Genetic variability in Brazilian wheat cultivars assessed by microsatellite markers

Wheat (Triticum aestivum) is one of the most important food staples in the south of Brazil. Understanding genetic variability among the assortment of Brazilian wheat is important for breeding. The aim of this work was to molecularly characterize the thirty-six wheat cultivars recommended for various regions of Brazil, and to assess mutual genetic distances, through the use of microsatellite markers. Twenty three polymorphic microsatellite markers (PMM) delineated all 36 of the samples, revealing a total of 74 simple sequence repeat (SSR) alleles, i.e. an average of 3.2 alleles per locus. Polymorphic information content (PIC value) calculated to assess the informativeness of each marker ranged from 0.20 to 0.79, with a mean of 0.49. Genetic distances among the 36 cultivars ranged from 0.10 (between cultivars Ocepar 18 and BRS 207) to 0.88 (between cultivars CD 101 and Fudancep 46), the mean distance being 0.48. Twelve groups were obtained by using the unweighted pair-group method with arithmetic means analysis (UPGMA), and thirteen through the Tocher method. Both methods produced similar clusters, with one to thirteen cultivars per group. The results indicate that these tools may be used to protect intellectual property and for breeding and selection programs.     

Genetic relationships and population structure of local olive tree accessions from Northeastern Spain revealed by SSR markers

The olive (Olea europaea L. subsp. europaea) is an old traditional crop which was domesticated from the wild at various locations around the Mediterranean basin, resulting in a vast number of accessions worldwide. This is the case of the northeast part of Spain (Aragon), where 163 genotypes were prospected and characterised. The majority of these genotypes (90 %) seem to be autochthonous accessions of this area and the rest are synonyms of accessions from other areas of Spain. In this study, 11 nuclear SSR markers were used to discriminate between the accessions and to understand the genetic relationship among them. All primers produced a successful amplification giving a total of 176 fragments in the genotypes studied, with an average of 16 alleles per SSR, ranging from 5 (DCA13) to 24 (DCA11). Allele size ranged from 120 bp at locus UDO99 to 284 bp at locus DCA15. The dendrogram generated using the variability observed classified most of the genotypes according to their geographical origin (Huesca, Teruel and Zaragoza provinces), confirming the particular evolution of different olive ecotypes. Structure software was used to investigate the genetic population structure among olive accessions, showing that the maximum rate of change in the log probability of the data occurred at K = 2. In addition, the SSR markers have consequently shown their usefulness for cultivar identification in olive, for establishing the genetic closeness among its accessions, and for establishing genealogical relationships.     

Genetic diversity and population structure of Taxus cuspidata in the Changbai Mountains assessed by chloroplast DNA sequences and microsatellite markers

Taxus cuspidata, a tree species with high economic value because of its anticancer properties, is experiencing severe reduction in populations across its range in China. We examined one chloroplast DNA (cpDNA) region (petA-psbE) and 9 nuclear simple sequence repeats (SSRs) loci variations among seven populations in the Changbai Mountains of China to investigate the levels of genetic diversity and population structure. A moderate level of haplotype diversity (H_T = 0.625), low nuclear microsatellite diversity (H_E = 0.261 ± 0.028), significant genetic differences (F_ST = 0.065) and substantial gene flow (Nm = 2.806) were observed. Most of the total genetic variation was partitioned within the population (87.8% and 94.0% for cpDNA and SSRs, respectively). Our haplotype identification permutation tests revealed that G_ST > N_ST, indicating an absence of phylogeographic structure in T. cuspidata. Neither STRUCTURE nor UPGMA analyses showed any geographic pattern in T. cuspidata populations. By comparatively analyzing the genetic diversity and survival situation of T. cuspidata, our results provide a theoretical foundation for the resource protection, utilization cultivation and breeding of this valuable plant.     

Genetic diversity in the Pantaneiro horse breed assessed using microsatellite DNA markers

The genetic variability for a sample of 227 animals from three populations of Pantaneiro horses was estimated using data from 10 microsatellite loci. The number of alleles and the proportion of heterozygosity indicated high variability. A total of 91 alleles were found, with a significantly high mean number of alleles. The mean polymorphic information content was 0.7 and the paternity exclusion probability was 99.3%. The inbreeding coefficient (F(IS)) was low for the three populations: Ipiranga (F(IS) = 0.147), Nova Esperan?a (F(IS) = 0.094) and Promiss?o (F(IS) = 0.108). Genetic differentiation among all three populations was low (F(ST) = 0.008 to 0.064). Three methods were used to test for a recent bottleneck effect. The graphical method and the Wilcoxon test using the stepwise mutation model showed no bottleneck pattern for any of the populations. The test by two-phase mutation model showed genetic signatures of bottleneck for Ipiranga and Promiss?o. When we consider standard deviation value for Nova Esperan?a, the M-statistic detected a bottleneck pattern, but this result could be explained by a sample size effect. Therefore, there is no immediate cause for concern regarding loss of variation within the breed.     

Genetic diversity among cultivars, landraces and wild relatives of rice as revealed by microsatellite markers

Genetic diversity among 35 rice accessions, which included 19 landraces, 9 cultivars and 7 wild relatives, was investigated by using microsatellite (SSR) markers distributed across the rice genome. The mean number of alleles per locus was 4.86, showing 95.2% polymorphism and an average polymorphism information content of 0.707. Cluster analysis based on microsatellite allelic diversity clearly demarcated the landraces, cultivars and wild relatives into different groups. The allelic richness computed for the clusters indicated that genetic diversity was the highest among wild relatives (0.436), followed by landraces (0.356), and the lowest for cultivars. Allelic variability among the SSR markers was high enough to categorize cultivars, landraces and wild relatives of the rice germplasm, and to catalogue the genetic variability observed for future use. The results also suggested the necessity to introgress genes from landraces and wild relatives into cultivars, for cultivar improvement.     

Genetic diversity and population structure of wild olives from the North-Western Mediterranean assessed by SSR markers

BACKGROUND AND AIMS: This study examines the pattern of genetic variability and genetic relationships of wild olive (Olea europaea subsp. europaea var. sylvestris) populations in the north-western Mediterranean. Recent bottleneck events are also assessed and an investigation is made of the underlying population structure of the wild olive populations. METHODS: The genetic variation within and between 11 wild olive populations (171 individuals) was analysed with eight microsatellite markers. Conventional and Bayesian-based analyses were applied to infer genetic structure and define the number of gene pools in wild olive populations. KEY RESULTS: Bayesian model-based clustering identified four gene pools, which was in overall concordance with the Factorial Correspondence Analysis and Fitch-Margoliash tree. Two gene pools were predominantly found in southern Spain and Italian islands, respectively, in samples gathered from undisturbed forests of the typical Mediterranean climate. The other two gene pools were mostly detected in the north-eastern regions of Spain and in continental Italy and belong to the transition region between the temperate and Mediterranean climate zones. CONCLUSIONS: On the basis of these results, it can be assumed that the population structure of wild olives from the north-western Mediterranean partially reflects the evolutionary history of these populations, although hybridization between true oleasters and cultivated varieties in areas of close contact between the two forms must be assumed as well. The study indicates a degree of admixture in all the populations, and suggests some caution regarding genetic differentiation at the population level, making it difficult to identify clear-cut genetic boundaries between candidate areas containing either genuinely wild or feral germplasm.     

Genetic diversity and admixture analysis of Sanfratellano and three other Italian horse breeds assessed by microsatellite markers

Sanfratellano is a native Sicilian horse breed, mainly reared in the north east of the Island, developed in the 19th century from local dams and sires with a restricted introgression of Oriental, African and, more recently, Maremmano stallions. In this study, the genetic relationships and admixture among Sanfratellano, the other two Sicilian autochthonous breeds and Maremmano breed were assessed using a set of microsatellites. The main goals were to infer the impact of Maremmano breed in the current Sanfratellano horse and to provide genetic information useful to improve the selection strategies of the Sanfratellano horse. The whole sample included 384 horses (238 Sanfratellano, 50 Sicilian Oriental Purebred, 30 Sicilian Indigenous and 66 Maremmano), chosen avoiding closely related animals. A total of 111 alleles from 11 microsatellite loci were detected, from four at HTG7 to 15 at ASB2 locus. The mean number of alleles was the lowest in Oriental Purebred (6.7), the highest in Sanfratellano (8.3). All the breeds showed a high level of gene diversity (He) ranging from 0.71 ± 0.04 in Sicilian Oriental Purebred to 0.81 ± 0.02 in Sicilian Indigenous. The genetic differentiation index was low; only about 6% of the diversity was found among breeds. Nei's standards (DS) and Reynolds' (DR) genetic distances reproduced the same population ranking. Individual genetic distances and admixture analysis revealed that: (a) nowadays Maremmano breed does not significantly influence the current Sanfratellano breed; (b) within Sanfratellano breed, it is possible to distinguish two well-defined groups with different proportions of Indigenous blood.     

Genetic structure of seven European cattle breeds assessed using 20 microsatellite markers

Genotype data from 20 microsatellites typed in 253 animals is used here to assess the genetic structure of seven European pedigree cattle breeds. Estimation of genetic subdivision using classical drift-based measures shows that the average proportion of genetic variation among breeds varies between 10 and 11% of the total, depending on the estimator used. We demonstrate that a simple allele-sharing genetic distance parameter can be used to construct a dendrogram of relationships among animals. This phylogenetic tree displays a remarkable degree of breed clustering and reflects an extensive underlying kinship structure, particularly for the Swiss Simmental breed and four breeds originating from the British Isles. Condensation of allele frequencies and individual genotypic compositions using principal component analysis is also used to investigate genetic structure among breeds and individual animals. In addition, the underlying genetic demarcation of European cattle breeds is emphasized in simulations of breed assignment using allele frequency distributions from samples of microsatellite loci. Correct breed designation can be inferred with accuracies approaching 100% using data from a panel of 10 microsatellite loci.     

Genetic diversity among barley cultivars assessed by sequence-specific amplification polymorphism

We analyzed the genetic structure and relationships among barley cultivars (Hordeum vulgare L.) with sequence-specific amplification polymorphisms (S-SAPs). Polymorphisms were identified in 824 individual barley plants representing 103 cultivars (eight plants per cultivar) widely grown in Canada and the United States, using PCR primers designed from the long terminal repeat of the barley retrotransposon BARE-1 and a subset of four selective MseI primers. From the 404 bands scored, 150 were polymorphic either within or between cultivars. Genetic structure assessed with analysis of molecular variance attributed the largest component of variation to the within groups of cultivars (69–86%). Within-cultivar genetic variation was estimated as average gene diversity over loci and ranged from 0 (completely homogenous) to 0.076 (most heterogeneous cultivar). Only 17 out of 103 cultivars (16%) were judged to be homogenous by this criterion. Relationships among cultivars were analyzed by cluster analysis using unweighted pair-groups using arithmetic averages and found groups similar to those determined by agriculturally significant phenotypic traits such as spike morphology (two-rowed or six-rowed), cultivar type (malting or feed), seed characteristic (hull-less or hulled), and growth habit (winter or spring), with minor overlaps. Discriminant analysis of groups determined by these phenotypic traits fully supported the different groups with minor overlaps between the malting/feed. S-SAP markers generated from retrotransposons such as BARE-1 are invaluable tools for the study of genetic diversity in organisms with a narrow genetic base such as barley. In this study, S-SAP analysis revealed significant amounts of cryptic variation in closely related cultivars including somaclonal variation, which could not be inferred by the pedigree analysis.     

Genetic diversity and population structure of Sicilian sheep breeds using microsatellite markers

Genetic diversity studies in domestic animals aim at evaluating genetic variation within and across breeds mainly for conservation purposes. In Sicily, dairy sheep production represents an important resource for hilly and mountain areas economy. Their milk is used for the production of traditional raw milk cheeses, sometimes protected designation of origin (PDO) cheeses. In some cases, the quality of these products is linked to a specific breed, i.e. mono-breed labelled cheeses and it is therefore important to be able to distinguish the milk of a breed from that of others, in order to guarantee both the consumer and the breed itself. In order to investigate the genetic structure and to perform an assignment test, a total of 331 individuals (Barbaresca, BAR n = 57, Comisana, COM n = 65, Pinzirita, PIN n = 75, Sarda, SAR n = 64, and Valle del Belice, VDB n = 70) were analysed using a panel of 20 microsatellite markers. A total of 259 alleles were observed with average polymorphic information content equal to 0.76, showing that the microsatellites panel used was highly informative. Estimates of observed heterozygosity ranged from 0.65 in the BAR breed to 0.75 in the COM breed. The low value of genetic differentiation among breeds (F_st = 0.049) may indicate that these breeds are little differentiated probably due to common history and breeding practices. The low F_is and F_it values indicated low level of inbreeding within and among breeds. The unrooted neighbor-joining dendrogram obtained from the Reynold's genetic distances, and factorial correspondence analysis revealed a separation between BAR and the other sheep breeds. Recent migration rates were estimated, showing that four out of the five breeds have not received a significant proportion of migrants. Only for the PIN breed a recent introgression rate from the VDB breed (7.2%) was observed. The Bayesian assignment test showed that BAR and SAR breeds had a more definite genetic structure (proportion of assignment of 92% and 86.6%, respectively), whereas the lowest assignment value was found in the PIN breed (67.1%). Our results indicated high genetic variability, low inbreeding and low genetic differentiation, except for BAR breed, and were in accordance with geographical location, history, and breeding practices. The low robustness of the assignment test makes it unfeasible for traceability purposes, due to the high level of admixture, in particular for COM, PIN and VDB.     

Genetic diversity and population structure of Chinese pony breeds using microsatellite markers

China is one of the principal origins of ponies in the world. We made a comprehensive analysis of genetic diversity and population structure of Chinese ponies based on 174 animals of five indigenous Chinese pony breeds from five provinces using 13 microsatellite markers. One hundred and forty-four alleles were detected; the mean number of effective alleles among the pony breeds ranged from 5.38 (Guizhou) to 6.78 (Sichuan); the expected heterozygosity ranged from 0.82 (Guizhou) to 0.85 (Debao, Sichuan). Although abundant genetic variation was found, the genetic differentiation was low between the ponies, with 6% total genetic variance among the different breeds. All the pairwise F(ST) values were significant; they varied from 0.0424 for the Sichuan-Yunnan pair to 0.0833 for the Guizhou-Sichuan pair. All five pony breeds deviated from Hardy-Weinberg equilibrium, except the Yunnan pony. Phylogenetic trees of the five pony breeds based on genetic distances were constructed using a neighbor-joining method. The Sichuan and Yunnan ponies were grouped into the same branch, with a high bootstrap support value (97%). Guizhou and Ningqiang ponies were clustered into the same branch with a bootstrap value of 56%, whereas the Debao pony was placed in a separate group, with a bootstrap value of 56%. This grouping pattern was supported by genetic structure analysis.     

Genetic diversity and population structure of 10 Chinese indigenous egg-type duck breeds assessed by microsatellite polymorphism

The genetic structure and diversity of 10 Chinese indigenous egg-type duck breeds were investigated using 29 microsatellite markers. The total number of animals examined were 569, on average 57 animals per breed were selected. The microsatellite marker set analysed provided 177 alleles (mean 6.1 alleles per locus, ranging from 3 to 10). All populations showed high levels of heterozygosity with the lowest estimate of 0.539 for the Jinding ducks, and the highest 0.609 observed for Jingjiang partridge ducks. The global heterozygote deficit across all populations (F _IT) amounted to −0.363. About 10% of the total genetic variability originated from differences among breeds, with all loci contributing significantly. An unrooted consensus tree was constructed using the NeighborNet tree based on the Reynold’s genetic distance. The structure software was used to assess genetic clustering of these egg-type duck breeds. Clustering analysis provided an accurate representation of the current genetic relations among the breeds. An integrated analysis was undertaken to obtain information on the population dynamics in Chinese indigenous egg-type duck breeds, and to better determine the conservation priorities.     

Genetic maternity and paternity in a local population of armadillos assessed by microsatellite DNA markers and field data

Genetic data from polymorphic microsatellite loci were employed to estimate paternity and maternity in a local population of nine-banded armadillos (Dasypus novemcinctus) in northern Florida. The parentage assessments took advantage of maximum likelihood procedures developed expressly for situations when individuals of neither gender can be excluded a priori as candidate parents. The molecular data for 290 individuals, interpreted alone and in conjunction with detailed biological and spatial information for the population, demonstrate high exclusion probabilities and reasonably strong likelihoods of genetic parentage assignment in many cases; low mean probabilities of successful reproductive contribution to the local population by individual armadillo adults in a given year; and statistically significant microspatial associations of parents and their offspring. Results suggest that molecular assays of highly polymorphic genetic systems can add considerable power to assessments of biological parentage in natural populations even when neither parent is otherwise known.     

Genetic structure and diversity of Portunus trituberculatus in Chinese population revealed by microsatellite markers

By using six polymorphic microsatellite loci, the population genetic diversity and structure of 120 individuals of Portunus trituberculatus were examined in five wild populations representing different geographical and ecological ranges along the coast of China. Middle level of genetic diversity of P. trituberculatus was revealed by observed heterozygosity, allele number and Shannon information index. Pairwise population differentiations were found between all pairs of populations except Beihai (BH) and Qingdao (QD) (P < 0.05). The greatest differentiation was detected between Yingkou (YK) and QD populations. It was inconsistent with the result in previous study using mitochondrial control region, which showed no differentiation between QD and YK population. Recent bottleneck was identified in Ningbo (NB) population under TPM and IAM models, as well as Dandong (DD) and QD populations under IAM model. Additionally, compared with YK, QD, DD and BH populations, lower historical effective population size with lower genetic diversity were detected in NB population, which called for urgent assessment of the ecological and economic potential of NB fisheries.     

Genetic relatedness of Portuguese almond cultivars assessed by RAPD and ISSR markers

Randomly amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were used to analyse the genetic diversity of Portuguese Prunus dulcis cultivars and their relationship to important foreign cultivars. Of the primers tested, 6 (out of 60) RAPD and 5 (out of 18) ISSR primers were selected for their reproducibility and high polymorphism. Out of 124 polymerase chain reaction fragments that were scored, 120 (96.8%) were polymorphic. All the plants could be discriminated and constitute a very heterogeneous group. Five unidentified almond plants found in the region of Foz Côa (north Portugal) and wild almond (P. webbii) from Italy and Spain were also included. Four main groups of plants could be distinguished: P. dulcis cultivars; one Foz Côa plant; P. webbii; and P. persica (outgroup). The segregating Foz Côa plant may represent a feral individual or a hybrid between P. dulcis and P. webbii.Abbreviations dNTP Deoxynucleotide triphosphate - CTAB Cetyltrimethylammonium bromide - ISSR Inter-simple sequence repeats - PCR Polymerase chain reaction - RAPD Randomly amplified polymorphic DNA - RASTM Regional Agricultural Services of Trás-os-Montes - TE Tris-EDTA buffer - UPGMA Unweighted pair group method with arithmetical averagesCommunicated by P. Puigdoménech     

Genetic diversity and structure of natural fragmented Chamaecyparis obtusa populations as revealed by microsatellite markers

The genetic diversity and population structure of hinoki (Chamaecyparis obtusa) in Japan were investigated by examining the distribution of alleles at 13 microsatellite loci in 25 natural populations from Iwaki in northern Japan to Yakushima Island in southern Japan. On average, 26.9 alleles per locus were identified across all populations and 4.0% of the genetic variation was retained among populations (G _ST = 0.040). According to linkage disequilibrium analysis, estimates of effective population size and detected evidence of bottleneck events, the genetic diversity of some populations may have declined as a result of fragmentation and/or over-exploitation. The central populations located in the Chubu district appear to have relatively large effective population sizes, while marginal populations, such as the Yakushima, Kobayashi and Iwaki populations, have smaller effective population sizes and are isolated from the other populations. Microsatellite analysis revealed the genetic uniqueness of the Yakushima population. Although genetic differentiation between populations was low, we detected a gradual cline in the genetic structure and found that locus Cos2619 may be non-neutral with respect to natural selection.     

Genetic diversity and differentiation in nine West African local goat breeds assessed via microsatellite polymorphism

This study was carried out to study the genetic relationship among nine (9) West African goat breeds out of which 6 (Guinea dwarf, Kirdi, Ghana dwarf, Mossi, Togo dwarf, Senegal dwarf) were of the trypanotolerant type and 3 (Mali Sahel, Mauritania Sahel, Chad Sahel) were of the trypanosusceptible type. One hundred ninety nine (199) animals were genotyped using 10 microsatellites. The microsatellites loci analysed were highly polymorphic with a mean number of alleles of 11.7 ± 3.7 ranging from 6 to 20. Observed heterozygosities ranged from 0.603 ± 0.164 (Togo dwarf) to 0.726 ± 0.166 (Chad Sahel). It was lower than expected heterozygosity in most of the breeds indicating a deviation from Hardy Weinberg equilibrium. The Fst measure of genetic distance between pairs of populations showed the highest distance (0.098) between Kirdi and Guinea dwarf which presented an isolated position from the other breeds and the lowest distance (0.025) between Togo dwarf and Mossi. Our study showed that the genetic relationships among the concerned breeds correspond to their geographical distribution. In addition, Guinea dwarf is strongly separated from the other breeds.     

Population structure and genetic diversity of Lithocarpus litseifolius (Fagaceae) assessed using microsatellite markers

Lithocarpus litseifolius (Fagaceae), known as tree of Chinese sweet tea, is a native commercial plant distributed in south China and adjacent areas. This study used ten microsatellite markers to investigate the genetic variation and population structure of 11 populations of L. litseifolius from the main production regions in China. All of the tested loci proved to be effective for the species. The number of alleles per locus ranged between 3 and 39, and mean observed and expected heterozygosity was 0.43 and 0.52, respectively. The deficiency in heterozygosity may be the result of human interference and harvesting activities. Analysis of molecular variance revealed that most of the variation was within populations, whereas differentiation among populations was insignificant, and that there was no significant correlation between genetic distance and geographic distance (r = 0.4089, p = 0.9940). However, A Bayesian model‐based analysis sorted all 11 populations into two (when k = 2) or three (when k = 3) clusters with higher probabilities and showed a geographical pattern which is probably related to the topography of China. Our results provide important information for protecting, sustainably using, and improving the current resources of L. litseifolius.     

Genetic relationships between two homologous goat breeds from Portugal and Brazil assessed by microsatellite markers

Twenty microsatellite markers were used to investigate genetic diversity and relationships in the Portuguese Serpentina (SERP) and the Brazilian Moxotó (MOX) goat breeds, which present a striking resemblance and are believed to have a common origin. A total of 46 animals of the SERP breed were sampled in southern Portugal, and 233 MOX goats were sampled in five different regions of Northeast Brazil. Genetic diversity was high, with a mean number of alleles of nearly 8.7 in both breeds, but expected heterozygosity and allelic richness were higher in SERP. The genetic distance between the two breeds resulted in a global F_ST of 0.16, and a strong deficit in within-breed heterozygosity was observed in MOX, mostly because of population substructuring. The estimated inbreeding was about 0.05 in SERP and ranged from 0.02 to 0.08 in the different MOX subpopulations. These subpopulations showed considerable genetic differentiation from each other, and those sampled in Paraíba and Rio Grande do Norte are the ones with a closer relationship with SERP. The analysis with STRUCTURE confirmed that some MOX subpopulations may share a common, but distant, ancestry with SERP, both having contributions from the same potential ancestral population. Other MOX subpopulations, however, are identified with ancestral populations distinct from SERP. These analyses further confirmed that, with the exception of goats from Rio Grande do Norte, the other MOX subpopulations show very little evidence of admixture among them, which may reflect the typical goat raising system in Northeast Brazil, where herds are usually kept closed and isolated, with little opportunity for gene flow among subpopulations. Our results indicate that SERP is likely a distant ancestor of MOX, but founder effects, genetic drift, selection for different environmental constraints and the possible influence of other breeds in the more recent past, probably lead to a differentiation of MOX subpopulations from their distant ancestor, as well as from each other.