Development of 15 polymorphic microsatellite loci from <Emphasis Type="Italic">Garrulax morrisonianus</Emphasis> (Timaliidae), an endemic avian species of Taiwan

Garrulax morrisonianus, an endemic avian species of Taiwan, inhabits evergreen forests at high elevations of 2,000–3,952 m. In this study, we developed 15 microsatellite primer pairs for genetic study. These markers were screened for 52 samples collected from wild populations of different geographical regions. The number of alleles ranged from 4 to 13. The expected (H _E) and observed (H _O) heterozygosities were 0.430–0.725 and 0.000–0.500, respectively. All loci were significantly deviated from Hardy-Weinberg equilibrium due to the heterozygote deficiency. The authors Tzen-Yuh Chiang, Sao-Cheng Lao, Ya-Fu Lee contributed to the study equally.     

Isolation and characterization of 11 polymorphic microsatellite loci from <Emphasis Type="Italic">Fatsia polycarpa</Emphasis> (Araliaceae), an element of evergreen forests in Taiwan

Fatsia polycarpa Hayata (Araliaceae) is an element of evergreen forests at middle elevations in Taiwan, also representing the southern limitation in distribution of the genus. In this study, we described the development of 11 microsatellite loci from F. polycarpa in Taiwan for genetic studies. These new markers were tested in 20 individuals of 5 populations of the woody species. The number of alleles ranged from 9 to 18. The expected (H _E ) and observed (H _O ) heterozygosities were 0.80–0.95 and 0.10–0.75, respectively. Six of the 11 microsatellite loci are significantly deviated from Hardy–Weinberg expectations likely due to the population structure within samples. No linkage disequilibrium were observed from pairwise comparisons of loci. Chi-Chun Huang, Kuo-Hsiang Hung, and Tsai-Wen Hsu contributed equally to this article.     

Isolation and characterization of 12 microsatellite loci from <Emphasis Type="Italic">Suzukia shikikunensis</Emphasis> (Lamiaceae), a genus endemic to Taiwan and Ryukyus

Suzukia shikikunensis Kudo is an endemic plant in Taiwan and suffers habitat destruction caused by human overexploitation. In this study, we developed 12 microsatellite primer pairs for genetic study. These markers were screened for 24 samples collected from wild populations distributed in Taiwan, and for a sister species S. luchuensis collected from Yonaguni and Lutao Islands. The number of alleles ranged from 5 to 14. The expected (H _E) and observed (H _O) heterozygosities were 0.65–0.922 and 0–0.625, respectively. All loci were significantly deviated from Hardy–Weinberg equilibrium due to the heterozygote deficiency. These primers amplifying microsatellites in the two species may provide a useful tool for population genetics to establish conservation strategy.     

Isolation and characterization of 11 microsatellite loci from Camellia sinensis in Taiwan using PCR-based isolation of microsatellite arrays (PIMA)

We report 11 novel microsatellite primer pairs for the wild tea, Camellia sinensis (L.) O. Kuntze forma formosensis Kitamura. These simple sequence repeat markers were tested in 24 samples collected from wild tea populations, and in cultivars and C. japonica. The number of alleles ranged from 4 to18. The expected (H _E) and observed (H _O) heterozygosity were 0.687–0.946 and 0.042–0.792, respectively. All loci were significantly deviated from Hardy-Weinberg expectations due to the heterozygote deficiency, indicating a dramatic loss of genetic polymorphisms in the rare species. Significant LD was discovered in most loci. These primers may provide a tool for understanding demography and population structure in wild tea.     

Isolation and characterization of microsatellite loci from a potential biofuel plant Miscanthus sinensis (Poaceae)

Miscanthus sinensis (Poaceae) is typical of many dominant grasses of East Asia. Due to its characteristics of fast growth and high biomass, Miscanthus, a C4 plant, has been long explored for the potential usage as biofuel. In this study, we described the development of nine microsatellite loci from M. sinensis for genetic studies. These new markers were tested in 25 individuals of five populations in three varieties. The number of alleles ranged from 3 to 8. The expected (H _E) and observed (H _O) heterozygosities were 0.37–0.78 and 0.05–0.52, respectively. All microsatellite loci are significantly deviated from Hardy–Weinberg expectations likely due to the population structure within samples. Linkage disequilibrium between four loci pairs were observed from the total 36 pairwise comparisons of loci. Cross-species transferability revealed that all loci can be applied to its closely related species, M. floridulus.     

Isolation and characterization of microsatellite loci in the endangered freshwater fish <Emphasis Type="Italic">Varicorhinus alticorpus</Emphasis> (Cyprinidae)

Varicorhinus alticorpus (Cyprinidae) is an endangered freshwater fish with a scattered distribution in the southern and eastern regions of Taiwan. We described the development of nine polymorphic microsatellite loci in V. alticorpus for genetic studies. These new markers were tested in 20 individuals of the rare species. The number of alleles ranged from 4 to 13. Expected (H _E) and observed (H _O) heterozygosities ranged from 0.692 to 0.892 (averaged at 0.821) and from 0.000 to 0.350 (averaged at 0.088), respectively. All loci are significantly deviated from Hardy–Weinberg expectations due to the heterozygote deficiency. This suite of highly polymorphic microsatellites provides the first chance to undertake a conservation program for this species in Taiwan. It is essential to evaluate its genetic diversity and the fine-scale population structure. Tzen-Yuh Chiang, Teh-Wang Lee, and Feng-Jiau Lin contributed equally to this work.     

Development and characterization of microsatellite loci in the endangered Tricyrtis ishiiana (Convallariaceae), a local endemic plant in Japan

Nine microsatellite loci were isolated and characterized for the endangered and tertiary relict perennial herb, Tricyrtis ishiiana. The number of alleles ranged from 2 to 33. The expected (H _E) and observed (H _O) heterozygosities were 0.207–0.944 and 0.215–0.813, respectively, from 96 individuals on one population. Five loci exhibited significantly fewer heterozygotes than expected under Hardy–Weinberg equilibrium (P < 0.05). These primers amplifying microsatellites in this species may provide a useful tool for population genetics to establish conservation strategy.     

Development and characterization of microsatellite markers for an endangered shrub, <Emphasis Type="Italic">Ammopiptanthus mongolicus</Emphasis> (Leguminosae) and cross-species amplification in <Emphasis Type="Italic">Ammopiptanthus nanus</Emphasis>

We developed 11 polymorphic microsatellite markers for an endangered shrub Ammopiptanthus mongolicus using the Fast Isolation by AFLP of Sequences Containing repeats protocol. Polymorphism of each locus was assessed in 24 individuals from five natural populations of A. mongolicus. The average allele number of these markers was 4.2, ranging from 2 to 7. The observed (H _O) and expected (H _E) heterozygosities were 0.044–0.956 and 0.044–0.827, respectively. Further assessment in the congeneric species A. nanus revealed that nine of eleven loci were successfully amplified, but only two loci showed polymorphism. These markers provide powerful tools for the conservation genetics studies of A. mongolicus.     

Isolation and characterization of microsatellite markers from Brown Eared-pheasant (Crossoptilon mantchuricum)

The Brown Eared-pheasant (Crossoptilon mantchuricum) is an endangered species endemic to China. Here we developed eight microsatellite loci using a modified biotin-capture method. In the analyses of 28 individuals sampled, the number of alleles per locus ranged from four to nine. The expected (H _E) and observed (H _O) heterozygosities were 0.466–0.825 and 0.619–0.847, respectively. Results that eight microsatellite loci were highly polymorphic indicated that these markers are sufficiently powerful to address such questions as genetic diversity and population genetic structure of C. mantchuricum.     

Development and characterization of new microsatellite markers for ginseng (<Emphasis Type="Italic">Panax ginseng</Emphasis> C. A. Meyer)

Panax ginseng C.A. Meyer, commonly known as Korean or Asian ginseng, is a perennial herb native to Korea and China. Its roots are highly prized for several medicinal properties. The present study describes development and characterization of twenty-two polymorphic microsatellite markers for this species. A total of 99 alleles were detected with an average of 4.5 alleles per locus across 20 accessions. Values for observed (H _O ) and expected (H _E ) heterozygosities ranged from 0.05 to 1.00 and from 0.18 to 0.73, respectively. Eleven loci deviated from Hardy–Weinberg equilibrium (P < 0.001). Significant (P < 0.05) heterozygote deficiency was observed at 13 loci. Exact test for linkage disequilibrium showed significant values (P < 0.05) between 12 pairs of loci. These microsatellite markers provide powerful tools for understanding population and conservation genetics of this species and also for genetic differentiation and authentication of different Panax species being used in commercial ginseng products.     

Isolation and characterisation of microsatellite loci in Calystegia soldanella (Convolvulaceae), an endangered coastal plant isolated in Lake Biwa,Japan

Eight microsatellite loci of Calystegia soldanella useful for comparisons of the genetic structure of isolated populations in the ancient Lake Biwa and coastal populations in Japan were isolated and characterised. The number of alleles ranged from 2 to 5. The expected (H _E) and observed (H _O) heterozygosities were 0.097–0.583 and 0.000–0.380, respectively, from 100 individuals from Lake Biwa and coastal populations. Seven of the eight loci exhibited significantly fewer heterozygotes than expected based on the Hardy–Weinberg equilibrium (P < 0.05). These primers amplifying microsatellites in C. soldanella may provide a population genetics tool useful in the establishment of a conservation strategy.     

Isolation and characterization of eight microsatellite loci from <Emphasis Type="Italic">Lycorma delicatula</Emphasis> (White) (Hemiptera: Fulgoridae) for population genetic analysis in Korea

Lycorma delicatula (White) is native to China but is becoming an important insect pest in Korea. Polymorphic DNA markers like microsatellites are widely used for characterizing dispersal patterns and capacity of invasive insect pests which can contribute to designing effective management of the species. To facilitate such population genetic studies of L. delicatula in Korea, we isolated and characterized eight microsatellite loci for L. delicatula using a hybridization-biotin enrichment method. We further used these novel microsatellite loci to determine population genetic parameters for 33 L. delicatula specimens collected from Cheonan, South Korea where outbreaks of this species were first reported in Korea. The number of alleles per locus ranged from three to ten, with an average of 6.25. The mean expected (H _E) and observed heterozygosities (H _O) were 0.575 and 0.626, respectively. The eight loci showed no deviation from Hardy–Weinberg equilibrium according to the adjusted significance threshold (P = 0.00625), and there was no linkage disequilibrium between each pair of these eight markers. Bayesian cluster analysis using the program structure revealed no evidence of genetic structuring in L. delicatula samples from Cheonan. These new microsatellite markers will be widely applicable to future ecological genetic studies of L. delicatula, including assessment of the level of gene flow and genetic connectivity among populations that are necessary for effective management and monitoring of the species.     

Isolation and characterization of microsatellite loci in Fagus longipetiolata Seem. (Fagaceae)

Eleven microsatellite loci have been developed from Fagus longipetiolata and the loci were characterized for 21 individuals. All eleven loci were polymorphic, with 2–8 alleles and an average of 4.8 per locus. The observed (H _O) and expected (H _E) heterozygosities were 0.053–0.714 and 0.355–0.856, respectively. There was significant deviation from Hardy–Weinberg equilibrium at two loci. No locus pair had significant linkage disequilibrium. Cross-species amplifications of the markers were also tested in three other congeneric species.     

Isolation and characterization of microsatellite loci in Shortia rotundifolia (Diapensiaceae), an endangered relict plant on the Ryukyu Islands and Taiwan

Ten microsatellite loci were isolated and characterized for Shortia rotundifolia, a relict temperate plant on the subtropical Ryukyu Islands and Taiwan. The number of alleles ranged from 2 to 21. The expected (H _E) and observed (H _O) heterozygosities were 0.027–0.877 and 0.027–0.622, respectively, from 74 individuals on Iriomotejima Island of Ryukyus. One locus exhibited significantly fewer heterozygotes than expected under Hardy–Weinberg equilibrium (P < 0.05). These primers amplifying microsatellites in this species may provide a useful tool for population genetics to establish conservation strategy.     

Isolation and characterization of 18 polymorphic microsatellite loci from freshwater pearl mussel (<Emphasis Type="Italic">Cristaria plicata</Emphasis>)

Cristaria plicata was an important freshwater mussel for pearl culture in China. 18 polymorphic microsatellite markers were isolated and characterized using (CA)₁₅-enriched genomic library of C. plicata. These loci showed high levels of genetic polymorphism testing on 60 individuals sampled from Poyang Lake of Jiangxi Province, China. The number of alleles per locus ranged from 4 to 18. The expected (H _E) and observed heterozygosities (H _O) were 0.7232–0.8961 and 0.0000–1.0000, respectively. Four microsatellite loci were significantly deviated from Hardy–Weinberg equilibrium and no linkage disequilibrium was found. These microsatellite loci will be useful for assessment of genetic diversity and population structure in C. plicata.     

Development and characterisation of microsatellite loci in Farfugium japonicum (Asteraceae)

Eight microsatellite loci for the perennial herb Farfugium japonicum, including the rheophytic variety luchuense endemic to riparian areas of the Ryukyu Islands, Japan, were isolated and characterised. The number of alleles ranged from 5 to 14. The expected (H _E) and observed (H _O) heterozygosities were 0.344–0.885 and 0.121–0.754, respectively, from 69 individuals in one population. Six loci exhibited significantly fewer heterozygotes than expected under Hardy–Weinberg equilibrium (P < 0.05). The primers amplifying microsatellite sequences in F. japonicum may provide a population genetics tool useful in the establishment of a conservation strategy.     

Isolation and characterization of eight microsatellite loci from the endangered plant species Hypochaeris salzmanniana (Asteraceae)

We report the isolation and characterization of eight microsatellite loci for Hypochaeris salzmanniana, an endangered species endemic to the southwestern coast of Spain and the Atlantic coast of Morocco. A total of 32 alleles were detected across a sample of 45 individuals, with an average number of 4.0 alleles per locus. The average polymorphic information content (PIC) was 0.533 and the observed (H _O) and expected (H _E) heterozygosity values varied from 0.022 to 0.978 and from 0.434 to 0.759, respectively. Five loci exhibited significant deviation from Hardy–Weinberg equilibrium (P ≤ 0.001) and three pairs of loci showed significant linkage disequilibrium (P ≤ 0.01). The eight loci were tested for transferability in three others species (H. arachnoidea, H. glabra, and H. radicata) belonging to the same section of H. salzmanniana. With the exception of locus Hsalz7, all loci successfully amplified in the three species. These preliminary data confirm the usefulness of microsatellite markers for assessing the ecology and genetic structure of H. salzmanniana and to understand the evolution of species within the section Hypochaeris.     

Isolation and characterization of 15 microsatellite loci from Caridina gracilipes (Atyidae, Decapoda)

Caridina gracilipes, an atyid shrimp of Taiwan, inhabits the fresh and brackish waters. Its life history can be classified into amphidromous and landlocked types. Recent human’s transportation has caused concerns of genetic contamination between these long evolved lineages and ecological instabilities. Molecular markers with high sensitivities are required for detecting the possible introgression due to human disturbance. In the study, we described the development of 15 polymorphic microsatellite loci in C. gracilipes for genetic and ecological studies. These new markers were tested in 20 individuals of three populations. The number of alleles ranged from 6 to 17. Expected (H _E) and observed (H _O) heterozygosities ranged from 0.653 to 0.954, averaged at 0.865, and from 0.000 to 0.4, averaged at 0.146, respectively. All loci are significantly deviated from Hardy–Weinberg expectations due to the heterozygote deficiency. Significant genetic differentiation among populations was detected based on F _ST values, which were estimated from 0.82 to 0.88. Chiao-Chuan Han and Cheng-Sheng Chang are contributed equally to this work.     

Isolation and characterization of 12 microsatellite loci for Michelia coriacea (Magnoliaceae), a critically endangered endemic to Southeast Yunnan, China

By using a modified biotin–streptavidin capture method, a total of 12 microsatellite loci were developed and characterized for Michelia coriacea (Magnoliaceae), a critically endangered endemic to Southeast Yunnan, China. The number of alleles (A) ranged from two to six in 30 samples of this species. The ranges of observed (H _O ) and expected (H _E ) heterozygosities were 0.033–0.8000 and 0.033–0.7910, respectively. Cross-species amplification in M. yunnanensis showed that a subset of these markers holds promise for congeneric species. These polymorphic SSR markers would be useful tools for population genetics studies on M. coriacea and other congeneric species.     

Isolation and characterization of first microsatellite markers for the noble crayfish, <Emphasis Type="Italic">Astacus astacus</Emphasis>

Noble crayfish (Astacus astacus) is listed as vulnerable by the IUCN Red List of Threatened Species in Europe. However, very little is known about genetic diversity and structuring of noble crayfish populations, mainly because of the lack of informative genetic markers. We describe the isolation and characterization of the first microsatellite markers for this species, which were obtained by screening 4,000 recombinant clones. Eight loci revealed polymorphisms in a panel of 172 individuals from seven populations in Northern Europe. Number of alleles per locus ranged from two to 10 (average 4.4) and heterozygosity levels among populations varied from 0 to 0.80 for H _o and from 0 to 0.72 for H _e.