Polymorphic Dinucleotide Microsatellite Research Articles

PSX-30 Late-Breaking Abstract: Genotypes database of pedigree cattle population in Kazakhstan and the introduction of a calculating mechanism for livestock parentage verification

Abstract A significant increase in pedigree cattle in Kazakhstan has led to scientific work on the conservation of breed genotypes in Kazakhstan and the development of imported breeds. Aim of work: creation a national database and collection of genotyped DNA samples of cattle in Kazakhstan and the introduction of an automated statistical and mathematical calculation of the local pedigree cattle parentage verification. The methodology of work has been the selection of biological samples and registration of data on the animal. DNA and STR locus testing have been performed at the University of Queensland’s Animal Genetics Laboratory (Australia). DNA genotyping was carried out using 21 polymorphic dinucleotide microsatellite loci. The database of DNA genotypes of breeding animals has been written and introduced into the database of breeding livestock in Kazakhstan (www.plem.kz). The method of determining kinship and confirming the origin of the animal was implemented by calculating the probability of random coincidence of genetic markers, taking into account their distribution in the population according to the “product rule.” The frequency distribution of homozygous alleles for 12 microsatellite loci (table 1) recommended by ICAR and 21 DNA loci (table 2) in the context of breeds was calculated, which indicates an increase in reliability when calculating the origin of livestock, with an increase in the number of microsatellite loci studied to 21 markers. The lower probability threshold is established, with 95% statistical significance confirms the relationship between the compared animals (table 3). A database of genotyped DNA samples of beef and dairy cattle breeds in Kazakhstan has been created and is currently used in pedigree beef cattle breeding. Access to the DNA database is provided to Kazakhstan genetic laboratories, and an automated calculation of the mathematical probability of parentage verification for a breeding animal and the generation of a genetic certificate was introduced on the website www.plem.kz

Development and Characterization of 15 Polymorphic Dinucleotide Microsatellite Markers for Tule Elk Using HiSeq3000.

The tule elk (Cervus elaphus nannodes) experienced a severe bottleneck in the 1800s, resulting in low genetic diversity. There is a need for high-resolution genetic assays that can be used to differentiate individual elk, including close relatives, with high confidence. An efficient assay requires multiple markers both polymorphic and that can be amplified in concert with other markers in multiplex reactions. To develop such markers, we employed 150-bp paired-end whole genome shotgun sequencing on an Illumina HiSeq3000 platform to discover dinucleotide microsatellite markers. After preliminary screening of these markers, we selected and screened 15 candidate loci and 5 existing tetra nucleotide markers in 56 tule elk. We combined these markers in 2 multiplex reactions and report primer concentrations and PCR conditions enabling their efficient amplification.

ISOLATION AND CHARACTERIZATION OF 15 MICROSATELLITE LOCI FOR Ophicephalus argus Cantor.

The isolation and development of 15 polymorphic dinucleotide microsatellite loci were described for Ophicephalus argus from the Huaihe River. All loci were polymorphic in the 30 individuals tested. The number of alleles per variable locus ranged from nine to seventeen, with a mean of 12.00. These novel microsatellite loci showed high level of polymorphism. Observed and expected heterozygosities ranged from 0.793 to 0.929 and from 0.841 to 0.952, respectively. Two loci were found deviated from HWE in the sampled population after Bonferroni correction. These microsatellite loci will be useful for revealing population structure, genetic diversity, and phylogeography of Ophicephalus argus.

Head injury, alpha‐synuclein Rep1, and Parkinson's disease

To test the hypothesis that variability in SNCA Rep1, a polymorphic dinucleotide microsatellite in the promoter region of the gene encoding α-synuclein, modifies the association between head injury and Parkinson's disease (PD) risk. Participants in the Farming and Movement Evaluation (FAME) and the Study of Environmental Association and Risk of Parkinsonism using Case-Control Historical Interviews (SEARCH), 2 independent case-control studies, were genotyped for Rep1 and interviewed regarding head injuries with loss of consciousness or concussion prior to Parkinson's disease (PD) diagnosis. Logistic regression modeling adjusted for potential confounding variables and tested interaction between Rep1 genotype and head injury. Consistent with prior reports, relative to medium-length Rep1, short Rep1 genotype was associated with reduced PD risk (pooled odds ratio [OR], 0.7; 95% confidence interval [CI], 0.5-0.9), and long Rep1 with increased risk (pooled OR, 1.4; 95% CI, 0.95-2.2). Overall, head injury was not significantly associated with PD (pooled OR, 1.3; 95% CI, 0.9-1.8). However, head injury was strongly associated with PD in those with long Rep1 (FAME OR, 5.4; 95% CI, 1.5-19; SEARCH OR, 2.3; 95% CI, 0.6-9.2; pooled OR, 3.5; 95% CI 1.4-9.2, p-interaction = 0.02). Individuals with both head injury and long Rep1 were diagnosed 4.9 years earlier than those with neither risk factor (p = 0.03). While head injury alone was not associated with PD risk, our data suggest head injury may initiate and/or accelerate neurodegeneration when levels of synuclein are high, as in those with Rep1 expansion. Given the high population frequency of head injury, independent verification of these results is essential.

Isolation and characterization of polymorphic microsatellite markers in the yellow catfish, Pelteobagrus fulvidraco

The yellow catfish (Pelteobagrus fulvidraco) is one of the important economic freshwater fish in China and widely distributed along Yangtze River. In this survey, we have developed eighteen polymorphic dinucleotide microsatellite markers. They were characterized using 36 samples collected from Chishui and Leshan fragment of Yangtze River. Number of alleles per locus ranged from 2 to 11. Mean observed heterozygosity over all eighteen loci was 0.559 (Chishui population) and 0.848 (Leshan population), respectively. Mean expected heterozygosity was 0.562 (Chishui population) and 0.613 (Leshan population), respectively. These microsatellite markers will be useful in studies of population genetic structure, genome mapping and marker-assisted selection of yellow catfish.

Isolation and characterization of 17 microsatellite loci for the Chinese longsnout catfish (Leiocassis longirostris)

We describe the isolation and development of 17 polymorphic dinucleotide microsatellite loci for the Chinese longsnout catfish (Leiocassis longirostris). All loci were polymorphic in the 30 individuals tested. The number of alleles per variable locus ranged from 6 to 18, with a mean of 11.71. Observed and expected heterozygosities ranged from 0.467 to 1.000 and from 0.540 to 0.929, respectively. Eight loci were found to have deviated from HWE in the sampled population after Bonferroni correction. Linkage disequilibrium tests revealed significant linkage between two loci (LLW5 and LL27). These microsatellite loci will be useful for revealing population structure, genetic diversity, and phylogeography of the Chinese longsnout catfish.

Characterization of microsatellite loci in the subsocial spider Stegodyphus lineatus (Araneae: Eresidae)

Stegodyphus lineatus spiders live in groups consisting of closely related individuals. There appears to be no discrimination against related individuals as mates but females mate multiply, despite the fact that matings are shown to carry a cost. We have developed eight polymorphic dinucleotide microsatellite markers that allow us to assess levels of heterozygosity and relatedness among individuals of this species. These molecular markers are likely to prove highly effective tools for estimating levels of inbreeding and thus allow us to test hypotheses about the relationships between social structure, mating strategies and inbreeding avoidance.

Polymorphic microsatellite loci for the common marmoset (Callithrix jacchus) designed using a cost‐ and time‐efficient method

We describe a cost- and time-efficient method for designing new microsatellite markers in any species with substantial genomic DNA sequence data available. Using this technique, we report 14 new polymorphic dinucleotide microsatellite loci isolated from the common marmoset. The relative yield of new polymorphisms was higher with less labor than described in previous marmoset studies. Of 20 loci initially evaluated, 14 were polymorphic and amplified reliably (70% success rate). The number of alleles ranged from 3 to 9 with heterozygosity varying from 0.48 to 0.83.

A set of polymorphic dinucleotide and tetranucleotide microsatellite markers for the Indo-Pacific humpback dolphin (Sousa chinensis) and cross-amplification in other cetacean species

Eleven dinucleotide and five tetranucleotide polymorphic microsatellite loci were presented for the Indo-Pacific humpback dolphin Sousa chinensis, a species categorized as ‘data deficiency’ in the IUCN Red List. These markers were developed to allow future population studies, such as characterization of population structure and genetic diversity that are important for the species’ conservation. The number of alleles ranged from 2 to 9 with observed heterozygosity ranging from 0.167 to 0.917, and expected heterozygosity ranging from 0.159 to 0.913. Test of these loci in five additional cetacean species found that 10–13 loci have successful cross-amplifications.

PERMANENT GENETIC RESOURCES: Isolation and characterization of polymorphic trinucleotide and dinucleotide microsatellite loci for the ice goby, Leucopsarison petersii

Eleven microsatellite markers were isolated and characterized for the ice goby, Leucopsarion petersii, from genomic libraries enriched for (ATG)(9) and (CA)(16) . Twenty individuals from a single population were used to screen polymorphism in these loci. The number of alleles per locus and observed heterozygosity ranged from four to 22 and from 0.35 to 1.00, respectively. All loci did not significantly deviated from Hardy-Weinberg equilibrium, and there was no evidence of linkage disequilibrium between all loci-pairs. These loci showed Mendelian inheritance in a full-sib family. The high level of polymorphism of these loci will be useful for studies of population genetics.

PERMANENT GENETIC RESOURCES: Ten novel polymorphic dinucleotide microsatellite loci cloned from the Antarctic fur seal Arctocephalus gazella

Ten new dinucleotide microsatellite loci were isolated from the Antarctic fur seal Arctocephalus gazella. These markers should prove useful for studying the reproductive ecology of Antarctic fur seals and other related pinniped species.

Twelve new polymorphic microsatellite markers from the loggerhead sea turtle (Caretta caretta) and cross-species amplification on other marine turtle species

We describe 12 new polymorphic dinucleotide microsatellite loci and multiplex Polymerase Chain Reaction conditions from the loggerhead sea turtle Caretta caretta. Levels of polymorphism were assessed in 50 individuals from the nesting population of the Cape Verde Islands. Number of alleles ranged from 3 to 13 (average of 7.33) and the values of observed heterozygosities from 0.32 to 0.80 (average of 0.61). Cross-species amplification on three other marine turtles, Chelonia mydas, Eretmochelys imbricata and Dermochelys coriacea, revealed polymorphism and variability at eight, eleven and three loci, respectively.

Isolation and characterization of nine polymorphic microsatellite loci from the desert locust, Schistocerca gregaria

AbstractBecause of the scarcity of polymorphic genetic markers, only a few genetic studies on the population structure of the desert locust, Schistocerca gregaria, have been carried out. We isolated and characterized nine polymorphic dinucleotide microsatellite loci. These markers were evaluated using individuals from Niger and Senegal. Seven of these microsatellite markers are also applicable to the nongregarious subspecies Schistocerca gregaria flaviventris. Cross‐species applicability was limited to one of the loci in the sister species S. americana and in the locust Locusta migratoria.

Isolation and characterization of microsatellite markers in pangolins (Mammalia, Pholidota, Manis spp.)

AbstractThirty‐four polymorphic dinucleotide microsatellite loci were developed in the Malayan pangolin Manis javanica. Of the 34 markers, 32 and 18 were also amplified, respectively, in the Chinese pangolin (Manis pentadactyla) and the African tree pangolin (Manis tricuspis). Analysis of 24 Malayan, 12 Chinese and 2 African tree pangolins showed high levels of variability (heterozygosity ranging from 0.321 to 0.708). These are the first available microsatellite markers in Pholidota and will be an invaluable tool for evolutionary and conservation genetic studies in pangolins.

Microsatellite loci in the Propertius duskywing,Erynnis propertius(Lepidoptera: Hesperiidae), and related species

AbstractFifteen polymorphic dinucleotide microsatellite loci were characterized forErynnis propertiususing an enrichment protocol. The number of alleles varied from nine to 28 for a sample of 24 individuals. Observed heterozygosities ranged from 0.25 to 0.96. Homozygote excess was detected for 10 loci. Twelve markers successfully amplified in relatedErynnisspecies and eight loci were polymorphic in at least one other species.

Cross-amplification of 10 new isolated polymorphic microsatellite loci for red mullet (Mullus barbatus) in striped red mullet (Mullus surmuletus)

Ten polymorphic dinucleotide microsatellite loci were isolated and characterized for the red mullet (Mullus barbatus). Allele variability was tested on both the red mullet and its congener the striped red mullet (Mullus surmuletus). Characterization of 30 individuals of both species from the western Mediterranean showed moderate to high allelic diversity ranging from two to 26 alleles per locus (mean 10.9). Three loci showed departures from Hardy–Weinberg proportions. No evidence of significant association between genotypes at pairs of loci was observed. These polymorphic loci could be suitable for population genetic assessments of both species.

Polymorphic microsatellite primers developed from DNA of the endangered Mekong giant catfish, Pangasianodon gigas (Chevey) and cross‐species amplification in three species of Pangasius

AbstractThe critically endangered Pangasianodon gigas is endemic to the Mekong River. Despite its importance, little is known about its genetic diversity and conservation efforts are hampered. Ten polymorphic dinucleotide microsatellite primer pairs were developed from DNA of P. gigas. The analysis of 20 individuals from hatchery stocks using these primers resulted in two to six alleles/locus; HO = 0.05–0.95; HE = 0.05–0.81. All but one locus (Pg‐3) conformed to Hardy–Weinberg expectation. Eight, six and seven primer pairs were amplified with the DNA from Pangasianodon hypophthalmus, Pangasius larnaudii and Pangasius sanitwongsei, respectively. These markers will be useful for genetic monitoring of wild and hatchery stocks of these pangasiids.

Microsatellites from the burbot (Lota lota), a freshwater gadoid fish (Teleostei)

AbstractWe developed 21 polymorphic dinucleotide microsatellite loci, (CA)n and (CT)n, for the Holarctic freshwater fish, Lota lota, using an enriched genomic library protocol. The species has an interesting life history because winter‐spawning adults migrate over long distances to form spawning aggregations, a behaviour which should maintain genetic homogeneity across large spatial scales. Availability of the reported microsatellites will facilitate the investigation of population genetic structure with regard to postglacial colonization history and conservation strategies. The primers were screened on 30 individuals from a natural population (Lake Constance, southern Germany), revealing three to 24 alleles per locus with expected heterozygosities ranging from 0.48 to 0.93.

Polymorphic CA/GT and GA/CT microsatellite loci forOstrinia nubilalis(Lepidoptera: Crambidae)

AbstractTen polymorphic dinucleotide (CA/GT and GA/CT) microsatellite loci suitable for population genetic screening were characterized from enriched partialOstrinia nubilalisgenomic libraries. Sequence from 126 enriched small insert genomic library clones identified 25 CA/GT and 58 GA/CT loci that were unique. Perfect repeats tended to be short (n = 10–12). Ten microsatellites, PCR amplified from a Crawfordsville Iowa population showed a mean of 10 alleles per locus (range six to 20), and six of 10 loci showed heterozygote deficiency. Amplification of eight loci was observed in the sister speciesO.furnicalis.

Isolation and characterization of highly polymorphic microsatellite markers in Hypochaeris radicata (Asteraceae)

AbstractWe developed five highly polymorphic dinucleotide microsatellite loci for the grassland species Hypochaeris radicata (Asteraceae). Polymorphism of these markers was examined in six populations in the Netherlands. All loci were polymorphic in all populations. The number of alleles per locus varied between 18 and 43. Expected heterozygosity was between 0.86 and 0.91. Cross‐species amplification was tested in six Hypochaeris species and was successful for three different loci in four species. These microsatellites are a useful tool in population genetic, dispersal and metapopulation studies or in testing levels of inbreeding.