Development and characterization of 20 new polymorphic microsatellite markers from Mangifera indica (Anacardiaceae)

? Premise of the study: Twenty microsatellite loci for mango (Mangifera indica), an important commercial fruit tree in East Asia, were developed to evaluate the genetic diversity and identification of cultivars. ? Methods and Results: The 20 new microsatellite markers were isolated from mango using a magnetic bead enrichment method, and polymorphisms were identified in 22 mango cultivars. The number of alleles ranged from one to nine, with expected heterozygosity ranging from 0 to 0.826. The polymorphism information content ranged from 0 to 0.756 with a mean of 0.525. ? Conclusions: These new microsatellite loci should be useful and convenient for further studies of the genetic diversity and identification of cultivars in mango.     

Isolation and characterization of 15 microsatellite loci from mango (Mangifera indica L.) and cross‐species amplification in closely related taxa

We report here on the development and characterization of 15 microsatellite loci isolated from Mangifera indica L. These markers were evaluated using 59 Florida cultivars and four related species from the USDA germplasm collection for mango. Two loci were monomorphic and 13 polymorphic, with two to seven alleles per locus. Four loci departed significantly from Hardy–Weinberg equilibrium and have significant heterozygote deficiency. Nine loci exhibited significant linkage disequilibrium. Cross‐species amplification was successful in four related species. These loci are being used to investigate patterns of genetic variation within M. indica and between closely related species.     

Development of microsatellite markers for mango (Mangifera indica L.)

Microsatellite markers for mango (Mangifera indica L.) were developed using a genomic library enriched for (GA)_n and (GT)_n dinucleotide repeats. A subset of 41 positive clones was sequenced and primers were designed. Twenty‐eight primer pairs produced polymorphic amplification products for a diversity sample including 15 mango cultivars and two accessions from the related species Mangifera laurina and Mangifera applanata. Nineteen simple sequence repeat (SSR) loci with clear scorable patterns were chosen to study diversity in the mango germplasm bank of Guadalupe (FWI). The number of alleles ranged from three to 13 with observed levels of heterozygosity ranging from 0.059 to 0.857.     

Isolation and characterization of novel microsatellite markers for Avena sativa (Poaceae) (oat)

? Premise of the study: A new set of microsatellite primers was developed for Avena sativa and characterized to assess the level of genetic diversity among cultivars and wild genotypes. ? Methods and Results: Using an enrichment genomic library, 14 simple sequence repeat markers were identified. The loci of these markers were characterized and found to be polymorphic in size among 48 genotypes of oat from diverse geographical locations. The number of alleles per locus ranged from two to eight, while the observed heterozygosity ranged from 0.031 to 0.75. ? Conclusions: These newly identified microsatellite markers will facilitate genetic diversity studies, fingerprinting, and genetic mapping of oat. Moreover, these new primers for A. sativa will aid future studies of polyploidy and hybridization in other species in this genus.     

Isolation of microsatellite markers in the tiger mosquito Aedes albopictus (Skuse)

Among the numerous molecular markers available in population genetics, microsatellites are one of the most powerful tools developed in recent years. This paper describes the isolation of six polymorphic microsatellite loci in the tiger mosquito Aedes albopictus using an enriched genomic library technique. Such loci should be an efficient tool in population genetic studies for this mosquito species.     

Isolation and characterization of six novel microsatellite markers for mulberry (Morus indica)

Genetic characterization of germplasm resources is necessary for their effective management and efficient utilization, especially for species like mulberry in which the available germplasm exhibits rich phenotypic diversity with almost no information about its genetic base. Here we present the first report on the isolation of six novel microsatellite markers of mulberry, developed from an enriched genomic library of Morus indica. These markers revealed a high degree of polymorphism (14–26 alleles per locus; polymorphic information content, 0.85–0.90) and a broad cross‐species affinity when tested on a set of 43 elite genotypes including 13 related Morus species. The data thus demonstrate their utility as potentially efficient genetic markers for germplasm characterization, crop improvement and molecular systematics of mulberry.     

Isolation and characterization of microsatellite markers in two basidiomycete species: Pleurotus eryngii and P. ferulae

Six polymorphic microsatellite loci were developed for the closed basidiomycete species Pleurotus eryngii and P. ferulae using an enriched genomic library. The number of alleles per locus varied between two and eight and the expected heterozygosity ranged from 0.30 to 0.72 in P. eryngii and from 0.20 to 0.73 in P. ferulae. This enrichment technique is useful in those species where microsatellite loci are rare in the genome.     

Isolation and characterization of microsatellite markers in Bonelli's eagle (Hieraaetus fasciatus)

To obtain polymorphic molecular markers for population genetics and conservation studies in Bonelli's eagle populations, we screened a partial genomic library enriched for microsatellites with di‐ and tri‐nucleotide motifs [(GT) (CT) (AAC) and (GCC)]. A total of 15 polymorphic markers were obtained. The number of alleles ranged from two to eight. These markers will be very useful for paternity tests and population structure studies as well as for evaluating the outcome of conservation programs.     

Isolation of 10 microsatellite markers in Crepidula coquimbensis (Gastropod,Calyptraeideae) for parentage analyses

Marine invertebrates show a high diversity of embryonic developmental modes and reproductive behaviour. However, little is known about the factors that are promoting the evolution of different strategies. The gastropod Crepidula coquimbensis has a direct embryonic development with encapsulated embryos showing a cannibalistic behaviour. Preliminary observations suggest that victim embryos are not randomly chosen. With intracapsular multipaternity, the intensity of cannibalism might vary according to the relatedness of the siblings. Paternity analyses based on highly polymorphic markers can provide valuable insights into this cannibalism behaviour. Ten polymorphic microsatellite loci were thus developed: the level of polymorphism is large enough to perform paternity analysis and investigate relatedness in this species.     

Isolation of 11 microsatellite markers in Crepidula convexa (Gastropoda, Calyptraeideae) for parentage analyses

Crepidula convexa, a calyptreid gastropod with direct embryonic development, changes sex from male to female in the course of its lifetime (protandry). Under sex-allocation theory, male reproductive success should be independent from age and size (a proxy used for age). However, this may be counterbalanced by female cryptic choice or gregarious behaviour. Eleven polymorphic microsatellite loci were thus developed to examine paternity of embryos and larvae. This set of loci appears suitable to carry out paternity analyses due to the high exclusion probability of unrelated males given the maternal genotype.     

Isolation and characterization of microsatellite markers in Perilla frutescens Brit

We isolated and characterized 13 polymorphic microsatellite primers from Perilla frutescens Brit. var. frutescens by using a modified method that involves one‐way PCR amplification with single primer prior to enrichment with an ‘oligo hook’. The efficiency of this procedure for isolating unique microsatellite sequences was approximately 77%. The number of alleles per microsatellite locus ranged from three to 10 with an average of 6.5 alleles per locus while fragment size varied from 156 to 298 bp. The observed and expected heterozygosities ranged from 0.52 to 0.86 and 0.52 to 0.89, respectively. These newly isolated microsatellite markers are expected to provide valuable resources for different genetic studies currently underway in our Perilla genome research program.     

Characterization of polymorphic microsatellite DNA markers in Hakea oldfieldii Benth. (Proteaceae)

A genomic library was constructed and 13 polymorphic microsatellite markers were developed for Hakea oldfieldii, a woody shrub endemic to southwest Western Australia. Polymorphism was investigated for these markers in 28 individuals from a single population located in restricted habitat at the base of the Whicher Range south of Busselton. Expected heterozygosity ranged from 0.279 to 0.770 and averaged 0.633. Observed heterozygosity ranged from 0.321 to 0.786 and averaged 0.598. The number of alleles per locus ranged from 2.0 to 6.0 and averaged 4.5. These markers will be used to assay genetic diversity and pollen dispersal in this species.     

Characterization of microsatellite markers in the mosquito Ochlerotatus caspius (Diptera: Culicidae)

Seven polymorphic microsatellite loci were developed for the mosquito species Ochlerotatus caspius, using an enriched genomic library. The number of alleles per locus varied between two and 11; the expected heterozygosity (H_E) ranged from 0.18 to 0.77. These microsatellite primers should prove useful for population genetic studies of this mosquito species.     

Development of microsatellite markers in polyploid persimmon (Diospyros kaki Lf) from an enriched genomic library

The oriental persimmon (Diospyros kaki Lf) is believed to have originated in China with subsequent introduction into Japan and Korea in ancient times. The species was then brought to Europe, Brazil and the USA from Japan in the 19th century. Recent studies highlighted the poor state of identification of cultivars in these countries due to incorrect labelling and presence of synonyms among local varieties. Thus, molecular marker characterization of germplasm resources is of great value for genetic resource preservation and plant breeding of persimmon. Therefore, to identify accessions for further plant breeding and germplasm management, 37 microsatellite loci were developed from a CT/AG‐enriched persimmon genomic library.     

Development of microsatellite markers in the Japanese pear (Pyrus pyrifolia Nakai)

Thirteen polymorphic microsatellite loci were developed in the Japanese pear (Pyrus pyrifolia Nakai) by using an enriched genomic library. The obtained microsatellite loci showed a high degree of polymorphism in the Japanese pear with 3–6 alleles per locus. The average values of observed and expected heterozygosities among these 13 loci were 0.69 and 0.71, respectively. Ten microsatellites could successfully amplify loci in the European pear (Pyrus communis L.), which were highly polymorphic as well.     

Characterization of nine novel microsatellite markers from Brandt's vole (Lasiopodomys brandtii)

Highly polymorphic microsatellite markers are the most powerful tools to infer kinship relations. In this study, a library enriched for (AC)(n) (AG)(n) and (AGAT)(n) was constructed for screening microsatellite markers in Brandt's vole (Lasiopodomys brandtii), and nine novel polymorphic microsatellite markers were developed and characterized. The number of alleles ranged from 4 to 11 per locus and the mean polymorphism information content was 0.7535. The observed and the expected heterozygosity values averaged 0.760 (0.554-0.908) and 0.7914 (0.718-0.845), respectively. Average nonexclusion probability for one candidate parent varied from 0.485 to 0.716. These nine novel markers are highly polymorphic and powerful enough for our future kinship analysis.     

Isolation and characterization of microsatellite markers from the budgerigar,Melopsittacus undulatus

The budgerigar, Melopsittacus undulatus, belongs to a group of birds that are among the most common captive‐held avian species that are also most threatened. Budgies are also a valuable resource as model species for many important vertebrate characteristics including auditory and ageing. Several microsatellite‐enriched genomic libraries were developed using standard protocols. Twelve of 20 primer pairs developed from microsatellite‐containing sequences were codominant and polymorphic when tested on 21 birds from three different states. The number of alleles detected ranged from two to four, with the expected and observed heterozygosities ranging from 0.09 to 0.61 and from 0.00 to 0.85, respectively. The genomic libraries and the primers described here will be useful in characterizing budgies.     

Preliminary linkage map of the turkey (Meleagris gallopavo) based on microsatellite markers

The turkey is an agriculturally important species for which, until now, there is no published genetic linkage map based on microsatellite markers--still the markers most used in the chicken and other farm animals. In order to increase the number of markers on a turkey genetic linkage map we decided to map new microsatellite sequences obtained from a GT-enriched turkey genomic library. In different chicken populations more than 35-55% of microsatellites are polymorphic. In the turkey populations tested here, 43% of all turkey primers tested were found to be polymorphic, in both commercial and wild type turkeys. Twenty linkage groups (including the Z chromosome) containing 74 markers have been established, along with 37 other unassigned markers. This map will lay the foundations for further genetic mapping and the identification of genes and quantitative trait loci in this economically important species. Genome comparisons, based on genetic maps, with related species such as the chicken would then also be possible. All primer information, polymerase chain reaction (PCR) conditions, allele sizes and genetic linkage maps can be viewed at http://roslin.thearkdb.org/. The DNA is also available on request through the Roslin Institute.