteo (Aves: Accipitridae)

We isolated 56 common buzzard (Buteo buteo) microsatellite loci from (AC)_n‐ and (AAAG)_n‐enriched genomic libraries. Eleven loci were tested on 90 individuals from Eastern Westphalia in Germany, yielding two to 17 alleles per locus (average 5.7) and expected heterozygosities ranging from 0.11 to 0.93 (average 0.53). These highly variable microsatellite markers provide a powerful means for investigating population genetic diversity in the common buzzard, a little‐understood aspect of this widespread species.     

Characterization of microsatellite loci in the Blue‐and‐gold Macaw,Ara ararauna (Psittaciformes: Aves)

Microsatellite loci were isolated from the Blue‐and‐gold Macaw (Ara ararauna), a Neotropical parrot, from a GT_n and CT_n enriched genomic library. Six loci were characterized varying from one to 11 alleles per locus. Five loci exhibited greater than 50% heterozygosity within the 49 individuals genotyped. Furthermore, the primers also amplified the DNA from two additional genera of Neotropical parrots, indicating the potential utility of these markers for population‐level studies and conservation efforts of Neotropical parrots.     

Microsatellite markers for the Mexican spotted owl (Strix occidentalis lucida)

A genomic cosmid library was used to develop seven highly polymorphic microsatellite markers for the Mexican spotted owl (Strix occidentalis lucida). These are the first reported microsatellite markers derived from this species. The cloned and sequenced repeat motifs include a triplet repeat of (AAT)_n, two tetranucleotide repeats of (GATA)_n, a tetranucleotide repeat of (ATCC)_n, a compound repeat of (GA)_n(GATA)_n and the two pentanucleotide repeats (AGAAT)_n and (ATTTT)_n. The microsatellites described represent six presumably independent loci with the two pentanucleotide repeats having originated from a single cosmid. Primer pairs allow locus‐specific amplification of each marker from Mexican spotted owl genomic DNA.     

Isolation of polymorphic tetranucleotide microsatellite markers for the ornate nursery frog Cophixalus ornatus

We have isolated 18 polymorphic microsatellite loci for Cophixalus ornatus from genomic libraries enriched for (AAAG)_n, (AACC)_n and (AAGG)_n repetitive elements. The number of alleles ranges from five to 22 per locus with the observed heterozygosity ranging from 0.10 to 0.92. These markers will be useful for the analysis of population structure in C. ornatus and testing alternative models of speciation.     

Isolation and characterisation of 11 tetranucleotide microsatellite loci in the common genet (Genetta genetta)

We report the isolation and characterization of 11 polymorphic tetranucleotide microsatellite loci in the common genet (Genetta genetta) from genomic libraries enriched for (AAAG) _n and (AGAT) _n repeat sequences. We chose to develop tetranucleotide repeats because they can be scored less ambiguously. In a sample of 25 individuals, we observed between four and thirteen alleles per locus and their observed and expected heterozygosities ranged from 0.60 to 0.84 and from 0.68 to 0.92, respectively. All genotypic frequencies conformed to Hardy–Weinberg equilibrium expectations and there were no instances of linkage disequilibrium detected between pairs of loci. These loci will be of use in studies of population genetics, historical demography, and molecular ecology of the common genet.     

Ten polymorphic dinucleotide microsatellite markers of the noble scallop Chlamys nobilis

The primers flanking 22 microsatellites isolated from a genomic library enriched for (CA)_n and (GA)_n were designed in the noble scallop Chlamys nobilis. Ten primer pairs provided clear and polymorphic amplification products. Based on characterization with 48 individuals, the number of alleles ranged from three to six. The values of observed heterozygosity and expected heterozygosity varied from 0 to 0.88 and from 0.29 to 0.76, respectively. No significant linkage disequilibrium between pairs of loci was found and six of 10 loci conformed to the Hardy–Weinberg equilibrium. These markers are therefore potentially useful for studies of the population structure of the species.     

Isolation and characterization of Brachymystax lenok microsatellite loci and cross‐species amplification in Hucho spp. and Parahucho perryi

We isolated and characterized eight polymorphic microsatellite markers for Brachymystax lenok (Pallas, 1773) from genomic libraries enriched for (GATA)_n, (GACA)_n and (ATG)_n microsatellites. The number of alleles per locus ranged from two to 17. Heterozygosity ranged from 0.2 to 0.95. In addition, cross‐species amplification was successful for seven loci in Hucho hucho, eight in H. taimen and seven in Parahucho perryi.     

Polymerase chain reaction primers for polymorphic microsatellite loci from the túngara frog Physalaemus pustulosus

We developed eight PCR?primer pairs of polymorphic microsatellite loci for the túngara frog Physalaemus pustulosus. Genomic libraries were enriched for one of four microsatellite repeat sequences (CA_n, GA_n, ATG_n and TAGA_n). Following characterization of microsatellite loci by sequencing, primers were designed and PCR conditions optimized. Microsatellite PCR‐amplification was tested in 37 frogs from 8 populations in Costa Rica and Panama. Primer sequences, PCR conditions, allelelic diversities and observed as well as expected heterozygosities in the screened populations are described.     

Polymorphic microsatellite loci isolated from the marine sponge Scopalina lophyropoda (Demospongiae: Halichondrida)

A total of seven microsatellites out of 88 isolated from a genomic library enriched for (CA)_n and (GA)_n repeats were characterized in the Mediterranean marine sponge Scopalina lophyropoda. The microsatellite motifs were large (34.81 ± 13.9 bp) and imperfect. The seven microsatellite loci were screened in 30 individuals collected from Blanes, northwestern Mediterranean. All of them were polymorphic (allele numbers and observed heterozygosities ranged from 3 to 6 and from 0.16 to 0.76, respectively). No significant linkage disequilibrium between pairs of loci and no departure from Hardy–Weinberg equilibrium were found. These markers are therefore promising for studies of the population structure of the species.     

Isolation and characterization of polymorphic microsatellite loci from Wilsonia backhousei (Convolvulaceae)

Wilsonia backhousei is a clonal saltmarsh plant restricted to the southern latitudes of Australasia and threatened in New South Wales. We have identified eight informative microsatellite loci in the species from (AG)_n‐ and (AC)_n‐enriched libraries. In 48 samples from two populations we detected an average of five alleles per locus (range 2–8, average H_E = 0.45), of which 72% were unique to one population or the other. Six of the eight loci were also amplifiable in Wilsonia rotundifolia under the same reaction conditions. The markers will be excellent tools for use in the management and conservation of both species.     

Microsatellites for tree of heaven (Ailanthus altissima)

Nine (CT)_n microsatellites were developed for tree of heaven, Ailanthus altissima, from invasive populations on the Mediterranean islands. These loci had seven to 12 alleles in 96 trees from five islands. Two loci had significant deficits of heterozygotes within islands while the other loci were in Hardy–Weinberg equilibrium, and four pairs of loci had significant linkage disequilibrium within a single island. These loci were also polymorphic in one to three individuals of the tree of heaven varieties, Ailanthus altissima erythrocarpa, Ailanthus altissima sutchuenensis and Ailanthus altissima tanakai, and the related species Ailanthus giraldii and Ailanthus vilmariniana.     

Instability of (GATA)<Subscript> n</Subscript> microsatellite loci in the parthenogenetic Caucasian rock lizard<Emphasis Type="Italic"> Darevskia unisexualis</Emphasis> (Lacertidae)

Mini- and microsatellites, comprising tandemly repeated short nucleotide sequences, are abundant dispersed repetitive elements that are ubiquitous in eukaryotic genomes. In humans and other bisexual species hypervariable mini- and microsatellite loci provide highly informative systems for monitoring of germline and somatic instability. However, little is known about the mechanisms by which these loci mutate in species that lack effective genetic recombination. Here, multilocus DNA fingerprinting was used to study M13 minisatellite and (GATA) _n microsatellite instability in the parthenogenetic Caucasian rock lizard Darevskia unisexualis (Lacertidae). DNA fingerprinting of 25 parthenogenetic families, from six isolated populations in Armenia (comprising a total of 84 siblings), using the oligonucleotide (GATA)₄ as a hybridization probe, revealed mutant fingerprinting phenotypes in 13 siblings that differed from their mothers in several restriction DNA fragments. In three families (8 siblings), the mutations were present in the germline. Moreover, the mutant fingerprint phenotypes detected in siblings were also present in population DNA samples. No intrafamily variations in DNA fingerprint patterns were observed with the M13 minisatellite probe. Estimates of the mutation rate for (GATA) _n microsatellite loci in D. unisexualis showed that it was as high as that seen in some bisexual species, reaching 15% per sibling or 0.95% per microsatellite band. Furthermore, in one case, a somatic (GATA) _n microsatellite mutation was observed in an adult lizard. These findings directly demonstrate that mutations in (GATA) _n microsatellite loci comprise an important source of genetic variation in parthenogenetic populations of D. unisexualis.Communicated by G. P. Georgiev     

Isolation and characterization of microsatellite loci from Psidium guajava L.

A (GA)_n and (GT)_n microsatellite‐enriched library was constructed and 23 nuclear simple sequence repeat (SSR) loci were characterized in the guava species (Psidium guajava L.). All SSR loci were found to be polymorphic after screening for diversity in different cultivars, and across‐taxa amplification tests showed the potential transferability of most SSR markers in three other Psidium species. First to be published for P. guajava, this new SSR resource will be a powerful tool for genetic studies of guava, including cultivars identification and linkage mapping, as well as potentially for interspecific genetic studies within the genus Psidium.