Karyological analyses of 33 species of the tribe Ophiopogoneae (Liliaceae) from Southwest China

Chromosome numbers and karyotypes of 26 Ophiopogon species, 2 Liriope species and 5 Peliosanthes species of the family Liliaceae from Southwest China, were investigated. The study revealed a detailed picture of chromosome features and their pattern of karyotype variation in Ophiopogoneae. Karyotype asymmetry in different species and different populations of the same species varied greatly due to different locality conditions. Our analyses may support the separately monophyly of Ophiopogon, Liriope and Peliosanthes.     

台湾海峡鲐鱼种群遗传结构

以往研究表明,台湾海峡的鲐鱼分属2个地理种群,即东海种群和闽南——粤东地方种群.为研究这2个种群的遗传结构,对鲐鱼闽东(30尾)和闽南(30尾)种群进行了AFLP分析,8对选择性引物在2个种群60个个体中,共扩增出497个位点,其中多态位点343个.闽东和闽南种群的多态位点比例、Nei遗传多样性指数和Shannon遗传多样性指数分别为57.75％、64.59％,0.1779、0.2123,0.2725和0.3228,2个种群的遗传多样性处于同一水平.与其他鱼类对比显示,台湾海峡鲐鱼种群的遗传多样性水平高.生境广及生命周期短被认为是台湾海峡鲐鱼具有较高遗传变异水平的原因;基因分化系数Gst、Shannon遗传多样性指数和AMOVA分析均显示鲐鱼的遗传变异主要来源于种群内,而种群间无明显的遗传分化.Nm显示2个种群间基因交流频繁.种群的显性基因型频率分布显示2个种群有基本相同的种群遗传结构.结果表明,鲐鱼闽东和闽南种群间无明显的遗传差异.幼体较强的扩散能力、海洋环流及洄游特性可能是造成台湾海峡鲐鱼种群间遗传同质性较高的原因.     

Phytochemical constituents and chemotaxonomic study of Polygonatum prattii Baker

Fifteen compounds were obtained from Polygonatum prattii Baker, which could be divided into four steroids, nine flavonoids, one lignanoid and one acidamide. The chemotaxonomic significance of P. prattii and the some other species among Asparagaceae was also discussed. As a result, Polygonatum may have close relationships with Liriope, Disporopsis, Ophiopogon and Aspidistra. Additionally, homoisoflavanones and steroids can be recognized as typical biomarkers for the above genus.     

Development, characterization, and inheritance of 113 novel EST-SSR markers in the Pacific oyster (Crassostrea gigas)

A total of 113 novel EST-derived simple sequence repeat (EST-SSR) markers were developed in the Pacific oyster (Crassotrea gigas). Polymorphisms of these markers were evaluated in a wild population of 30 individuals. The number of alleles per locus ranged from 2 to 27 with an average of 6.3, and the observed and expected heterozygosities varied from 0 to 0.9667 and from 0.0333 to 0.9701, respectively. Mendelian segregations were tested for 24 of the markers that were polymorphic in one family produced by single-pair mating. Null alleles were discovered at four loci. Nine tests of segregation ratios revealed significant departures from expected Mendelian ratios. As a useful addition to the collection of the microsatellites that are now available for C. gigas, these EST-SSR markers will help the advance in investigation of QTL mapping and genetic diversity in this species.     

Characterization of 11 novel polymorphic microsatellite loci in the threatened Korean loach,Iksookimia koreensis,isolated using a next-generation sequencing method

To investigate the genetic diversity and population structure of Iksookimia koreensis, we characterized 11 novel polymorphic microsatellite markers developed using next-generation sequencing. The number of alleles per locus ranged from 4 to 10 (average = 6.26). Observed and expected heterozygosities ranged from 0.333 to 0.866 and from 0.375 to 0.866, respectively. No loci showed significant deviation from Hardy–Weinberg equilibrium (HWE). These loci were also used successfully to study the genetic diversity of a closely related species, Iksookimia longicorpa. Four of the 11 loci amplified in the two species showed different allele frequency and distribution, indicating deep genetic divergence between I. koreensis and I. longicorpa. The newly developed microsatellite markers reported here will provide a useful tool for examining gene flow, population genetic structure, and genetic diversity of these species.     

Eleven new microsatellite markers in jack mackerel (Trachurus japonicus) derived from an enriched genomic library

Jack mackerel (Trachurus japonicus, Carangidae) are a commercially important fisheries resource in Korea. To understand patterns of genetic variation for conservation and management efforts, we developed microsatellite DNA markers fromT. japonicus. We report the isolation and characterization of eleven microsatellite loci isolated using an enrichment method based on magnetic/biotin capture of microsatellite sequences from a size-selected genomic library. To characterize each locus, 50 individuals from a naturalT. japonicus population in southern Korea were genotyped. All loci except one, KTJ38, were polymorphic with an average of 14 alleles per locus (range 6–23). The mean observed and expected heterozygosities were 0.70 (range 0.46–0.92) and 0.81 (range 0.49–1.00), respectively. Significant deviation from Hardy-Weinberg equilibrium was observed at three loci, KTj3, KTJ20 and KTJ28. Such high variability indicates that these microsatellites are useful markers for high-resolution analysis for population gemetic studies.     

Characterization of expressed sequence tag‐derived microsatellites from the kuruma prawn,Marsupenaeus japonicus

The kuruma prawn, Marsupenaeus japonicus, is a commercially important benthic marine crustacean in East Asia. Understanding the species’ population structures will be very important for its proper stock assessment and management strategy. Herein, 13 polymorphic microsatellite loci originating from expressed sequence tag libraries of M. japonicus were isolated and characterized. Number of alleles per locus ranged from two to 15, observed and expected heterozygosities ranged from 0.250 to 0.906 and from 0.310 to 0.932, respectively. The adequate level of variability within the population renders these microsatellite loci ideal markers for population genetic analysis of M. japonicus.     

Discrimination and genetic diversity of cultivated and wild safflowers (Carthamus spp.) using EST-microsatellites markers

In spite of being one of the major oilseed crops, little is known about genetic diversity and relationships between species of safflower. In this study EST-SSR markers were used to evaluate and characterize 42 genotypes from six species including Carthamus tinctorius, Carthamus palaestinus, Carthamus oxyacanthus, Carthamus lanatus, Carthamus dentatus, and Carthamus boissieri. Thirty three primer pairs produced 123 polymorphic bands with 2–8 alleles per locus. The EST-SSR markers showed different level of gene diversity. The highest Polymorphism Information Content (PIC) values were observed for primers EL510507 and EL390720 (0.49 and 0.45, respectively). The highest genetic diversity and heterozygosity were observed for C. oxyacanthus. Both cluster and principal coordinate analysis (PCoA) clearly separated species into distinct groups. Within each species the accessions were clustered in different subgroups that mainly supported the known origins. The result showed that C. palaestinus had the most genetic similarity with cultivated safflower and C. oxyacanthus was next in this respect. In general, EST-SSR markers effectively revealed the genetic relationships and diversity of Carthamus species. This information is valuable for safflower improvement since C. palaestinus and C. oxyacanthus are both crossable with the cultivated species C. tinctorius.     

SSR markers development and their application in genetic diversity evaluation of garlic (Allium sativum) germplasm

Garlic (Allium sativum), an asexually propagated vegetable and medicinal crop, has abundant genetic variation. Genetic diversity evaluation based on molecular markers has apparent advantages since their genomic abundance, environment insensitivity, and non-tissue specific features. However, the limited number of available DNA markers, especially SSR markers, are insufficient to conduct related genetic diversity assessment studies in garlic. In this study, 4372 EST-SSR markers were newly developed, and 12 polymorphic markers together with other 17 garlic SSR markers were used to assess the genetic diversity and population structure of 127 garlic accessions. The averaged polymorphism information content (PIC) of these 29 SSR markers was 0.36, ranging from 0.22 to 0.49. Seventy-nine polymorphic loci were detected among these accessions, with an average of 3.48 polymorphic loci per SSR. Both the clustering analyses based on either the genotype data of SSR markers or the phenotypic data of morphological traits obtained genetic distance divided the 127 garlic accessions into three clusters. Moreover, the Mantel test showed that genetic distance had no significant correlations with geographic distance, and weak correlations were found between genetic distance and the phenotypic traits. AMOVA analysis showed that the main genetic variation of this garlic germplasm collection existed in the within-population or cluster. Results of this study will be of great value for the genetic/breeding studies in garlic and enhance the utilization of these garlic germplasms.     

Development of new EST-derived SSRs in Salvia miltiorrhiza (Labiatae) in China and preliminary analysis of genetic diversity and population structure

Salvia miltiorrhiza, known as Dan-shen in China, is a well-known and important medicinal herb in Asia. Its root and rhizome has been used to treat cardiovascular diseases in China for centuries. Recently, it had been widely cultivated in China due to a rapid decline in wild resources. However, the germplasm resources of different cultivated regions are extremely variable. Here, new simple sequence repeats (SSRs) from expressed sequence tags (ESTs) were mined to analyze the frequency and distribution of SSRs, assess the validity of developed EST-SSR marker and identify new EST-SSR marker for the preparatory investigation of the population structure in cultivated S. miltiorrhiza. This study showed that 5160 unique ESTs were accumulated from database, 760 SSRs combined with 41 types of SSR motifs were mined. High polymorphism and transferability were presented in newly developed EST-SSR marker. It also showed that cultivated S. miltiorrhiza were mainly divided into three large groups. The results suggested that S. miltiorrhiza may has high abundance of SSRs in transcribed regions of genome and the developed EST-SSR marker may be useful for analysis of genetic diversity and population structure.     

Population genetic structure of the sea bass (Lateolabrax japonicus) in Korea based on multiplex PCR assays with 12 polymorphic microsatellite markers

Population genetics has been recognized as a key component of policy development for fisheries and conservation management. In this study, natural sea bass (Lateolabrax japonicus) populations in three ocean basins in Korea were assessed using multiplex assays with 12 highly polymorphic microsatellite loci; 203 alleles and similarly high levels of genetic diversity [mean number of alleles (N_A) = 14.43, mean expected heterozygosity (He) = 0.84] were detected. All populations showed significant heterozygote deficiency at four loci, which could be explained by the presence of null alleles. The genetic population subdivision was low and was significantly different according to F-statistics (overall F _ST = 0.003, R _ST = 0.005). However, this substructure was not supported by an analysis of molecular variance test, analyses of isolation by distance or Bayesian analysis. The passive dispersal of eggs/larvae via the main currents appears to facilitate gene flow. The possibility of a recent genetic bottleneck was observed in all three populations of L. japonicus, indicating that overfishing and degradation of the environment in recent years has led to a decline in the sea bass populations in Korea. Our study demonstrates that sea bass in Korea do not appear to be genetically partitioned and should be managed as a single unit; however, the potential for a rapid loss of genetic diversity remains. Information regarding the genetic characteristics of Korean sea bass populations has important implications for fishery management and conservation efforts and will aid in the sustainable exploitation of fishing resources and the preservation of biodiversity.     

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 thirteen polymorphic microsatellite loci for Lithocarpus glaber

We isolated and characterized polymorphic microsatellite loci in Lithocarpus glaber (Fagaceae), an evergreen broadleaved monoecious tree, to provide tool for analyzing genetic structure and diversity. Thirteen polymorphic microsatellite loci were developed and tested in two L. glaber populations. The number of alleles per locus varied from 2 to 19. The observed and expected heterozygosities within populations were 0.037–0.833 and 0.316–0.931, respectively. Four loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction in each population and no significant linkage disequilibrium between pairs of loci was found. These polymorphic loci showed high levels of polymorphism within tested populations and will be useful in further population genetic studies.     

Development of EST-SSR markers through data mining and their use for genetic diversity study in Indian accessions of Jatropha curcas L.: a potential energy crop

Jatropha curcas L. is gaining importance as a potential energy crop. However, lack of sufficient numbers of molecular markers hinder current research on crop improvement in Jatropha. The expressed sequences tags (EST) sequences deposited in public databases, offers an excellent opportunity to identify simple sequence repeats (SSRs) through data mining, for further research on molecular breeding. In the present study 42,477 ESTs of J. curcas were screened, out of which 5,673 SSRs were identified with 48.8 % simple (excluding mononucleotide repeats) and 52.2 % compound repeat motifs. Amongst these repeat motifs, dinucleotide repeats were abundant (26.5 %), followed by trinucleotide (23.1 %) and tetranucleotide repeats (0.8 %). From these microsatellites, 32 EST-SSR (genic microsatellite) primer pairs were designed. These primers were used to analyze the genetic diversity among 42 accessions collected from different parts of India. Out of the 32 EST-SSR primers, 24 primer pairs exhibited polymorphism among the genotypes, with amplicons varying from one to eight, giving an average of 2.33 alleles per polymorphic marker. Polymorphic information content value ranged from 0.02 to 0.5 with an average of 0.402 indicating moderate level of informativeness within these EST-SSRs markers. The EST-SSR markers developed here will serve as a valuable resource for genetic studies, like linkage mapping, diversity analysis, quantitative trait locus/association mapping, and molecular breeding. The current study also revealed low diversity in the screened Indian Jatropha germplasm. Therefore, the future efforts must be made to broaden the gene pool of Jatropha for the creation of genetic diversity that can be further used for crop improvement through breeding.     

山麦冬及其近缘种DNA条形码研究

通过分析山麦冬及其近缘种cpDNA trnL-F、psbA-trnH间隔区序列特点,探讨trnL-F、psbA-trnH序列作为山麦冬及其近缘种DNA条形码的潜力。分别对两者进行PCR扩增后,并进行纯化测序。其中山麦冬及其近缘种9个物种18个样品的trnL-F序列长度为355~356 bp,在山麦冬属内序列完全一致,但在沿阶草属中存在特异变异位点,可对麦冬与沿阶草两物种进行鉴别。而psbA-trnH序列长度为543~544 bp,仅在麦冬中存在特异鉴别位点。结果表明,trnL-F、psbA-trnH序列由于进化速率低、保守性强,仅适合麦冬类植物属间鉴别的DNA条形码,而在属下水平的应用有一定的局限。     

EST-SSR markers derived from Laminaria digitata and its transferable application in Saccharina japonica

The public availability of numerous expressed sequence tag (EST) enables EST-based SSR (simple sequence repeat) markers to be widely used for genetics and breeding studies. In the present study, EST-SSR markers were developed from ESTs of Laminaria digitata and were transferred to the non-congeneric species Saccharina japonica. Among the 2,668 non-redundant ESTs, 83 (3.1%) ESTs containing SSR were identified totally, with an average of one SSR per 13.6 kb. Analysis of SSR motifs revealed that the trinucleotide and tetranucleotide were major motifs, accounted for 44.58% and 16.87%, respectively. Based on the 83 ESTs containing SSR, we designed 45 pairs of primers in the flanking regions of the SSR, of which 13 pairs showed polymorphism in a wild S. japonica population, and the mean alleles per locus was 3.6 (ranging from 2 to 6). The observed (Ho) and expected (He) heterozygosities of these EST-SSRs were 0.234–0.632 and 0.260–0.635, respectively. All loci were in Hardy–Weinberg equilibrium in the wild population and no linkage disequilibrium was detected among loci. The obtained EST-SSR markers can facilitate and promote related research such as ecological investigation, genetic diversity assessment and breeding practice of S. japonica as well.     

Genetic diversity of switchgrass and its relative species in Panicum genus using molecular markers

Switchgrass (Panicum virgatum), a warm season C4 grass, is a promising crop for bioenergy-dedicated biomass production. Understanding of genetic diversity within Panicum genus will facilitate switchgrass breeding. Genetic relationships of 22 Panicum species from six continents including ninety-one USDA germplasm accessions were investigated by Sequence-Related Amplified Polymorphism (SRAP) and Expressed Sequence Tags-Simple Sequence Repeat (EST-SSR) markers. Eight hundred and twenty-six markers from 28 pairs of SRAP and 25 pairs of EST-SSR Primers were used to differentiate between accessions of a bulk of 25 genotypes. The results showed that there was high genetic diversity found in Panicum species. Most genetic variation was present among the different species and cluster analysis indicated that all the Panicum accessions could be distinguished by SRAP or EST-SSR. Dendrogram results reflected the phylogenetic relationships between Panicum species and Panicum amarum was found to be the closest species to switchgrass. Comparison between molecular markers revealed that SRAP methods were considered more efficient than EST-SSR for screening Panicum accessions.     

Isolation and Characterization of Twenty Microsatellite Loci in Japanese Sea Cucumber (Stichopus japonicus)

Twenty microsatellite markers were first developed from the Japanese sea cucumber Stichopus japonicus using an enrichment protocol. Of the 20 microsatellite loci, 19 loci were polymorphic in the population examined. At these polymorphic loci, the number of alleles per locus varied from 2 to 15, and the observed heterozygosities ranged from 0.03 to 0.97, which is considerably higher than those previously found for allozymes. The high variability of the microsatellite markers identified in this study will make them excellent tools for genetic analyses of S. japonicus.