Expressed sequence tag-simple sequence repeat-based molecular variance in two Salicornia (Amaranthaceae) populations

Salicornia spp is one of the most salt-tolerant vascular plants and is native to salt marshes and estuaries. We developed expressed sequence tag derived-simple sequence repeat (EST-SSR) markers for estimating genetic diversity and marker-assisted Salicornia breeding. Six polymorphic EST-SSRs of 40 detected 27 alleles, ranging from three to five alleles per locus. The average number of alleles per locus was 4.33 and 4.17, and the major allele frequency at locus DY529765 was high, being 0.859 and 0.857 in S. bigelovii and S. europea, respectively. Gene diversity, heterozygosity and polymorphism information content were highest at locus DY529950 and similar in these two species. Gene diversity increased with increase in the number of alleles that had a low major allele frequency at a locus. Six polymorphic loci effectively discriminated 46 taxa into three clusters via different analyses. Significant deviation of F(ST) from zero in three suggested populations for six loci indicated population differentiation and limited gene flow among them. A reduced median network established that taxon SB65 is primitive. SMART (simple modular architecture research tool) analysis of peptide sequences of six EST-SSRs showed that loci DY529765, DY529950 and EC906203 contained transmembrane, TLC, AgrB and NTR domains and might be involved in salinity stress tolerance. These EST-SSRs are a valuable resource for marker development and may be useful in marker-assisted Salicornia breeding.     

Genetic structure of Asian populations of Bombus ignitus (Hymenoptera: Apidae)

The genetic structure of seven mainland and island Asian populations of Bombus ignitus was investigated using nine microsatellite markers and the sequences of part of the mitochondrial cytochrome b (cytb) gene. While microsatellite markers showed high genetic variability, no sequence variation was found in the cytb gene fragment analyzed. The number of microsatellite alleles ranged from 9 to 24. Gene diversities per locus per population ranged from 0.378 to 0.992. Analysis of molecular variance (AMOVA) and most pairwise F(ST) values showed significant genetic differentiation between mainland and island populations. Cytb sequences data and microsatellite bottleneck tests indicated that almost all populations were subjected to recent bottlenecks. Our results suggest that B. ignitus populations diverged due to recent bottlenecks and geographic isolation.     

Development of compound microsatellite markers in red-tide-causing dinoflagellate Akashiwo sanguinea (Dinophyceae)

We isolated 13 polymorphic microsatellites from the red-tide causing dinoflagellate Akashiwo sanguinea. These loci were highly variable, with between 2 and 10 alleles per locus, and estimated gene diversity ranging from 0.08 to 0.82. These loci have the potential to reveal genetic structure and estimate gene flow among A. sanguinea populations.     

Allozyme diversity in natural populations of Viola palmensis Webb & Berth. (Violaceae) from La Palma (Canary Islands): implications for conservation genetics

Genetic diversity was measured by allozyme electrophoresis in eight natural populations of the threatened Canarian endemic Viola palmensis Webb & Berth. (Violaceae). Nineteen alleles corresponding to 11 gene loci were detected. High levels of genetic diversity were found, ranging from 36.3 to 45.4 % for the percentage of polymorphic loci (P), from 1.45 to 1.60 for the average number of alleles per locus (A) and from 0.128 to 0.200 for the expected heterozygosity (H(e)). Between 85.5 and 96.6 % of genetic variability was apportioned within populations. As a whole, populations were not at Hardy-Weinberg equilibrium, with a deficit of heterozygous individuals attributable to the existence of genetic structuring in the populations analysed. The levels of interpopulation genetic differentiation were low (mean F(ST) = 0.100), while genetic identity pair-wise comparisons were high (mean I = 0.973) suggesting considerable levels of gene flow among populations. No relationship was detected between genetic differentiation and geographical distances between populations. An outcrossing insect-mediated breeding system might contribute to pollen dispersion of this species. For conservation genetics we suggest in situ preservation areas are defined that are free of disturbance and that include populations with the highest genetic diversity.     

Development of microsatellite markers for the critically endangered Limonium dufourii (Girard) Kuntze (Plumbaginaceae)

Limonium dufourii is an endemic plant from the eastern Mediterranean coast of Spain with a triploid chromosome number and apomictic reproduction. We have isolated and characterized 13 polymorphic microsatellite loci from an enriched library in order to investigate its population genetic structure. Simple sequence repeat (SSR) loci were screened in 120 individuals from the six extant populations of this species. They show an average of 5.76 alleles per locus, ranging from 2 to 18, with seven loci exhibiting heterozygosities larger than 0.60. Three loci present one single allele in each individual, whereas one locus presents three alleles in every individual analysed.     

Patterns of allozyme diversity in the threatened plant Erigeron parishii (Asteraceae)

Thirty-one occurrences of Erigeron parishii, a narrowly endemic plant threatened by mining, were sampled for allozyme diversity. This taxon held considerable genetic variation at the 14 allozyme loci surveyed. Species (e.g., alleles per locus [A] = 4.3 and proportion of polymorphic loci [P] = 0.64) and population (e.g., A = 2.15 [SD = 0.3] and P = 0.53 [SD = 0.12]) genetic diversity measures were higher than expected for narrowly endemic plant taxa. The proportion of polymorphic loci and numbers of alleles per locus indicated that E. parishii has not experienced severe or long-lasting population bottlenecks. Within-population f indicated low to moderate levels of inbreeding. Populations were only moderately differentiated (theta-p = 0.12), suggesting either that there is substantial gene flow among populations or that populations have not been isolated long enough to detect effects of genetic drift. There was no significant differentiation among populations in different vegetation types nor was there a relationship between genetic distance and geographic distance among sites. Continued fragmentation by mining activities would isolate populations, disrupting gene flow, exacerbating loss of diversity, and increasing inbreeding in the remaining fragments. Protection of large, interconnected populations throughout the range of the taxon is warranted to maintain processes that have sustained the observed diversity.     

Isolation and characterization of polymorphic microsatellite loci in the gudgeon,Gobio gobio (Cyprinidae)

A total of seven polymorphic microsatellite loci from Gobio gobio were isolated and characterized. A preliminary population survey of 82 specimens from four populations, located in a downstream pollution gradient of cadmium and zinc, demonstrated the usefulness of these primers both in population genetic studies in general, as well as in evaluating the effects of anthropogenic pollution on genetic structure. Overall locus polymorphism ranged from two to 13 alleles. Observed heterozygosity per locus ranged from 0.39 to 0.73.     

Population genetic structure of wild Phaseolus lunatus (Fabaceae), with special reference to population sizes

To set up an in situ conservation strategy for Phaseolus lunatus, we analyzed the genetic structure of 29 populations in the Central Valley of Costa Rica. Using 22 enzyme loci, we quantified the proportion of polymorphic loci (P(p)), the mean number of alleles per locus (A), and the mean effective number of alleles per locus (A(e)), which equaled to 10.32%, 1.10, and 1.05, respectively. The total heterozygosity (H(T)), the intrapopulation genetic diversity (H(S)), and the interpopulation genetic diversity (D(ST)) were 0.193, 0.082, and 0.111, respectively. The genotypic composition of the analyzed populations showed a deviation from the Hardy-Weinberg proportions (F(IT) = 0.932). This disequilibrium was due to either genetic differentiation between populations (F(ST) = 0.497) or nonrandom mating within populations (F(IS) = 0.866). From the level of genetic differentiation between populations and the private alleles frequencies estimates, gene flow was calculated: Nm(W) = 0.398 and Nm(S) = 0.023, respectively. The results suggested that wild Lima bean maintains most of its isozyme variation among populations. Significant positive correlation was observed between population size and P(p), A, and H(o) (observed heterozygosity), whereas no correlation was observed with the average fixation index of population (F). The loss of genetic variability in populations was attributed to inbreeding and the bottleneck effects that characterized the target populations. In situ conservation and management procedures for wild Lima bean are discussed.     

Genetic structure of endangered Emmenopterys henryi Oliv. based on ISSR polymorphism and implications for its conservation

Inter-simple sequence repeat (ISSR) markers were used to determine the genetic variation and genetic differentiation of nine populations of Emmenopterys henryi Oliv., an endangered plant endemic to China. Relatively low genetic diversity was detected at population level (the percentage of polymorphic loci P=22.56%, the number of alleles per locus A=1.183+/-0.045, the effective number of alleles per locus A(E)=1.007+/-0.345, Nei's gene diversity h=0.071+/-0.017, Shannon information index I=0.104+/-0.025). However, the genetic diversity at species level was relatively high (P=56.05%; A=1.561+/-0.498, A(E)=1.325+/-0.371, h=0.191+/-0.199, I=0.287+/-0.284). Analysis of molecular variance showed that most of the ISSR variation (68.03%) in E. henryi occurred among populations. The estimated Nm from F (ST )was 0.235. It indicated that the fragmentation and isolation of populations might result from specific evolutionary history and anthropogenic activity. Consequently, genetic drift might play an important role in determining the genetic structure of E. henryi. Conservation strategies for this endangered species are proposed based on the genetic data.     

Polymorphic chloroplast microsatellite loci in Nelumbo (Nelumbonaceae)

? Premise of the study: To study population genetics, phylogeography, and hybridization of Nelumbo (Nelumbonaceae), chloroplast microsatellite markers were developed. ? Methods and Results: Seventeen microsatellite loci were identified from the chloroplast genomes of N. nucifera and N. lutea. Polymorphisms were assessed in three populations of N. nucifera and one population of N. lutea. Nine loci were found to be polymorphic in N. nucifera, and all 17 loci were found to be polymorphic in N. lutea. In N. nucifera, the number of alleles per locus ranged from two to six, and the unbiased haploid diversity per locus ranged from 0.198 to 0.790. In N. lutea, the number of alleles ranged from two to four, and the unbiased haploid diversity per locus ranged from 0.245 to 0.694. ? Conclusions: The identified chloroplast simple sequence repeat markers will be useful for the study of genetic diversity, phylogeography, and identification of Nelumbo cultivars.     

Deeply divergent lineages of the widespread New Zealand amphipod Paracalliope fluviatilis revealed using allozyme and mitochondrial DNA analyses

1. We evaluated the population genetic structure of the common New Zealand amphipod Paracalliope fluviatilis using eight allozyme loci, and the mitochondrial cytochrome oxidase c subunit I (COI) gene locus. Morphological analyses were also conducted to evaluate any phenotypic differences. Individuals belonging to P. fluviatilis were collected from a total of 14 freshwater fluvial habitats on the North and South Islands, New Zealand. 2. We found evidence for strong genetic differentiation among locations (Wright's F_ST > 0.25), and fixed differences (non‐shared alleles) at two of the eight allozyme loci indicating the possibility of previously unknown species. Analysis of a 545‐bp fragment of the COI locus was mostly congruent with the allozyme data and revealed the same deeply divergent lineages (sequence divergences up to 26%). 3. Clear genetic breaks were identified between North Island and South Island populations. North Island populations separated by <100 km also showed genetic differences between east and west draining watersheds (sequence divergence >12%). Accordingly, present‐day dispersal among hydrologically isolated habitats appears minimal for this taxon. 4. Although population differences were clearly shown by allozyme and mtDNA analyses, individuals were morphologically indistinguishable. This suggests that, as in North American and European taxa (e.g. Hyalella and Gammarus), morphological conservatism may be prevalent among New Zealand's freshwater amphipods. We conclude that molecular techniques, particularly the COI gene locus, may be powerful tools for resolving species that show no distinctive morphological differences.     

GENETIC VARIABILITY OF GELIDIUM CANARIENSIS (RHODOPHYTA) DETERMINED BY ISOZYME ELECTROPHORESIS1

The populations of Gelidium canariensis (Grunow) Seoane-Camba from the Canary Islands were analyzed for genetic variability by isozyme electrophoresis in 1989 and 1990. Each population was divided into sporophytic and gametophytic subpopulations. Twenty-three to 27 putative alleles corresponding to 22 gene loci were analyzed. Sev-enteen loci were monomorphic in all six subpopulations, and five were polymorphic in at least one subpopulation. Significant deviations from Hardy-Weinberg equilibrium were found. The amount of genetic variability (percentage of polymorphic loci, mean number of alleles per locus, and average gene diversity) of haploid subpopulations was lower than that of diploid subpopulations. No correlation between genetic distance and geographical distance was found. Low genetic differentiation between sporophytic and gametophytic subpopulations of the same locality was obsewed in two populations. The low genetic diversity and genetic differentiation suggest that the genetic structure of the populations of G. canariensis from the Canary Islands is due to a combination of founder effects and the predominance of asexual reproduction. Initial differences in gene frequencies may have persisted because of insufficient time to reach a higher level of differentiation.     

Allozyme differentiation and genetic structure of the Lady's slipper (Cypripedium calceolus) island populations in north-east Poland

The population genetic structure in the rare and endangered perennial plant, Cypripedium calceolus , from north-east Poland was investigated. A total of 11 loci were analysed, of which five were polymorphic for the species and in every population. The 11 loci resolved gave a total of 19 alleles with the mean number of 1.73 alleles per locus. The proportion of polymorphic loci (P) was the same for the species and in every population, 45.5%. The average observed and expected heterozygosities for the total Lady's slipper data set were 0.156 and 0.184, respectively. The average expected heterozygosity for the populations under study ranged from 0.176 to 0.188 and did not differ significantly among populations. Genetic diversity within C. calceolus populations from the Biebrza Valley is relatively high compared with rare taxa and taxa with similar life histories. The genetic diversity among populations of C. calceolus studied ( F _ST = 0.016) is much smaller than the genetic diversity within populations. In the species studied, there was no significant correlation between genetic and geographical distances ( r = −0.73, Mantel test). This may indicate the relatively recent origin of the populations studied, or a high level of gene flow among populations. © 2002 The Linnean Society of London, Botanical Journal of the Linnean Society , 2002, 138 , 433–440.     

Development of microsatellite markers in the toxic dinoflagellate Alexandrium tamarense (Dinophyceae)

Outbreaks of paralytic shellfish poisoning caused by the toxic dinoflagellate Alexandrium tamarense (Dinophyceae) are currently a serious problem from an economic and food hygiene point of view throughout the world. We isolated 13 polymorphic microsatellite loci from this species. These loci provided microsatellite markers with high polymorphism ranging from four to 15 alleles per locus and gene diversity between 0.632 and 0.974. The markers are available for more detailed investigations of genetic structure and gene flow of A. tamarense populations.     

四个不同地理区域的库蚊复合组居群的遗传多样性研究

库蚊种群在农药强选择压下的生存及抗性进化是与其种群的遗传多样性密切相关的。本文主要从居群的遗传多样性角度出发 ,研究其抗性遗传行为。分别对采自 4个不同地理区域的库蚊复合组样群 ,采用水平切片淀粉凝胶电泳的方法 ,分析了 3个酶系统 (苹果酸酶Me、苹果酸脱氢酶Mdh、甘油醛-3-磷酸脱氢酶Gpd) ,获得了 5个位点共 2 7个等位基因的资料。并计算了 4个库蚊居群的遗传多样性指标为 (A ,P ,He,Ho) ,同时分析了 5种居群间的遗传距离。分析结果表明 :遗传多样性主要存在于各居群之内 (FIS=0 .5 4 37) ,但居群之间的遗传分化程度也相对较大 (Gst =0 .4 498)。     

Allozyme frequencies in New Jersey and North Carolina populations of Atlantic white-cedar, Chamaecyparis thyoides (Cupressaceae)

The present study was conducted to assess the distribution of genetic variation within and among populations of Chamaecyparis thyoides in both marginal and centrally located populations. Allozyme frequency analyses of ten loci from foliage of four New Jersey populations and two North Carolina populations of C. thyoides showed polymorphic loci = 50%, mean number of alleles per locus = 2.8, effective number of alleles per locus = 1.17, and expected heterozygosity = 0.14. Diversity was highest in two populations from southern New Jersey. The isolated population at High Point, New Jersey had only two polymorphic loci and expected heterozygosity of 0.03. There was no correlation between genetic and geographic distances among populations, implying that cedar must have possessed some means of long-distance dispersal at the end of the last glacial period, rather than advancing northward step by step.     

Genetic variation of Avicennia marina (Forsk.) Vierh. (Avicenniaceae) in Vietnam revealed by microsatellite and AFLP markers

Genetic variation of Avicennia marina in the costal area of Vietnam was examined using microsatellite and AFLP markers. By using five microsatellite loci a total of 21 alleles were detected. The average number of alleles per locus per population ranged from 1.667 to 3.000. The observed heterozygosity varied from 0.180 to 0.263, with an average of 0.210 indicating relatively low level of genetic variation comparing to the previous studies on A. marina in the worldwide range. The expected heterozygosity was larger than the observed heterozygosity leading to positive inbreeding coefficients in all the six populations. Highly significant departures from Hardy-Weinberg Equilibrium were detected in four populations. AFLP analysis revealed a total of 386 loci, of which 232 (60.1%) were polymorphic. In congruent with microsatellite markers relatively low levels of genetic variation were detected at both gene and nucleotide levels (H = 0.086; pi = 0.0054). Reduced level of genetic variation was found in the central population, and in the southern populations. Both microsatellite and AFLP markers revealed large genetic differentiation (F(ST) = 0.262 and 0.338, respectively) indicating strong genetic structure among regional populations. Pairwise genetic distance by AFLP showed two populations in the north and the other two in the south are closely related each other.     

Selection of high heterozygosity popcorn varieties in Brazil based on SSR markers

We analyzed genetic structure and diversity among eight populations of popcorn, using SSR loci as genetic markers. Our objectives were to select SSR loci that could be used to estimate genetic diversity within popcorn populations, and to analyze the genetic structure of promising populations with high levels of heterozygosity that could be used in breeding programs. Fifty-seven alleles (3.7 alleles per locus) were detected; the highest effective number of alleles (4.21) and the highest gene diversity (0.763) were found for the Umc2226 locus. A very high level of population differentiation was found (F(ST) = 0.3664), with F(ST) for each locus ranging from 0.1029 (Umc1664) to 0.6010 (Umc2350). This analysis allowed us to identify SSR loci with high levels of heterozygosity and heterozygous varieties, which could be selected for production of inbred lines and for developing new cultivars.     

意大利蜜蜂产浆性状10个微卫星位点的遗传分析

我国培育成功的世界上蜂王浆产量最高的蜂种（Ea）是从20世纪30年代引进我国的意大利蜜蜂（Eb）中选育的。采用10个微卫星位点对蜂王浆高产蜜蜂、原种意大利蜜蜂（Ee）和本地意大利蜜蜂进行研究,以探明由于人工选择和地理隔离造成的其分子进化上的一些特征。结果,10个微卫星位点在3个蜂种中共扩增到96个等位基因,其中有48个等位基因是不同的,表明10个微卫星位点在3个蜂种中的高度多态性,而且由于人工选择和地理隔离已造成3蜂种在遗传基础上的一些分化。3蜂种的多态信息含量（PIC）分别为0．57、0．50、0．57,杂合度分别为0．60、0．57、0．61,二者均无显著差异。遗传距离的分析结果显示,Ee和Eb（0．14）、Eb和Ea（0．16）之间的遗传距离较近,而Ea和Ee（0.25）之间的遗传距离较远。等位基因频率的分析结果表明,6个位点的7个等位基因的频率（A29的159bp、A24的100bp和104bp、A7的110bp、A43的126bp、A14的221bp和A113的221bp）以Ee、Eb、Ea的顺序递增,Ea的这7个等位基因频率分别显著高于Ee和Eb。同时4个位点的4个等位基因的频率（A24的106bp、A43的140bp;A113的215bp和A14的219bp）以Ea、Eb、Ee的Ⅲ页序下降,Ea在这4个位点的频率分别显著低于Eb和Ee。这些位点的等位基因可能与蜂王浆产量有关。     

Genetic variation in remnant populations of Dalbergia nigra (Papilionoideae), an endangered tree from the Brazilian Atlantic Forest

BACKGROUND AND AIMS: Dalbergia nigra, known as Brazilian rosewood, is an endangered tree species restricted to the Brazilian Atlantic Forest and has been intensively logged for five centuries due to its high-quality wood. The objective of the present study was to assess the genetic variation and structure in adults and saplings of the species from a large reserve of the Atlantic Forest, the Rio Doce State Park, and from two small surrounding fragments, one better preserved and another with a high degree of anthropogenic disturbance. METHODS: Analyses of genetic variation and structure were conducted by studying allozyme markers. Seven putative enzymatic loci were resolved, five of them being polymorphic. KEY RESULTS: The mean numbers of alleles per locus (A) were 1.93 and 1.73, while the percentages of polymorphic loci (P) were 93 and 73 % for adults and saplings, respectively. Saplings from the fragment with high anthropogenic disturbance exhibited the lowest values of A and P. The fragment that constitutes a conservation area exhibited genetic variation similar to the population from the large reserve. The observed (H(o)) and expected (H(e)) heterozygosities were not significantly different among the three populations. Only sapling populations showed F(ST) values (divergence among populations) significantly different from zero over all studied loci. The fragment with high anthropogenic disturbance exhibited considerable genetic divergence in relation to the above-cited populations. CONCLUSIONS: The evaluated populations displayed mean levels of genetic variation intermediate to those expected for narrow and widespread species. The results suggest that fragments with similar area and geographical distance from a large protected reserve can exhibit different levels of genetic variation, depending on the degree of anthropogenic disturbance. The considerable genetic variation in the protected fragment points to the importance of adequate conservation of small fragments for the preservation of genetic variation in D. nigra.