Microsatellite loci for paternity assessment in the black swan (Cygnus atratus: Aves)

Seven polymorphic tetranucleotide microsatellite loci were isolated and characterized from the black swan Cygnus atratus Latham, using nonradioactive PCR‐based techniques to screen an enriched genomic library. These loci were highly variable (mean H _E  = 0.75, mean number of alleles = 9.1), and showed no evidence of linkage disequilibrium or null alleles. This locus set is suitable for detecting extra‐pair copulations (combined exclusion probability = 0.999) and for other population genetic applications.     

Novel microsatellite loci for the study of the black‐capped vireo (Vireo atricapillus)

We identified 14 novel polymorphic microsatellite loci in the black‐capped vireo (Vireo atricapillus). We also attempted to amplify and genotype these loci in other Vireo species, including the white‐eyed vireo (Vireo griseus), red‐eyed vireo (Vireo olivaceus), and blue‐headed vireo (Vireo solitarius). In 33 genotyped black‐capped vireos from two locations, total alleles ranged from six to 20, with observed heterozygosity ranging from 0.58 to 0.91 and expected heterozygosity from 0.65 to 0.93. Two loci had detectable levels of null alleles. Many of the loci were able to be amplified in the related Vireo species.     

Microsatellite loci in the house fly,Musca domestica L. (Diptera: Muscidae)

Genomic libraries from house flies enriched for (CA)₁₅ and (CAG)₁₀ repeats were constructed by using biotinylated probes. Twenty‐five loci were isolated and evaluated for polymorphisms in wild flies representing two geographically diverse populations. Fourteen of 19 dinucleotide loci, and one of six trinucleotide loci were polymorphic. One hundred and twenty‐seven alleles were detected, 39 of which were private. Average number of alleles per polymorphic locus was 8.4 ± 2.5 and average heterozygosity was 72 ± 4%. F_ST by the private allele method was 0.73. Three of 15 loci showed significant heterozygote deficiencies, attributed to null alleles. Five of 15 loci were amplified in the face fly, Musca autumnalis.     

Isolation and characterization of microsatellite markers in musk duck (Biziura lobata: Aves), and their application to other waterfowl species

We isolated and characterized 11 novel microsatellite loci to study paternity in the Australian musk duck (Biziura lobata), using nonradioactive PCR‐based techniques to screen GA and GAAA repeats enriched genomic DNA libraries. Nine of 11 loci showed no evidence of null alleles and were variable (mean H_E = 0.825, mean number of alleles = 9). This set of nine loci is suitable for paternity assignment (exclusion probability for nine unlinked loci = 0.9999). We also demonstrated that many of these loci cross‐amplify in various other waterfowl species.     

Isolation and characterization of microsatellite markers in the southern emu‐wren (Stipiturus malachurus: Aves)

We isolated and characterized eight novel microsatellite loci in the southern emu‐wren (Stipiturus malachurus). We used nonradioactive polymerase chain reaction (PCR)‐based techniques to screen an enriched genomic DNA library. Based on genotypes from a single population, six loci showed no evidence of null alleles and were polymorphic (allele range = 2–9, mean heterozygosity = 0.57), and one locus was sex‐linked (N_A = 4). These loci were variable and had different allele size ranges in three other populations of southern emu‐wrens, and are therefore useful for determining levels of genetic diversity within and between populations of the species.     

Development of EST‐derived microsatellite markers for Arabidopsis lyrata subspecies petraea (L.)

A set of expressed sequence tag–simple sequence repeat (EST‐SSR) loci has been developed for Arabidopsis lyrata ssp. petraea. From 768 root cDNA clones, 126 microsatellites, including di‐, tri‐, tetra‐ and pentanucleotide repeat motifs were identified and primers were designed to 24 EST‐SSRs. Eleven loci were subsequently screened on 150 individuals sampled from five natural populations, which revealed three to nine alleles per locus (mean 5.36) and expected heterozygosity (H_E) estimates ranging from 0.046 to 0.698. Significant deviations from random mating were observed at 10 EST‐SSR loci, likely due to inbreeding (global F_IS = 0.151) and population structure (global F_ST = 0.246).     

Polymorphic microsatellite loci in the Coffee Berry Borer,Hypothenemus hampei (Coleoptera,Scolytidae)

Microsatellite loci were isolated from the coffee berry borer, Hypothenemus hampei, which is regarded as the major pest in coffee cultures. Seven polymorphic loci were obtained from an enriched genomic library. A low to moderate genetic diversity was observed per locus, with an observed number of alleles ranging from two to five in the 39 unrelated individuals sampled from Ethiopia. A clear deficit of heterozygotes within the population (mean heterozygosities, H_O = 0.10/H_E = 0.50) and an extreme inbreeding (F_IS, 0.70–1.00) were demonstrated. Cross‐species amplifications showed that some of the markers could be useful in two closely related Hypothenemus species.     

Characterization of 27 novel microsatellite loci in Sinibotia superciliaris (Günther 1892) and cross‐species transferability in Sinibotia reevesae (Chang 1944)

The Chinese golden loach, Sinibotia superciliaris and its congener Sinibotia reevesae, are endemic to China and extraordinary similar in morphological traits. However, few genetic studies have been conducted on them, especially those based on nuclear markers. Here, we developed 27 novel polymorphic microsatellite markers from S. superciliaris and further tested the cross‐species transferability of those loci in S. reevesae. The cross‐species amplification test showed that 19 loci demonstrate polymorphism in S. reevesae. The mean number of alleles (N_A) were both 3 in S. superciliaris and S. reevesae, the mean expected heterozygosities (H_E) were 0.57 and 0.54, the Shannon‐Wiener Diversity Indices (SW) were 0.84 and 0.82, and the evenness (E) were 0.09 and 0.08, respectively. The polymorphic information content (PIC) showed a high level of polymorphism for the two species (mean 0.54 and 0.50, respectively). In addition, the cross‐species transferability (100%) of those markers was clearly high, which confirmed that the microsatellite markers developed here could be used effectively for other related species.     

Isolation of microsatellites from two species of scleractinian coral

Microsatellites were isolated from two broadcast‐spawning species of scleractinian coral (Platygyra daedalea and Goniastrea favulus) from Australia's Great Barrier Reef. We found 27 microsatellites across both species, although only five loci were polymorphic in each species. Microsatellite loci displayed a wide range of diversity levels with four to 11 alleles per locus (H_O = 0.26–0.91) in P. daedalea and two to seven alleles per locus (H_O = 0.16–0.96) in G. favulus. Most loci showed departures from Hardy–Weinberg equilibrium which may reflect nonrandom mating but may also be related to difficulties associated with coral DNA.     

Isolation and characterization of polymorphic microsatellite markers in Anthyllis vulneraria

To study the reproductive system of the putative autogamous species Anthyllis vulneraria in Belgian calcareous grasslands, eight polymorphic microsatellite loci were developed using the sequence‐tagged microsatellite profiling technique and 35 plants were typed for all loci. No linkage disequilibrium was found between any pair of loci. Over the loci, the number of alleles ranged from two to five. Observed and expected heterozygosities ranged from 0.053 to 0.688 and from 0.053 to 0.631, respectively. The estimated F_IS values (F_IS = 0.031 and 0.122) are not in accordance with a predominantly selfing reproductive system.     

Isolation and characterization of polymorphic microsatellite loci for the study of paternity and population structure in the little penguin Eudyptula minor

We isolated and characterized eight novel microsatellite loci in the little penguin Eudyptula minor, using nonradioactive polymerase chain reaction‐based techniques to screen GA and GAAA repeats from enriched genomic DNA libraries. All eight loci were polymorphic and seven were variable in our main study population (mean H_E = 0.613, mean N_A = 7.14). Cross‐amplification using a microsatellite primer developed in Spheniscus demersus (African penguin) yielded one additional polymorphic locus. This locus combined with six of the little penguin loci is suitable for paternity assignment in little penguins (exclusion probability for seven unlinked loci = 0.993).     

Polymorphic microsatellite loci for paternity analysis in the Madagascar paradise flycatcher (Terpsiphone mutata: Aves)

Nine polymorphic microsatellite loci from the Madagascar paradise flycatcher Terpsiphone mutata were isolated using nonradioactive polymerase chain reaction (PCR)‐based techniques to screen an enriched genomic library. Seven polymorphic loci showed no evidence of null alleles and exhibited high levels of variation in 18 unrelated individuals (mean diversity = 0.80, mean number of alleles = 13.6). These loci are therefore suitable for analysis of population structure and paternity (exclusion probability for six unlinked loci = 0.9998).     

Differential introgression from a sister species explains high FST outlier loci within a mussel species

Scanning genomes for loci with high levels of population differentiation has become a standard of population genetics. F_ST outlier loci are most often interpreted as signatures of local selection, but outliers might arise for many other reasons too often left unexplored. Here, we tried to identify further the history and genetic basis underlying strong differentiation at F_ST outlier loci in a marine mussel. A genome scan of genetic differentiation has been conducted between Atlantic and Mediterranean populations of Mytilus galloprovincialis. The differentiation was low overall (F_ST = 0.03), but seven loci (2%) were strong F_ST outliers. We then analysed DNA sequence polymorphism at two outlier loci. The genetic structure proved to be the consequence of differential introgression of alleles from the sister‐hybridizing species Mytilus edulis. Surprisingly, the Mediterranean population was the most introgressed at these two loci, although the contact zone between the two species is nowadays localized along the Atlantic coasts of France and the British Isles. A historical contact between M. edulis and Mediterranean M. galloprovincialis should have happened during glacial periods. It proved difficult to disentangle two hypotheses: (i) introgression was adaptive, implying edulis alleles have been favoured in Mediterranean populations, or (ii) the genetic architecture of the barrier to edulis gene flow is different between the two M. galloprovincialis backgrounds. Five of the seven outliers between M. galloprovincialis populations were also outliers between M. edulis and Atlantic M. galloprovincialis, which would support the latter hypothesis. Differential introgression across semi‐permeable barriers to gene flow is a neglected scenario to interpret outlying loci that may prove more widespread than anticipated.     

Isolation and characterization of microsatellite loci from mother‐of‐millions,Bryophyllum delagoense (Crassulaceae), and its hybrid with Bryophyllum daigremontianum, ‘Houghton's hybrid’

Nine microsatellite loci were developed, and are transferable, across the Madagascan succulents Bryophyllum daigremontianum, Bryophyllum delagoense (mother‐of‐millions) and their horticultural hybrid (Houghton's), from enriched libraries of the later two species. For B. delagoense, a tetraploid, three to 13 alleles per locus were found for native Madagascan (H_O = 0.4–1.0), and one to nine in invasive Australian (H_O = 0.0–1.0) samples. In addition for 91 Australian samples, only five multilocus genotypes were found (95% of individuals were of two genotypes), suggesting extensive clonality in its introduced range. These loci will be used to examine genetic diversity, hybrid origin and mating system in natural and introduced populations.     

Polymorphic microsatellite loci for the trematode Diplostomum pseudospathaceum

We present primers for five polymorphic microsatellite loci in the eye fluke Diplostomum pseudospathaceum (Trematoda), a widely distributed parasite with a complex life cycle used as a model for parasitology and fish immunology. The loci were identified using a GA/CT‐enriched genomic library by subtractive hybridization with magnetic particles. All five loci were highly polymorphic, displaying 17 to 61 alleles and heterozygosities ranging from 0.53 to 0.92. We isolated populations of parasites within the first (snail) and second (fish) intermediate host and found small but significant genetic differentiation (F_ST = 0.012) between the two life stages of the parasite.     

Isolation and characterization of microsatellites in yellow perch (Perca flavescens)

A total of 45 microsatellite loci from yellow perch, Perca flavescens, were isolated and characterized. Among the 45 microsatellite loci, 32 had more than two alleles. A wild population of P. flavescens (n = 48) was used to examine the allele range of the microsatellite loci. Mendelian inheritance of alleles was confirmed by examining the amplified products in pair‐mated families. The number of alleles for the 32 polymorphic loci varied from two to 16, and observed heterozygosity ranged between 0.024 (YP79) and 0.979 (YP60). Cross‐species polymorphic amplification in four other Percidae species was successful for 22 loci.     

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.     

Genetic drift outweighs natural selection at toll‐like receptor (TLR) immunity loci in a re‐introduced population of a threatened species

During population establishment, genetic drift can be the key driver of changes in genetic diversity, particularly while the population is small. However, natural selection can also play a role in shaping diversity at functionally important loci. We used a well‐studied, re‐introduced population of the threatened Stewart Island robin (N = 722 pedigreed individuals) to determine whether selection shaped genetic diversity at innate immunity toll‐like receptor (TLR) genes, over a 9‐year period of population growth following establishment with 12 genetic founders. We found no evidence for selection operating with respect to TLR diversity on first‐year overwinter survival for the majority of loci, genotypes and alleles studied. However, survival of individuals with TLR4_BE genotype was significantly improved: these birds were less than half as likely to die prior to maturity compared with all other TLR4 genotypes. Furthermore, the population frequency of this genotype, at a two‐fold excess over Hardy–Weinberg expectation, was increased by nonrandom mating. Near‐complete sampling and full pedigree and reproductive data enabled us to eliminate other potential causes of these patterns including inbreeding, year effects, density dependence, selection on animals at earlier life history stages or genome‐level association of the TLR4_E allele with ‘good genes’. However, comparison of observed levels of gene diversity to predictions under simulated genetic drift revealed results consistent with neutral expectations for all loci, including TLR4. Although selection favoured TLR4_BE heterozygotes in this population, these effects were insufficient to outweigh genetic drift. This is the first empirical study to show that genetic drift can overwhelm natural selection in a wild population immediately following establishment.     

Isolation and characterization of microsatellite markers for paternity assessment in the golden whistler (Pachycephala pectoralis: Aves)

We isolated and characterized six novel microsatellite markers for paternity analysis in the golden whistler Pachycephala pectoralis, by screening an enriched genomic library using nonradioactive PCR techniques. The six loci exhibited little or no evidence of null alleles and showed high levels of polymorphism (mean H_E = 0.85, mean number of alleles = 15.2), making them suitable for paternity assessment in this species (exclusion probability of six unlinked loci = 0.9997).     

Genetic variation and hybridization in SwedishSchoenus (Cyperaceae)

Schoenus ferrugineus andS. nigricans have restricted distributions in Sweden and are almost exclusively confined to calcareous fen habitats. AtS. nigricans sites,S. ferrugineus is usually also present, and hybrids are frequently found. In this report, I used allozymes to estimate the amount of gene flow between the two species, and to compare the partitioning of genetic diversity in each of them. Thirteen loci were analysed at eight different enzyme systems. Seven loci were variable between or within the species. The two species had completely different alleles at two of the seven variable loci, whereas there was overlap at five loci. In all, 22 different alleles were found. Six of these alleles were confined toS. nigricans, and five alleles were confined toS. ferrugineus. Nei's genetic identity was 0.55.—InS. ferrugineus, three loci (23%) were polymorphic, and the average number of alleles per polymorphic locus was 2.0 (each polymorphic locus had two alleles). InS. nigricans, three loci (23%) were polymorphic, and the average number of alleles per polymorphic locus was 2.3.—The proportion of genetic diversity due to variation among sites (G _ST) was fairly similar in the two species, mean over loci = 0.12 inS. ferrugineus and 0.15 inS. nigricans. However, the proportion of genetic diversity due to variation among individuals within sites (G _IS) differed markedly between the two species, mean over loci = 0.54 inS. ferrugineus and 0.17 inS. nigricans. Accordingly, there was a much higher individual heterozygosity inS. nigricans than inS. ferrugineus. — Most hybrids were interpreted as F₁ hybrids. However, a small proportion, 0.5–1.6 %, were F_n hybrids or back-crosses.—On the Swedish mainland, all former occurrences ofS. nigricans are extinct, but viable hybrids are still present at a few sites in southernmost Sweden.