Polymorphic Tetranucleotide Microsatellite Loci Research Articles

Population Genetics of the Copperhead at Its Most Northeastern Distribution

Population genetic data are an important standard with which to gauge demography and gene flow within and among biodiversity units, and are often gathered on species of conservation concern. Yet an exclusive focus on ‘conservation immediacy' can also have negative consequences. For example, it can shift monitoring efforts away from more abundant and/or widely distributed clades and, by so doing, promote a more myopic approach to conservation and management. A current example concerns North American pitvipers (Viperidae: Crotalinae), within which listed species of Crotalus and Sistrurus receive considerable population genetic attention whereas broadly distributed Agkistrodon is largely overlooked. To address this disparity, we used 22 polymorphic tetra-nucleotide microsatellite loci to explore genetic structure, diversity, and relatedness in a Connecticut population of Copperhead (A. contortrix). Three admixed genetic clusters were identified across five winter dens, with overall and sex-specific relatedne...

Development of microsatellite loci for the endangered seagrass Zostera japonica (Zosteraceae).

Premise of the study:New microsatellite markers were developed for the Asian endangered seagrass Zostera japonica (Zosteraceae) to assess genetic diversity and population structure of this species. In China, Z. japonica populations have drastically decreased since the 1970s.Methods and Results:A total of 12 polymorphic tetranucleotide microsatellite loci were isolated and characterized in Z. japonica. The number of alleles per locus ranged from one to 11. The expected and observed heterozygosity ranged from 0 to 0.772 and from 0 to 1.000, respectively.Conclusions:The new microsatellites will be useful in evaluating clonality and population structure of Z. japonica and aiding in conservation and management of the endangered seagrass in Asia.

Isolation and characterization of twelve novel polymorphic tetranucleotide microsatellite loci for Tsinling Tree Toad (Hyla tsinlingensis)

The Tsinling Tree Toad, Hyla tsinlingensis is a tree frog endemic to the Tsinling–Dabie mountains, with a restricted and patchy distribution that is currently shrinking. Twelve novel polymorphic tetranucleotide microsatellite loci markers were isolated and characterized for H. tsinlingensis. The number of alleles per locus is range from 2 to 10. Values of observed heterozygosity (Ho) and expected heterozygosity (He) ranged from 0.000 to 0.579 and from 0.272 to 0.883, respectively. One locus showed deviation from Hardy–Weinberg equilibrium and no linkage disequilibrium was detected. These markers should be a useful tool for further population genetic studies of H. tsinlingensis.

Genome-wide survey and analysis of microsatellites in giant panda (Ailuropoda melanoleuca), with a focus on the applications of a novel microsatellite marker system.

BackgroundThe giant panda (Ailuropoda melanoleuca) is a critically endangered species endemic to China. Microsatellites have been preferred as the most popular molecular markers and proven effective in estimating population size, paternity test, genetic diversity for the critically endangered species. The availability of the giant panda complete genome sequences provided the opportunity to carry out genome-wide scans for all types of microsatellites markers, which now opens the way for the analysis and development of microsatellites in giant panda.ResultsBy screening the whole genome sequence of giant panda in silico mining, we identified microsatellites in the genome of giant panda and analyzed their frequency and distribution in different genomic regions. Based on our search criteria, a repertoire of 855,058 SSRs was detected, with mono-nucleotides being the most abundant. SSRs were found in all genomic regions and were more abundant in non-coding regions than coding regions. A total of 160 primer pairs were designed to screen for polymorphic microsatellites using the selected tetranucleotide microsatellite sequences. The 51 novel polymorphic tetranucleotide microsatellite loci were discovered based on genotyping blood DNA from 22 captive giant pandas in this study. Finally, a total of 15 markers, which showed good polymorphism, stability, and repetition in faecal samples, were used to establish the novel microsatellite marker system for giant panda. Meanwhile, a genotyping database for Chengdu captive giant pandas (n = 57) were set up using this standardized system. What’s more, a universal individual identification method was established and the genetic diversity were analysed in this study as the applications of this marker system.ConclusionThe microsatellite abundance and diversity were characterized in giant panda genomes. A total of 154,677 tetranucleotide microsatellites were identified and 15 of them were discovered as the polymorphic and stable loci. The individual identification method and the genetic diversity analysis method in this study provided adequate material for the future study of giant panda.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1268-z) contains supplementary material, which is available to authorized users.

Isolation and characterization of 16 highly polymorphic tetranucleotide microsatellite DNA markers in Paa spinosa

The giant spiny frog, Paa spinosa is a vulnerable species. By using a modified FIASCO (Fast Isolation by AFLP Sequences Containing repeats) method, we isolated and characterized 16 polymorphic tetranucleotide microsatellite loci for P. spinosa. These loci had high polymorphism when screened in 33 randomly caught individuals. The average number of alleles for 16 loci was 7.13 (range 2–14). Observed and expected heterozygosities were 0.152–0.818 and 0.144–0.918, respectively, and did not deviate from Hardy–Weinberg expectations. There was no evidence for scoring error due to stutter, null alleles, or large allele dropout. No significant association between pairs of loci was found after Bonferroni correction. These polymorphic loci will be effective for evaluating genetic diversity, population structure, gene flow and determining paternity in this frog, and also for conservation management.

Development of twenty-one polymorphic tetranucleotide microsatellite loci for Schizothorax o'connori and their conservation application

Schizothorax o'connori, of the subfamily Schizothoracinae, is an endemic fish in the Yarlung Tsangpo River in Tibet, China. To evaluate the population genetic diversity as a means of devising conservation strategies, a total of 21 polymorphic tetranucleotide microsatellite loci were isolated from S. o'connori using the fast isolation by AFLP of sequences containing repeats (FIASCO) protocol. The characterization of these loci was assessed against a panel of 42 individuals. The number of alleles per locus ranged from 4 to 29 with an average of 17.76. Expected locus-specific heterozygosity and the multilocus Shannon–Wiener diversity index ranged from 0.114 to 0.951 and from 0.280 to 3.131, with an average of 0.851 and 2.338, respectively. The cross-species amplification and applicability of these loci were tested against the two other Schizothorax species from the same river. These microsatellite loci will be valuable for further studies of population genetic diversity and genetic structure, and assessments of the artificial propagation release effect of S. o'connori and other related species.

Isolation and characterization of fifteen polymorphic tetranucleotide microsatellite loci in Schizopygopsis younghusbandi Regan

Schizopygopsis younghusbandi Regan is an important endemic fish in Tibet, China. Its wild population has been rapidly declining for the past few decades due to overfishing, dam construction and invasion of exotic fishes. To conserve natural resources and develop breeding stocks of S. younghusbandi, fifteen polymorphic tetranucleotide microsatellite loci were isolated and characterized in 45 individuals collected from the Yarlung Tsangpo River in Tibet. The number of alleles per locus ranged from 6 to 32 with an average of 17.87. The expected heterozygosity and Shannon–Wiener diversity index ranged from 0.454 to 0.951 and from 0.931 to 3.201, respectively. These microsatellite loci will be useful for future population genetic analysis and resource conservation of S. younghusbandi.

Isolation and characterization of twelve polymorphic tetranucleotide microsatellite loci from Yerana yei

Twelve highly polymorphic microsatellite markers were isolated and characterized for the frog Yerana yei. Loci were isolated from a genomic library from Y. yei enriched for (ATAG)n repetitive elements. We designed primers that reliably amplified twelve polymorphic loci and tested them on 32 individuals from two populations. The number of alleles per locus ranged from 3–20, the observed and expected heterozygosity ranged within 0.500–1.000 and 0.752–0.964 respectively, and the polymorphic information content (PIC) value ranged from 0.683–0.930. These microsatellite markers could be valuable in future research concerning the conservation genetics and molecular ecology of this species.

Research Note Characterization of polymorphic microsatellite markers for the Neotropical leaf-frog Phyllomedusa burmeisteri and cross-species amplification

Twelve polymorphic tetranucleotide microsatellite loci were isolated and characterized for the leaf-frog Phyllomedusa burmeisteri, an endemic species of the Brazilian Atlantic forest. These loci were screened in 25 individuals from two populations of the Minas Gerais State (Carangola and Juiz de Fora). The number of alleles per locus ranged from 3 to 16 (mean = 8). Observed and expected heterozygosities ranged from 0.25 to 0.92 and 0.56 to 0.92, respectively. Evidence for both the presence of null alleles and Hardy- Weinberg equilibrium deviations were found in loci Phybu4, Phybu17, and Phybu21. Genotypic disequilibrium for each pair of loci across populations was not significant. Cross-species amplification was successful for 11 of the 12 developed loci for the sister-species, P. bahiana. These microsatellites will be important for future fine-scale population structure analyses.

Development of 34 new microsatellite markers for the endangered Chinese sucker (Myxocyprinus asiaticus) using 454 sequencing

Chinese sucker (Myxocyprinus asiaticus) is an endangered fish species in China which is only distributed in Yangtze River basin. Here we isolated and characterized 34 polymorphic tri and tetra-nucleotide microsatellite loci in the genome of M. asiaticus using 454 sequencing. Of the 73 loci screened, 47 were amplified successfully and 34 were polymorphic. The genetic diversity in M. asiaticus Wanzhou population was moderate with the number of alleles per locus ranged from 2 to 8 (mean 3.97), and the observed and expected heterozygosities ranged from 0.08 to 1.000 (mean 0.50) and from 0.12 to 0.83 (mean 0.55), respectively. These polymorphic microsatellite markers should be useful for further studies of population structure, parentage analysis, and conservation management for this species.

Characterisation of tetra-nucleotide microsatellite loci in the ostrich (Struthio camelus)

1. Eleven polymorphic tetra-nucleotide microsatellite loci were identified in the ostrich (Struthio camelus) using a selective enrichment protocol. 2. The average number of alleles observed was 6·6 with an average heterozygosity of 0·4. 3. The population was found to be in Hardy-Weinberg equilibrium and two of the loci had a greater than 95% probability of having null alleles. 4. These microsatellite loci will add to the existing pool of markers available for the ostrich and help to facilitate analysis of population structure and pedigree determination.

Isolation and characterization of eleven polymorphic tetranucleotide microsatellite loci for Quasipaa boulengeri (Anura: Dicroglossidae)

We isolated and characterized eleven polymorphic tetranucleotide microsatellite loci in the spiny-bellied frog (Quasipaa boulengeri). The loci were screened in 62 individuals from two natural populations. All loci were variable, with the number of alleles ranging from 8 to 26. The average observed and expected heterozygosity per locus ranged from 0.500 to 0.947 and from 0.735 to 0.932, respectively. Two loci were found to be significant departure from Hardy–Weinberg equilibrium and no significant linkage disequilibrium was detected after sequential Bonferroni corrections. These polymorphic loci will be useful for conservation and population genetics studies of this threated species.

Microsatellite markers for the Iberian endemic Bosca’s newt, Lissotriton boscai (Caudata, Salamandridae)

The new generation Roche/454 sequencing of DNA fragments enriched for microsatellite loci was used to isolated microsatellite markers for the Iberian endemic Bosca’s newt, Lissotriton boscai. Two multiplex PCR sets were optimized in order to genotype ten polymorphic tetranucleotide microsatellite loci. The level of genetic diversity of these loci was assessed in 42 individuals from three central-south populations of Portugal (Nazare, Alcobaca and Evora). The number of alleles per locus ranged from 11 to 20 (mean; Na = 16). Observed and expected heterozygosities ranged from 0.50 to 1.0 and 0.55 to 0.92, respectively. We found no deviations from Hardy–Weinberg equilibrium, nor did we find linkage disequilibrium between pairs of loci after Bonferroni correction. We found no evidence for large allele dropouts or stuttering, although null alleles were detected for loci Ltb17 and Ltb4 in Alcobaca population. These markers will be useful for resolving fine-scale population genetic structure, especially in contact zones between highly divergent lineages of L. boscai.

Development and characterization of 15 polymorphic tetranucleotide microsatellite loci for Townsend’s big-eared bat (Corynorhinus townsendii) and cross amplification in Rafinesque’s big-eared bat (Corynorhinus rafinesquii)

Townsend’s big-eared bat, Corynorhinus townsendii, is a species of conservation concern with two subspecies (C. t. ingens, C. t. virginianus) listed as federally endangered. We developed and characterized 15 tetranucleotide microsatellite loci across four of the five C. townsensdii subspecies and Rafinesque’s big-eared bat, C. rafinesquii, which is also a species of conservation concern. Most loci were highly polymorphic (polymorphic information content >0.5) in all C. townsendii subspecies analyzed and in C. rafinesquii. The markers presented in this study will be useful to those managing populations of Townsend’s and Rafinesque’s big-eared bats.

Isolation and characterization of 13 tetranucleotide microsatellite loci in the Stone marten (Martes foina)

We isolated and characterised 13 novel polymorphic tetranucleotide microsatellite loci in the stone marten (Martes foina) from genomic libraries enriched for (AAAG)n and (AGAT)n repeat sequences. Thirty-three individuals were genotyped and the number of alleles per locus recorded varied between two and eight and their observed and expected heterozygosities ranged from 0.242 to 0.879 and from 0.335 to 0.817, respectively. All genotypic frequencies were in Hardy–Weinberg equilibrium and no linkage disequilibrium was detected between pairs of loci.

Isolation and characterization of 11 tetranucleotide microsatellite loci in the Egyptian mongoose (Herpestes ichneumon)

We report the isolation of 11 polymorphic tetranucleotide microsatellite loci in the Egyptian mongoose (Herpestes ichneumon). In a sample of 27 individuals, we observed between 4 and 7 alleles per locus and their observed and expected heterozygosities ranged from 0.37 to 0.85 and from 0.44 to 0.79, respectively. All genotypic frequencies conformed to Hardy-Weinberg equilibrium expectations and there were no instances of linkage disequilibrium detected between pairs of loci.

Isolation and characterization of polymorphic tetranucleotide microsatellite loci in the chestnut‐crowned babbler (Pomatostomus ruficeps)

We describe 18 microsatellite markers isolated in the cooperatively breeding chestnut-crowned babbler (Pomatostomus ruficeps). The number of alleles ranged from seven to 16 per locus (mean N(a) =10.4±0.54 SE) and the expected heterozygosity ranged from 0.732 to 0.889 (mean H(E) =0.836±0.01 SE). Three of the 18 loci exhibited significant heterozygote deficiency, but the remaining 15 will be used to analyse population genetic structure and the mating system of this highly social species.

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.

Isolation and characterization of microsatellite markers in tree lizards (Urosaurus ornatus)

Tree lizards (Urosaurus ornatus) are a highly accessible species useful in testing theories of life-history evolution and behavioural ecology. A polymerase chain reaction-based method was used to isolate nine polymorphic tetranucleotide microsatellite loci from the genome of these lizards. The isolated loci displayed 5 to 9 alleles in the populations screened. Observed heterozygosity ranged from 0.333 to 0.900. Results indicate these loci now provide a basis to study mate choice, parentage, and population genetic structure within this species.

Development of tetranucleotide microsatellite markers for the cushion star, Asterina gibbosa, and cross‐species amplification

We describe nine polymorphic tetranucleotide microsatellite loci from the starfish, Asterina gibbosa. Loci were isolated from a partial genomic library that had been enriched for AAAC repeat sequences. Number of alleles per locus ranged from two to 14 in a sample of 85 individuals from three populations (two from Spain and one from the UK). Observed and expected heterozygosities per population ranged from 0.000 to 0.400 and from 0.040 to 0.784, respectively. All loci presented significant heterozygote deficits in one or more populations. Eight of these loci were amplified and variable in A. pancerii and A. phylactica. These loci will be used to study population structure in A. gibbosa.