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Premise of the Study Simple sequence repeat (SSR) and expressed sequence tag (EST)–SSR markers were developed as tools for marker‐assisted selection of Chamaecyparis formosensis and for the molecular differentiation of cypress species. Methods and Results Based on the SSR‐enriched genomic libraries and transcriptome data of C. formosensis, 300 primer pairs were selected for initial confirmation, of which 19 polymorphic SSR and eight polymorphic EST‐SSR loci were chosen after testing in 92 individuals. The number of alleles observed for these 27 loci ranged from one to 17. The levels of observed and expected heterozygosity ranged from 0.000 to 1.000 and from 0.000 to 0.903, respectively. Most markers also amplified in C. obtusa var. formosana. Conclusions The developed SSR and EST‐SSR sequences are the first reported markers specific to C. formosensis. These markers will be useful for individual identification of C. formosensis and to distinguish cypress species such as C. obtusa var. formosana.

Olive (Olea europaea ssp. europaea) is the most important oil fruit crop in temperate areas, but the origin of the cultivated olive remains unclear. The existence of one or several domestication events in the Mediterranean Basin (MB) is still debated. We analyzed a dataset of 387 cultivated and wild accessions that were genotyped at 25 simple‐sequence repeat (SSR) loci. The sample represented genetic diversity at the geographic extremes of the MB. We inferred relationships among samples and also applied approximate Bayesian computation to estimate the most probable demographic model of our samples. Cultivated olives clustered into three different gene pools (Q1, Q2 and Q3), corresponding loosely to the west, central and eastern MB, respectively. Q1 consisted primarily of accessions from southern Spain, retained the fingerprint of a genetic bottleneck, and was closely related to accessions from the eastern MB. Q2 showed signs of recent admixture with wild olives and may derive from a local domestication event in the central MB. Overall our results suggest that admixture shaped olive germplasm and perhaps also local domestication events.

Premise of the study: Microsatellite primers were developed for Begonia luzhaiensis (Begoniaceae) to assess genetic diversity and population genetic structure. Methods and Results: Based on the transcriptome data of B. luzhaiensis, 60 primer pairs were selected for initial validation, of which 16 yielded polymorphic microsatellite loci in 57 individuals. The number of alleles observed for these 16 loci ranged from one to nine. The observed and expected heterozygosity ranged from 0.000 to 1.000 and from 0.000 to 0.804 with averages of 0.370 and 0.404, respectively. Five loci could be successfully amplified in B. leprosa. Conclusions: The expressed sequence tag–simple sequence repeat markers are the first specifically developed for B. luzhaiensis and the first developed in Begonia sect. Coelocentrum. These markers will be useful for future studies of the genetic structure and phylogeography of B. luzhaiensis.

Premise of the study: The first set of expressed sequence tag–simple sequence repeat (EST-SSR) markers were developed and characterized for Speranskia tuberculata (Euphorbiaceae), a traditional medicinal plant endemic to northern China, to explore the effects of recent habitat fragmentation on the genetic diversity and structure of this species. Methods and Results: In this study, a total of 18 novel polymorphic microsatellite (EST-SSR) markers were developed for S. tuberculata using high-throughput transcriptome sequencing. Analysis of 24 individuals of S. tuberculata from four natural populations revealed their robust polymorphic reliability. The number of alleles per locus ranged from two to 11, while the expected and observed heterozygosity per marker varied from 0.187 to 0.827 and 0.042 to 0.917, respectively. Of these markers, 13 showed good amplification results in the closely related species S. cantonensis. Conclusions: These newly generated SSR markers are expected to provide novel tools for genetic studies of S. tuberculata, which will contribute to the conservation and sustainable use of the species' wild genetic resources.

Premise Camellia reticulata, which is native to southwestern China, is an economically important plant belonging to the family Theaceae. We developed expressed sequence tag–simple sequence repeat (EST‐SSR) markers for C. reticulata, which can be used to investigate its genetic diversity, population structure, and evolutionary history. Methods and Results We detected 4780 SSRs in C. reticulata from Camellia RNA‐Seq data deposited in the National Center for Biotechnology Information's expressed sequence tags database (dbEST). Primer pairs for 70 SSR loci were designed and used for PCR amplification using 90 individuals from four populations of C. reticulata. Of these loci, 50 microsatellite markers were successfully identified, including 11 polymorphic markers. The allele number per locus ranged from two to seven (mean = 4.182), and the levels of observed and expected heterozygosity ranged from 0.044 to 0.567 and from 0.166 to 0.642, respectively. Eleven primer pairs amplified PCR products in three other species of Camellia (C. saluenensis, C. pitardii, and C. yunnanensis). Conclusions The set of microsatellite markers developed here can be used to study the genetic variation and population structure of C. reticulata and related species and thereby help to develop conservation strategies for this species.

Premise of the Study Expressed sequence tag–simple sequence repeat (EST‐SSR) markers were developed for Carex angustisquama (Cyperaceae) to investigate the evolutionary history of this plant that is endemic to solfatara fields in northern Japan. Methods and Results Using RNA‐Seq data generated by the Illumina HiSeq 2000, 20 EST‐SSR markers were developed. Polymorphisms were assessed in C. angustisquama and the closely related species C. doenitzii and C. podogyna. In C. angustisquama, many loci were monomorphic within populations; the average number of alleles ranged from one to five, and levels of expected heterozygosity ranged from 0.000 to 0.580, while all markers were polymorphic in a population of C. doenitzii. This indicates that low genetic polymorphism of C. angustisquama is likely due to the species’ population dynamics, rather than to null alleles at the developed markers. Conclusions These markers will be used to assess genetic diversity and structure and to investigate evolutionary history in future studies of C. angustisquama and related species.

Premise of the Study Ephedra sinica (Ephedraceae) is a gymnosperm shrub with a wide distribution across Central and Eastern Asia. It is widely cultivated as a medicinal plant, but its wild populations are monitored to determine whether protection is needed. Methods and Results Thirty‐six microsatellite markers, including 11 polymorphic markers, were developed from E. distachya RNA‐Seq data deposited in the National Center for Biotechology Information dbEST database. Among 100 genotyped E. sinica individuals originating from five different population groups, the allele number ranged from three to 22 per locus. Levels of observed and expected heterozygosity ranged from 0 to 0.866 (average 0.176) and 0 to 0.876 (average 0.491), respectively. Allelic polymorphism information content ranged from 0.000 to 0.847 (average 0.333). Cross‐species amplifications were successfully conducted with two related Ephedra species for all 11 di‐ or trinucleotide simple sequence repeats. Conclusions This study provides the first set of microsatellite markers for genetic monitoring and surveying of this medicinal plant.

Premise Saxifraga fortunei (Saxifragaceae) includes several infraspecific taxa that are ecologically and morphologically distinct. To investigate the evolutionary history of phenotypic polymorphisms in this species, we developed expressed sequence tag–simple sequence repeat (EST‐SSR) markers for S. fortunei. Methods and Results We developed 26 polymorphic markers based on transcriptome data obtained from Illumina HiSeq 2000. Within three populations of S. fortunei var. incisolobata, the number of alleles ranged from four to 25, and the levels of observed and expected heterozygosity ranged from 0.200 to 0.847 and from 0.209 to 0.930, respectively. Furthermore, all 26 loci showed transferability for S. fortunei var. obtusocuneata and S. fortunei var. suwoensis, and 18 loci were also successfully amplified in S. acerifolia. Conclusions These newly developed EST‐SSR markers will prove useful to infer the evolutionary history of S. fortunei var. incisolobata and its relatives in population genetic studies.

Premise of the Study Vitex negundo var. heterophylla (Lamiaceae) is a dominant shrub in the warm temperate zone of northern China. Expressed sequence tag–simple sequence repeat (EST‐SSR) markers were developed to investigate its genetic diversity and structure. Methods and Results We detected 12,075 SSRs in V. negundo var. heterophylla using transcriptome sequencing. Primer pairs for 100 SSR loci were designed and amplified in three populations of V. negundo var. heterophylla. Sixty loci were amplified, of which 14 were polymorphic. The number of alleles per locus ranged from two to 15, and levels of observed and expected heterozygosity ranged from 0.241 to 0.828 and from 0.426 to 0.873, respectively. All primer pairs amplified PCR products from V. rotundifolia but only four of them amplified products from Leonurus japonicus. Conclusions The identified EST‐SSR markers will be useful for future molecular and reproductive ecology studies of V. negundo var. heterophylla and V. rotundifolia.

Chinese white poplar (Populus tomentosa), an important commercial tree species for timber and pulp production in northern China, has been used to examine the individual genes and allelic diversity responsible for complex traits controlling growth and lignocellulosic biosynthesis. Taking advantage of the low degree of linkage disequilibrium (LD) within P. tomentosa association populations, we examined associations between 15 cellulose synthase (PtoCesA) genes and traits including growth and wood properties. Thirty-six novel simple sequence repeat (SSR) markers within PtoCesA genes were detected by re-sequencing and genotyped in an association population (460 individuals). Single-marker and haplotype-based LD approaches were used to identify significant marker–trait associations. Family-based linkage studies and real-time PCR testing were conducted to validate the functional significance of SSR variation. Fifteen single-marker associations from seven PtoCesA genes and nine haplotype-based associations within six genes were identified in the association population (false discovery rate Q < 0.05). Next, five SSR marker–trait associations (Q< 0.05) from four PtoCesA genes were successfully validated in a linkage mapping population (1200 individuals). The results imply a functional role for these genes in mediating wood properties, demonstrating the potential of combining single-marker and haplotype-based LD approaches to detect functional allelic variation underlying quantitative traits in a low-LD population.