Set Of Polymorphic Microsatellite Markers Research Articles

The Development of a Fluorescent Microsatellite Marker Assay for the Pitaya Canker Pathogen (Neoscytalidium dimidiatum).

Pitaya canker, caused by Neoscytalidium dimidiatum, is a destructive disease that significantly threatens the safety of the pitaya industry. The authors of previous studies have mainly focused on its biological characteristics and chemical control. However, there are no molecular markers available thus far that can be used for the population genetics study of this pathogen. In the present study, a draft genome of N. dimidiatum with a total length of 41.46 MB was assembled in which 9863 coding genes were predicted and annotated. In particular, the microsatellite sequences in the draft genome were investigated. To improve the successful screening rate of potentially polymorphic microsatellite makers, another five N. dimidiatum isolates were resequenced and assembled. A total of eight pairs of polymorphic microsatellite primers were screened out based on the polymorphic microsatellite loci after investigating the sequencing and resequencing assemblies of the six isolates. A total of thirteen representative isolates sampled from different pitaya plantations were genotyped in order to validate the polymorphism of the resulting eight markers. The results indicated that these markers were able to distinguish the isolates well. Lastly, a neighbor-joining tree of 35 isolates, sampled from different pitaya plantations located in different regions, was constructed according to the genotypes of the eight molecular markers. The developed tree indicated that these molecular markers had sufficient genotyping capabilities for our test panel of isolates. In summary, we developed a set of polymorphic microsatellite markers in the following study that can effectively genotype and distinguish N. dimidiatum isolates and be utilized in the population genetics study of N. dimidiatum.

Genome-Wide Development of Polymorphic Microsatellite Markers and Genetic Diversity Analysis for the Halophyte Suaeda aralocaspica (Amaranthaceae).

Suaeda aralocaspica, which is an annual halophyte, grows in saline deserts in Central Asia with potential use in saline soil reclamation and salt tolerance breeding. Studying its genetic diversity is critical for effective conservation and breeding programs. In this study, we aimed to develop a set of polymorphic microsatellite markers to analyze the genetic diversity of S. aralocaspica. We identified 177,805 SSRs from the S. aralocaspica genome, with an average length of 19.49 bp, which were present at a density of 393.37 SSR/Mb. Trinucleotide repeats dominated (75.74%) different types of motifs, and the main motif was CAA/TTG (44.25%). We successfully developed 38 SSR markers that exhibited substantial polymorphism, displaying an average of 6.18 alleles with accompanying average polymorphism information content (PIC) value of 0.516. The markers were used to evaluate the genetic diversity of 52 individuals collected from three populations of S. aralocaspica in Xinjiang, China. The results showed that the genetic diversity was moderate to high, with a mean expected heterozygosity (He) of 0.614, a mean Shannon's information index (I) of 1.23, and a mean genetic differentiation index (Fst) of 0.263. The SSR markers developed in this study provide a valuable resource for future genetic studies and breeding programs of S. aralocaspica, and even other species in Suaeda.

Assessment of Microsatellite Markers for Parentage Testing in Jersey Crossbred Cattle in Tamil Nadu

A study was aimed to evaluate a set of polymorphic microsatellite markers for their efficiency in testing parentage in Jersey crossbred cattle available in Tamil Nadu. The objective of validation was based on the criteria such as polymorphism information content, various genetic diversity parameters and the ability of markers to exclude the wrong parent. A total of 21 microsatellite markers were included to verify the parentage in 24 Jersey crossbred trios (comprising 24 dams, 24 progenies and 2 sires). The microsatellite loci were amplified using fluorescently labelled primers by multiplex PCR and fragment analysis was done through capillary electrophoresis using automated DNA analyzer. Statistical analyses were done using Pop gene version 1.31, CERVUS 3.0.7 and GENALEX 6.503 software programs. The number of alleles for the markers utilized in the study ranged from 4 (ILSTS11) to 12 (INRA23 and TGLA122) with an overall mean of 8.0952 ± 0.47 alleles per locus. The markers also exhibited high expected heterozygosity (He) and polymorphism information content (PIC) ranging from 0.6362 (ETH3) to 0.8629 (SPS115) with a mean of 0.7681 ± 0.013 and 0.592(ETH3) to 0.83(CSSM66) with a mean of 0.7256±0.014 respectively, signifying them to be highly informative. Low overall probability of identity (PI) of 0.0943 ± 0.008 and high overall probability of exclusion (PE) of 0.9619±0.02 with the increasing locus combinations were observed. The probability of exclusion was cent per cent (PE=1.0000) when a combination of 8 markers were used with one known parent and a combination of 12 markers when excluding a putative pair, suggesting the efficacy and suitability of the markers used in the study for parentage testing on Jersey crossbreds of Tamil Nadu.

Design and development of a multiplex microsatellite panel for the genetic characterisation and diversity assessment of domestic turkey (Meleagris gallopavo gallopavo)

Domestic turkey production generally utilises only a few genetically improved lines, and local breeds are severely endangered as a result. Furthermore, the genetic resources of domestic turkeys have not been properly investigated, which could, ultimately, lead to the extinction of local breeds and negatively affect their corresponding genetic diversity and environmental adaptation. Although, several microsatellite markers have been designed for mapping and quantitative trait locus analysis, there is no standard panel of markers for genetic characterisation or genetic diversity assessment. Accordingly, the present study aimed to develop a set of polymorphic microsatellite markers that could be used for international turkey population studies. Thirty-nine microsatellites were selected based on polymorphism, DNA sequence and chromosome position, as well as on amplification efficiency, success rate and the absence of nonspecific amplification. The markers were screened using 105 DNA samples from local turkey breeds from Mexico, the United States, Italy, Brazil, Egypt and Spain. A total of 401 alleles were identified, with a mean number of alleles per marker of 10.28 ± 4.25. All microsatellites were polymorphic, with at least four alleles and no more than 19 alleles. Furthermore, allelic richness ranged from 3.810 to 17.985, mean heterozygosity ranged from 0.452 ± 0.229 to 0.667 ± 0.265, polymorphic information content values ranged from 0.213 (MNT264) to 0.850 (RHT0024) and the mean Fis value was 0.322. Overall, the panel was highly polymorphic and exhibited moderate Hardy–Weinberg disequilibrium, thereby indicating its value as a tool for biodiversity and population structure studies that could play an important role in promoting the conservation of local turkey breeds. Highlights Important genetic resources reside within indigenous turkey populations. These are linked to historic heritage production values and breeds. It is important to preserve this heritage and genetic diversity, which threatens to be lost as production systems focus on production characteristics. Microsatellite markers, even though, they are now replaced by single nucleotide polymorphism automatic genotyping platforms in many fields of genetics, remain a viable alternative thanks to their cheapness and simplicity of study which makes them particularly useful when the population to be studied lacks information of the prior genetic structure.

Isolation and cross-amplification of the first set of polymorphic microsatellite markers of two high-Andean cushion plants

In the southern Andes mountains (27-39◦S) Azorella madreporica and Laretia acaulis, two Apiaceae cushion plant species commonly known as yaretas, conform a well-established altitudinal vegetation belt along the lower Andean zone. These species have been considered as fundamental components of several ecological dynamics within their communities; however, high mountain ecosystems are increasingly threatened worldwide by natural and anthropogenic pressures and the southern Andes are not the exception. Recognizing that genetic information is crucial for the success of any conservation or restoration initiative inwild populations, we developed and cross-amplified 28 specifically designed microsatellite markers (14 in A. madreporica and 14 in L. acaulis), and also tested the cross amplification of 25 markers from the related species Azorella selago. In a region which is particularly vulnerable to global change trends, this new polymorphic microsatellite loci will be useful in the study of the genetic diversity of these high-mountain cushion plants, which are pivotal in the structuring of their native ecosystems.

Lessons from the macroinvertebrates: species‐genetic diversity correlations highlight important dissimilar relationships

Summary Species and genetic diversity patterns are predicted to co‐vary due to similar mechanistic processes. Previous studies assessing species and genetic diversity correlations (SGDCs) have focused primarily on local diversity patterns or island‐like systems and ignore the underlying dispersal network. Here we assessed local and regional SGDCs using freshwater macroinvertebrates sampled across the Rhine river network, a spatially large and highly connected system, in Switzerland. We utilised a set of polymorphic microsatellite markers to assess the genetic diversity of two amphipod species of the Gammarus fossarum complex, which were compared to species level diversities of Amphipoda, Ephemeroptera, Plecoptera, Trichoptera and family level macroinvertebrate diversity across 217 randomly selected sites. All sites were selected based on a representative and standardised species‐sampling scheme. We analysed within site (α‐SGDC) and between‐site SGDC (β‐SGDC). Against our expectation, we generally found negative or null α‐SGDCs and β‐SGDCs. However, we did find genetic diversity to be spatially structured, whereas species richness was related to local environmental factors. These findings suggest that the genetic and species levels of diversity observed are driven by different mechanisms (e.g., environment versus demography), or operate across different temporal or spatial scales (e.g., colonisation history or dendritic river network structure), and may be attributed to differences in the species' ecology or life history. Overall, conservation measures in riverine systems aiming at only one level of diversity may not necessarily benefit other levels of diversity.

Characterization of the complete mitochondrial genome and a set of polymorphic microsatellite markers through next-generation sequencing for the brown brocket deer Mazama gouazoubira.

The complete mitochondrial genome of the brown brocket deer Mazama gouazoubira and a set of polymorphic microsatellite markers were identified by 454-pyrosequencing. De novo genome assembly recovered 98% of the mitochondrial genome with a mean coverage of 9-fold. The mitogenome consisted of 16,356 base pairs that included 13 protein-coding genes, two ribosomal subunit genes, 22 transfer RNAs and the control region, as found in other deer. The genetic divergence between the mitogenome described here and a previously published report was ∼0.5%, with the control region and ND5 gene showing the highest intraspecific variation. Seven polymorphic loci were characterized using 15 unrelated individuals; there was moderate genetic variation across most loci (mean of 5.6 alleles/locus, mean expected heterozygosity = 0.70), with only one locus deviating significantly from Hardy-Weinberg equilibrium, probably because of null alleles. Marker independence was confirmed with tests for linkage disequilibrium. The genetic variation of the mitogenome and characterization of microsatellite markers will provide useful tools for assessing the phylogeography and population genetic patterns in M. gouazoubira, particularly in the context of habitat fragmentation in South America.

Discovery and characterization of a first set of polymorphic microsatellite markers in Siganus oramin

Nine microsatellite DNA markers were developed and characterized for Siganus oramin by the 5'-anchored polymerase chain reaction technique. A total of 42 alleles were identified in 30 individuals, and the number of alleles per locus ranged from 3 to 7, with an average of 4.7. The observed and expected heterozygosity per locus ranged from 0.5333 to 1.0000 and from 0.5254 to 0.8474, respectively, with an average of 0.7422 and 0.6906, respectively. A significant deviation from the Hardy-Weinberg equilibrium was detected at one microsatellite locus after a Bonferroni's correction (P < 0.0056). No significant linkage disequilibrium was found between any of the pairs of the nine loci. The microsatellite loci developed in this study will improve our understanding of the genetic background of S. oramin.

Isolation and characterization of ten microsatellite loci for wild Citrus japonica (Rutaceae)

The genus of Citrus Linnaeus (Rutaceae) includes several widely cultivated species, such as C. sinensis (L.) Osbeck, C. limon (L.) Burm.f. and C. maxima (Burm.) Merr., which are top ranked among all the fruit crops (Mabberley 1997). Citrus is believed to be native to Southeast Asia (Scora 1975; Gmitter and Hu 1990) and has been cultivated for fruit for over 4000 years (Webber et al. 1967). The long cultivation history, together with a great number of intraspecific and interspecific hybrids in Citrus, makes the genetic background extremely complex. The C. japonica is native to South China, with wild populations distributed in broad-leaved evergreen forests and cultivated forms widely grown in warm parts of China (Zhang et al. 2008). It is a species complex, integrated from the formerly segregated genus Fortunella, consisting of F. hindsii (Champ. ex Benth.) Swingle, F. venosa (Champ. ex Benth.) Huang, F. margarita (Lour.) Swingle, F. japonica (Thunb.) Swingle and F. bawangica Huang (Zhang et al. 2008). Despite this taxonomic revision, a considerable amount of variation in fruit size within some wild populations still exists. Further comprehensive field and molecular studies throughout the complex are needed. Recently, due to human activities and environmental changes, wild populations of C. japonica have been dramatically declining (Huang et al. 2010). Evaluating the genetic structure of these wild populations is urgent for their conservation. Additionally, various cultivated forms of C. japonica have important ornamental and medicinal, as well as food values. However, their origins have never been identified. Microsatellites have been proven to be efficient genetic markers for taxon delimitation, genetic structure analysis and detection of origins of cultivars. In the present study, a set of polymorphicmicrosatellite markers for

Characterisation of the complete mitochondrial genome and 13 microsatellite loci through next-generation sequencing for the New Caledonian spider-ant Leptomyrmex pallens

The complete mitochondrial genome and a set of polymorphic microsatellite markers were identified by 454 pyrosequencing (1/16th of a plate) for the New Caledonian rainforest spider-ant Leptomyrmex pallens. De novo genome assembly recovered the entire mitochondrial genome with mean coverage of 8.9-fold (range 1-27). The mitogenome consists of 15,591 base pairs including 13 protein-coding genes, 2 ribosomal subunit genes, 22 transfer RNAs, and a non-coding AT-rich region. The genome arrangement is typical of insect taxa and very similar to the only other published ant mitogenome from the Solenopsis genus, with the main differences consisting of translocations and inversions of tRNAs. A total of 13 polymorphic loci were also characterized using 41 individuals from a single population in the Aoupinié region, corresponding to workers from 21 nests and 16 foraging workers. We observed moderate genetic variation across most loci (mean number of alleles per locus=4.50; mean expected heterozygosity=0.53) with evidence of only two loci deviating significantly from Hardy-Weinberg equilibrium due to null alleles. Marker independence was confirmed with tests for linkage disequilibrium. Most loci cross amplified for three additional Leptomyrmex species. The annotation of the mitogenome and characterization of microsatellite markers will provide useful tools for assessing the colony structure, population genetic patterns, and dispersal strategy of L. pallens in the context of rainforest fragmentation in New Caledonia. Furthermore, this paper confirms a recent line of evidence that comprehensive mitochondrial data can be obtained relatively easily from small next-generation sequencing analyses. Greater synthesis of next-generation sequencing data will play a significant role in expanding the taxonomic representation of mitochondrial genome sequences.

Discovery and characterization of a first set of polymorphic microsatellite markers in red crab (Charybdis feriatus)

Discovery and characterization of a first set of polymorphic microsatellite markers in red crab (Charybdis feriatus)

Isolation by the 5′ anchored PCR technique and characterization of eighteen microsatellite loci in horseshoe crab (Tachypleus gigas)

The horseshoe crab, a well-known ‘living fossil’, is represented by two species (Tachypleus gigas and Carcinoscorpius rotundicauda) in peninsular Malaysia. One of these, T. gigas inhabits shallow marine waters and migrates to intertidal sandy beaches for breeding during high tides at every full and new moon throughout the year (Hajeb et al. 2009). This species is being used as bait by local fishermen to catch crabs or shrimps by netting or cage fishing methods. Due to its low commercial value in comparison to other marine economic species such as fishes, crabs and shrimps, most of the animals caught in fishing nets or cages are released immediately back to sea by fishermen or left to die in the sun. In certain states of Malaysia, such as Kelantan, Penang and Johore, adult female horseshoe crabs are sold to local seafood restaurants and in fish markets due to consumption of their eggs by the local people. Horseshoe crab populations are rapidly declining around the world due to various reasons such as water pollution, loss of their living and spawning habitats and human exploitation of adult horseshoe crabs (Li et al. 2009; Shin et al. 2009). Assessing the genetic variability of horseshoe crab is important as it has direct benefit for the conservation and management of wild populations of the species. However, there is no information available on the population genetic structure and diversity of T. gigas in Malaysia. To address this research need, we have isolated and developed the first set of polymorphic microsatellite markers for T. gigas by using the 5′ anchored PCR technique. A total of 130 individuals of horseshoe crabs were collected from five locations along the coastal areas of peninsular Malaysia, namely Port Dickson (Negeri Sembilan), Pantai Bersih (Penang), Sungai Muar (Johor), Pantai Balok

Identification of sugarcane microsatellites associated to sugar content in sugarcane and transferability to other cereal genomes

Microsatellites or simple sequence repeats (SSRs) markers are very informative for various applications in genetics and breeding. Information obtained with these markers has contributed to a better understanding of evolution and the complexity of the sugarcane genome. With the objective of identifying a large set of polymorphic microsatellite markers designated as Unigene derived Sugarcane Microsatellite (UGSM) and Sugarcane Enriched Genomic Microsatellite (SEGMS), 351 UGSM and 36 SEGMS were tested to find out informative SSRs marker for sugar content. These markers were screened and validated for their use in genetic diversity, cross transferability and comparative linkage potential in high and low sugar bulk of two segregating progenies and twenty each, cultivated high and low sugar cultivars. 158 (40.83%) of the microsatellite markers (144-UGSM: 14-SEGMS) were found to be highly robust and polymorphic. Cross amplification was estimated among nineteen accessions of six sugarcane cultivars, one inter specific hybrids, five related species, four related genera, and three divergent genera by using 27 UGSM primers. Analysis of 388 alleles, amplified by these markers, indicated the high number of observed allele ranged from 2 to 26, with an average of 14.37 alleles detected per locus. High level of polymorphism detected by these markers among sugarcane species, genera and cultivars was 96.3%, while cross-transferability rate was 98.0% within Saccharum complex and 88.27% to cereals. Wide range of genetic diversity (0.33–0.79 with an average of 0.56) assayed with UGSM markers suggested their importance in various genotypic applications in sugarcane.

Isolation and characterization of 19 highly polymorphic microsatellite markers in the devil stinger, Inimicus japonicus (Synanceiidae)

Inimicus japonicus, the devil stinger, has an extensive distribution along the coast of China, Japan and the Korean Peninsula. Nineteen highly polymorphic microsatellite markers were isolated and characterized in I. japonicus. Twenty-eight individuals from a wild population were tested for polymorphism using this set of polymorphic microsatellite markers. The number of alleles per locus ranged from 4 to 14. The ranges of observed and expected heterozygosity were 0.500-0.892 and 0.521-0.910, respectively. Significant deviations from Hardy-Weinberg equilibrium were detected at two loci. To the best of our knowledge, these were the first microsatellite loci characterized from the Synanceiidae; they can be used for estimating genetic diversity, population structure studies, parentage analysis, genetic linkage map construction, germplasm classification and identification, gene identification, quantitative trait loci mapping, and marker-assisted selection in breeding of I. japonicus and other species of this family.

Isolation and characterization of 12 microsatellite loci for the tropical tree species Shorea maxwelliana and S. laevis (Dipterocarpaceae)

Shorea maxwelliana and S. laevis are characteristic tree species of lowland dipterocarp forests. A set of polymorphic microsatellite markers was isolated for gene flow research, particularly targeting the endangered S. maxwelliana. Micro-satellites were isolated from a microsatellite-enriched library of S. laevis. Primer pairs were designed for 144 loci, 12 of which exhibited high levels of polymorphism in S. maxwelliana, nine of these were also polymorphic for S. laevis. The number of alleles and observed heterozygosity ranged from 3 to 13 and 0.467 to 0.900, respectively. These markers will facilitate the direct estimation of gene flow within and among populations of both species.

Construction of a Genetic Linkage Map and Mapping of a Female-Specific DNA Marker in Half-Smooth Tongue Sole (Cynoglossus semilaevis)

The half-smooth tongue sole (Cynoglossus semilaevis, hereafter, "tongue sole") is a marine flatfish with great commercial importance for fisheries and aquaculture in China. It has also been a promising model for the study of sex determination mechanisms in fish. Here, we report the construction of a genetic linkage map for the tongue sole, based on 137 markers including 103 AFLP markers, 33 microsatellite markers, and one female-specific DNA marker. Twenty-six linkage groups (LGs) were found. The total map length was 934.6 cM (Kosambi), with an average spacing of 8.4 cM, covering 64.4% of the estimated genome size. Furthermore, a female-specific SCAR marker, CseF-382, was mapped on LG5. This study represents the first genetic linkage map in the tongue sole. This map has great potential in the identification of quantitative traits loci and sex-related genes and marker-assisted selection in the tongue sole. Meanwhile, the new set of polymorphic microsatellite markers developed in this study is not only useful for genetic mapping but also of critical importance for studies on genetic diversity and broodstock management in tongue sole.

Isolation and charaterization of polymorphic microsatellite loci from so-iuy mullet (Mugil soiuy Basilewsky 1855)

The first set of polymorphic microsatellite markers were developed from so-iuy mullet (Mugil soiuy Basilewsky 1855). From a (GT)n-enriched genomic library, 53 microsatellites were selected for designing microsatellite primers, of which 36 gave working primer pairs. Ten of these loci were polymorphic in a test population of 24 individuals with alleles ranging from 3 to 9, and observed and expected heterozygosities from 0.2083 to 0.9167 and from 0.2651 to 0.8812, respectively. No significant linkage disequilibrium between pairs of loci was found, but two loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction. These polymorphic microsatellite loci should provide sufficient level of genetic diversity to investigate the fine-scale population structure and evaluate the breeding strategy in Mullet.

Isolation and characterization of eight microsatellite loci from the house finch (Carpodacus mexicanus)

AbstractI isolated the first set of polymorphic microsatellite markers from the house finch, Carpodacus mexicanus, a well‐studied North American bird species, as part of an effort to compare levels of genetic diversity in introduced and native populations. Here, I describe eight independently assorting microsatellite loci screened for polymorphism using 40 house finches. Polymorphism levels ranged from six to 14 alleles (mean = 10.6), making these markers a powerful tool for paternity and population level analyses of this widely distributed North American species.

Genetic linkage analysis supports the presence of two susceptibility loci for alcoholism and heavy drinking on chromosome 1p22.1-11.2 and 1q21.3-24.2

BackgroundIn order to confirm a previous finding of linkage to alcoholism on chromosome 1 we have carried out a genetic linkage study.MethodsDNA from eighteen families, densely affected by alcoholism, was used to genotype a set of polymorphic microsatellite markers at loci approximately 10 centimorgans apart spanning the short arm and part of the long arm of chromosome 1. Linkage analyses were performed using the classical lod score and a model-free method. Three different definitions of affection status were defined, these were 1. Heavy Drinking (HD) where affected subjects drank more than the Royal College of Psychiatrists recommended weekly amount. 2. The Research Diagnostic Criteria for alcoholism (RDCA) 3. Alcohol Dependence Syndrome (ADS) as defined by Edwards and Gross (1976) and now incorporated into ICD10 and DSMIV.ResultsLinkage analyses with the markers D1S1588, D1S2134, D1S1675 covering the cytogenetic region 1p22.1-11.2 all gave positive two point and multipoint lods with a maximum lod of 1.8 at D1S1588 (1p22.1) for the RDCA definition of alcoholism. Another lod of 1.8 was found with D1S1653 in the region 1q21.3-24.2 using the HD affection model.ConclusionThese results both support the presence of linkage in the 1p22.1-11.2 region which was previously implicated by the USA Collaborative Study of the Genetics of Alcoholism (COGA) study and also suggest the presence of another susceptibility locus at 1q21.3-24.2.

Hereditary Catalepsy: Genetic and Molecular Mechanisms of Catalepsy in Mice

The results of experiments on the inheritance and neurobiological mechanism of high predisposition to tonic immobility (catalepsy) in CBA mice are discussed. Genetic analysis has demonstrated a monogenic inheritance of the predisposition to catalepsy. A set of polymorphic microsatellite markers has been used to demonstrate that the predisposition to catalepsy is linked to the distal fragment of mouse chromosome 13, which contains the gene of the 5-HT1A-serotonin receptor. Pharmacological and biochemical evidence for the association between hereditary catalepsy and 5-HT1A-receptor dysfunction are presented. The use of CBA mice for studying the mechanisms of depression and the effects of antidepressants is discussed.