Molecular characterization of olive (Olea europaea L.) Sicilian cultivars using SSR markers

Olive (Olea europaea L.) is one of the economically most important fruit crops for the Mediterranean area, with production being mainly destined to oil extraction. In Sicily, olive has been cultivated since ancient times and its germplasm is characterized by a wide genetic diversity that could be related to its domestication and spread in ancient times, and to some reproductive biological peculiarities as self-incompatibility. This analysis was conducted on 65 genotypes with the purpose of characterizing a large collection of Sicilian accessions (47 genotypes) and to compare them with varieties coming from Southern Italy and from the most important countries of the Mediterranean basin. With this aim we used 8 simple sequence repeat (SSR) markers, which detected a total of 74 alleles and identified an average of 19.5 genotypes in the population investigated. A larger variability than expected was found in the analyzed genotypes, some synonymies already reported in literature were confirmed, but also some cultivars considered as identical were discriminated such as in the case of Castriciana, Ogliarola messinese and Passalunara. The whole study revealed a wide intraspecific variability within the gene pool examined, independently from the geographical origin.     

Genetic characterisation of traditional chestnut varieties in Italy using microsatellites (simple sequence repeats) markers

European chestnut (Castanea sativa) is an important multipurpose tree that has been cultivated for wood and fruit in the Mediterranean basin since ancient times. Cultivation of traditional chestnut varieties has a long tradition in Italy, where cultivars have been selected over centuries as a function of the best nut traits. In this study, 94 grafted chestnuts corresponding to 26 representative cultivars from Italy were evaluated by seven simple sequence repeat (SSR) markers to establish whether they corresponded to varieties in the narrow sense. The results allowed 20 genotypes to be identified that corresponded to the same number of clones. In total, 52 alleles were identified, eight of which were exclusive. Cases of homonymies and synonymies were detected. Moreover, our results highlighted a considerable genetic uniformity among ‘Marrone-type’ cultivars and, on the contrary, a high genetic diversity among the evaluated cultivars demonstrating that this is a valuable germplasm and an important genetic resource to be preserved.     

Genetic and landscape characterization of ancient autochthonous olive trees in northern Italy

Several ancient olive plants with different genetic, agronomic and morphological characteristics are widespread in the Emilia-Romagna region. In this study, details of 206 trees located in four provinces were collected and their phytometric traits were recorded. Easily recognizable varieties were identified, while plants of uncertain genetic identity were submitted for molecular analysis with SSR markers and compared with cultivars widespread in central Italy and Emilia-Romagna. The genetic analyses showed that most of the ancient olive trees belonged to 10 cultivars that are already characterized, with different cases of intra-varietal clones. The remaining accessions belonged to 19 different genotypes of unknown origin. The highest percentage of policormic trees was found in the Bologna and Rimini provinces. A high percentage of plants located under unfavourable aspects (west to east) was found only in the Rimini territory, which presents peculiar and milder climatic conditions compared to the other provinces. Slope appeared not to be a limiting factor for tree survival. The information obtained could help the rational reintroduction of olive growing in suitable areas of Emilia-Romagna, with the subsequent economic advantages, erosion preservation and germplasm enrichment.     

Centennial olive trees as a reservoir of genetic diversity

Background and Aims

Genetic characterization and phylogenetic analysis of the oldest trees could be a powerful tool both for germplasm collection and for understanding the earliest origins of clonally propagated fruit crops. The olive tree (Olea europaea L.) is a suitable model to study the origin of cultivars due to its long lifespan, resulting in the existence of both centennial and millennial trees across the Mediterranean Basin.

Methods

The genetic identity and diversity as well as the phylogenetic relationships among the oldest wild and cultivated olives of southern Spain were evaluated by analysing simple sequence repeat markers. Samples from both the canopy and the roots of each tree were analysed to distinguish which trees were self-rooted and which were grafted. The ancient olives were also put into chronological order to infer the antiquity of traditional olive cultivars.

Key Results

Only 9·6 % out of 104 a priori cultivated ancient genotypes matched current olive cultivars. The percentage of unidentified genotypes was higher among the oldest olives, which could be because they belong to ancient unknown cultivars or because of possible intra-cultivar variability. Comparing the observed patterns of genetic variation made it possible to distinguish which trees were grafted onto putative wild olives.

Conclusions

This study of ancient olives has been fruitful both for germplasm collection and for enlarging our knowledge about olive domestication. The findings suggest that grafting pre-existing wild olives with olive cultivars was linked to the beginnings of olive growing. Additionally, the low number of genotypes identified in current cultivars points out that the ancient olives from southern Spain constitute a priceless reservoir of genetic diversity.     

Assessment of genetic diversity and DNA profiling of linseed (<Emphasis Type="Italic">Linum usitatissimum</Emphasis> subsp. <Emphasis Type="Italic">usitatissimum</Emphasis> L.) germplasm using SSR markers

Access to genetic diversity is essential for any progress in adapting linseed (Linum usitatissimum subsp. usitatissimum L.) cultivation to changing environmental conditions or to the changing market needs. An attempt has been made in the present study to assess genetic diversity in 96 genotypes of linseed including varieties, landraces and exotic material. A total of 38 SSR primers amplified 153 alleles with 4.0 alleles per marker locus. The number of alleles ranged from 2 to 15 and the observed polymorphism ranged from 50 to 100%. Average genetic dissimilarity ranged from 2 to 50%. In order to analyze the efficiency for unambiguous identification of linseed germplasm, various statistical measures, viz., number of genotyping patterns, polymorphism information content, resolving power, discrimination power, probability of identity and probability of random identity, identified a set comprising of primers LU7, LU27, LU25, LU20 and LU31 (or LU637) for DNA fingerprinting of linseed germplasm. UPGMA cluster analysis showed that all genotypes could be grouped into four main clusters. Cluster 2 was the largest consisting of mainly landraces, whereas, Cluster 4 was the smallest. Cluster 1 consisted of mainly the released cultivars. Cluster 3 and Cluster 4 were smaller clusters and consisted of exotic genotypes. Principal co-ordinate analysis further substantiated the UPGMA clustering patterns of the observed genetic relationship. To explain 70–80% variability, 17–23 PCOs were needed, whereas 70 components were needed to explain the whole variability in the linseed material under study. Analysis of molecular variance indicated that most of the genetic variation is owing to the individuals within single population, whereas grouping of linseed material into varieties, landraces and exotics accounted for nearly 10% of the total genetic variation. The utility of SSR markers in diversity assessment and cultivar identification is discussed.     

Assessing Genetic Diversity in <Emphasis Type="Italic">Olea europaea</Emphasis> L. Using ISSR and SSR Markers

Olea europaea L. is one of the most economically important crops in the Mediterranean area, and known for having large genetic variability. In order to assess the genetic diversity, DNA from 41 olive cultivars, present in the protected denomination of origin (PDO) region of Trás-os-Montes, was screened using inter simple sequence repeat (ISSR) and microsatellite (SSR) markers. Eleven ISSR primers amplified 135 reproducible bands of which 108 were polymorphic. The percentage of polymorphic bands detected by ISSR was 79%. The highest number of polymorphic bands was obtained by the use of primers UBC807 (15) and UBC809 (16). A total of 67 alleles were detected by six SSR primers, with an average of 11 alleles per primer. The number of alleles per locus ranged from five (ssrOeUaDCA05) to 15 (ssrOeUaDCA03). The observed heterozygosity ranged from 0.219 (ssrOeUaDCA05) to 0.900 (ssrOeUaDCA04), while the expected heterozygosity varied between 0.426 (ssrOeUaDCA05) and 0.887 (ssrOeUaDCA03). The polymorphism information content (PIC) values ranged from 0.392 (ssrOeUaDCA05) to 0.863 (ssrOeUaDCA03). The collection of primers selected gave a reasonable number of amplification products for the genetic diversity analysis. Based on the results, the genetic diversity among 41 olive cultivars is discussed. This study reveals the great importance of guaranteeing the differentiation of olive cultivars and their application for certification purposes.     

AFLP reveals structural details of genetic diversity within cultivated olive germplasm from the Eastern Mediterranean

Amplified fragment length polymorphism (AFLP) analysis was used to assess genetic inter-relationships among olive varieties cultivated in the Eastern Mediterranean Basin. The genotypes sampled included most of the important cultivars from Turkey, Greece and the Middle East and selected genotypes from the Western Mediterranean area. A total of 119 polymorphic markers were generated from five selective primer-pair combinations. The combined data sets generated by just two primer-pairs were adequate to discriminate between all 65 genotypes, while each primer-pair could individually identify up to 64 genotypes. A factorial correspondence analysis (FCA) plot indicated that the cultivars clustered into two relatively modestly defined groups. The first broad group was dominated by cultivars from Turkey but also included genotypes originating from the Middle East (Syria and Lebanon) that collectively formed a tight subcluster. The second group comprised Greek cultivars and those originating from the Western Mediterranean. A significant genetic distance value between Greek and Turkish cultivars was provided by an analysis of molecular variance (amova). There was also evidence of substructure here, with an apparent separation of most Spanish and Italian clones. These findings are in general accordance to previous suggestions of an East-West divergence of olive cultivars, although the dichotomy is less extensive than reported previously and complicated by regional variation within each group.     

Genetic diversity and relationships in olive ( Olea europaea L.) germplasm collections as determined by randomly amplified polymorphic DNA

Genetic diversity studies using the RAPD technique were carried out in a set of 103 olive cultivars from the World Germplasm Bank of the Centro de Investigación y Formación Agraria (CIFA) "Alameda del Obispo" in Cordoba (Spain). A total of 126 polymorphisms (6.0 polymorphic markers per primer) out of 135 reproducible products (6.4 fragments per primer) were obtained from the 21 primers used. The number of bands per primer ranged from 4 to 11, whereas the number of polymorphic bands ranged from 3 to 10, corresponding to 83% of the amplification products. The dendrogram based on unweighted pair-group cluster analysis using Jaccard's index includes three major groups according to their origin: (1) cultivars from the Eastern and Central Mediterranean areas, (2) some Italian and Spanish cultivars, and (3) cultivars from the Western Mediterranean zone. The pattern of genetic variation among olive cultivars from three different Mediterranean zones (West, Centre and East) was analysed by means of the analysis of molecular variance (AMOVA). Although most of the genetic diversity was attributable to differences of cultivars within Mediterranean zones (96.86%) significant phi-values among zones (phi(st) = 0.031; p < 0.001) suggested the existence of phenotypic differentiation. Furthermore, the AMOVA analysis was used to partition the phenotypic variation of Spain, Italy (Western region), Greece and Turkey (Eastern region) into four categories: among regions, among countries (within regions), within countries, and among and within countries of each region. Most of the genetic diversity was attributable to differences among genotypes within a country. These results are consistent with the predominantly allogamous nature of Olea europaea L. species. This paper indicates the importance of the study of the amount and distribution of genetic diversity for a better exploration of olive genetic resources and the design of plant breeding programmes.     

Assessment of genetic diversity in cultivars of white clover (Trifolium repens L.) detected by SSR polymorphisms.

White clover (Trifolium repens L.) is an important temperate pasture legume that plays a key role as a companion to grass species, such as perennial ryegrass (Lolium perenne L.). Due to the outbreeding nature of white clover, cultivars are highly heterogeneous. Genetic diversity was assessed using 16 elite cultivars from Europe, North and South America, Australia, and New Zealand. Fifteen simple sequence repeat markers that detect single, codominant polymorphic genetic loci were selected for the study. The genetic relationships among individuals were compared using phenetic clustering, and those among cultivars were compared using nonmetric multidimensional scaling. Intrapopula tion variability exceeded interpopulation variability, with substantial overlap among populations and weak interpopula tion differentiation. No obvious or significant differentiation was observed on the basis of morphology or geographic origin of the cultivars. The number of parental genotypes used to derive each cultivar was not a major determinant of genome-wide genetic diversity. The outcomes of this assessment of genetic variation in elite white clover germplasm pools have important implications for the feasibility of molecular marker-based cultivar discrimination, and will be used to assist the design of linkage disequilibrium mapping strategies for marker-trait association.     

Introgression from modern hybrid varieties into landrace populations of maize (Zea mays ssp. mays L.) in central Italy

Landraces are domesticated local plant varieties that did not experience a deliberate and intensive selection during a formal breeding programme. In Europe, maize landraces are still cultivated, particularly in marginal areas where traditional farming is often practiced. Here, we have studied the evolution of flint maize landraces from central Italy over 50 years of on-farm cultivation, when dent hybrid varieties were introduced and their use was widespread. We have compared an 'old' collection, obtained during the 1950s, before the introduction of hybrids, and a recent collection of maize landraces. For comparison, a sample of maize landraces from north Italy, and of improved germplasm, including hybrids and inbred lines were also used. A total of 296 genotypes were analysed using 21 microsatellites. Our results show that the maize landraces collected in the last 5–10 years have evolved directly from the flint landrace gene pool cultivated in central Italy before the introduction of modern hybrids. The population structure, diversity and linkage disequilibrium analyses indicate a significant amount of introgression from hybrid varieties into the recent landrace populations. No evidence of genetic erosion of the maize landraces was seen, suggesting that in situ conservation of landraces is an efficient strategy for preserving genetic diversity. Finally, the level of introgression detected was very variable among recent landraces, with most of them showing a low level of introgression; this suggests that coexistence between different types of agriculture is possible, with the adoption of correct practices that are aimed at avoiding introgression from undesired genetic sources.     

Patterns of subspecies genetic diversity among oilseed <Emphasis Type="Italic">Brassica rapa</Emphasis> as revealed by agro-morphological traits and SSR markers

Brassica rapa (2n = 20, AA genome) is an important oil yielding species of the family Brassicaceae and characterized by wide range of genetic and morphological subtypes suitable for cultivation under diverse agro-climatic regions of India. In this study, genetic diversity among three subspecies of B. rapa including yellow sarson, toria and outlier brown sarson was estimated using various agro-morphological traits and simple sequence repeat (SSR) markers. Maximum variability was recorded for siliqua angle (Coefficient of variation = 30.9%), followed by seeds/siliqua (CV = 18.8%), leaf length (CV = 10%) and plant height (CV = 16.8%). Principal component analysis explained more than 50% of the total observed morphological variability for first two components. Of the 107 SSR markers tested, 80 generated reproducible, clear and distinct amplicons of which, 65 (81.25%) were found polymorphic. The number of alleles at each locus ranged from 2 to 7, with an average of 3.03 alleles per marker. A total of 197 alleles were detected at 65 SSR loci with average PIC value of 0.457 and a mean resolving power of 3.04. Neighbor-Joining cluster analysis based on morphological traits and SSR markers separately classified all the 28 genotypes into five major groups. The population structure analysis resulted into three sub-populations with certain extent of admixture among the earlier established taxonomic sub-groups. Twenty-three unique alleles were detected in thirteen B. rapa varieties. The clustering analysis and principal coordinate analysis outlined the genetic relationships among different varieties belonging to the three subspecies of B. rapa. Genetically diverse genotypes as illustrated by score plots and from the clustering patterns brought out the wide range of diversity present among B. rapa genotypes and the underlying options available for selecting parental genotypes for hybridization and developing high yielding cultivars suitable for Indian conditions.     

The utilization of unreplicated trials for conservation and quantification of intravarietal genetic diversity of rarely grown ancient grapevine varieties

The conservation of the genetic diversity within ancient grapevine varieties, which are rarely grown nowadays and are subject to genetic erosion, is extremely important in guaranteeing future utilization. This article studies the use of unreplicated trials as a way of preserving the genetic variability of those varieties and quantifying their intravarietal diversity as the raw material for future selections. Through simulation, several experimental situations were generated, varying the numbers of test and check treatments and the type of experimental design associated with the check treatments (augmented randomized complete block design and alpha–alpha design). The quality of the estimates of genotypic variance increased with the increase of the number of genotypes in the conservation collection and with the frequencies of plots with check genotypes. A greater precision was achieved in the quantification of the genetic diversity of ancient grapevine varieties when collections of over 250 genotypes and a minimum of 33 % of plots containing check genotypes were used.     

Integrating gene flow, crop biology, and farm management in on-farm conservation of avocado (Persea americana, Lauraceae)

Maintaining crop diversity on farms where cultivars can evolve is a conservation goal, but few tools are available to assess the long-term maintenance of genetic diversity on farms. One important issue for on-farm conservation is gene flow from crops with a narrow genetic base into related populations that are genetically diverse. In a case study of avocado (Persea americana var. americana) in one of its centers of diversity (San Jerónimo, Costa Rica), we used 10 DNA microsatellite markers in a parentage analysis to estimate gene flow from commercialized varieties into a traditional crop population. Five commercialized genotypes comprised nearly 40% of orchard trees, but they contributed only about 14.5% of the gametes to the youngest cohort of trees. Although commercialized varieties and the diverse population were often planted on the same farm, planting patterns appeared to keep the two types of trees separated on small scales, possibly explaining the limited gene flow. In a simulation that combined gene flow estimates, crop biology, and graft tree management, loss of allelic diversity was less than 10% over 150 yr, and selection was effective in retaining desirable alleles in the diverse subpopulation. Simulations also showed that, in addition to gene flow, managing the genetic makeup and life history traits of the invasive commercialized varieties could have a significant impact on genetic diversity in the target population. The results support the feasibility of on-farm crop conservation, but simulations also showed that higher levels of gene flow could lead to severe losses of genetic diversity even if farmers continue to plant diverse varieties.     

On the origin of the invasive olives (Olea europaea L., Oleaceae)

The olive tree (Olea europaea) has successfully invaded several regions in Australia and Pacific islands. Two olive subspecies (subspp. europaea and cuspidata) were first introduced in these areas during the nineteenth century. In the present study, we determine the origin of invasive olives and investigate the importance of historical effects on the genetic diversity of populations. Four invasive populations from Australia and Hawaii were characterized using eight nuclear DNA microsatellites, plastid DNA markers as well as ITS-1 sequences. Based on these data, their genetic similarity with native populations was investigated, and it was determined that East Australian and Hawaiian populations (subsp. cuspidata) have originated from southern Africa while South Australian populations (subsp. europaea) have mostly derived from western or central Mediterranean cultivars. Invasive populations of subsp. cuspidata showed significant loss of genetic diversity in comparison to a putative source population, and a recent bottleneck was evidenced in Hawaii. Conversely, invasive populations of subsp. europaea did not display significant loss of genetic diversity in comparison to a native Mediterranean population. Different histories of invasion were inferred for these two taxa with multiple cultivars introduced restoring gene diversity for europaea and a single successful founder event and sequential introductions to East Australia and then Hawaii for cuspidata. Furthermore, one hybrid (cuspidata x europaea) was identified in East Australia. The importance of hybridizations in the future evolution of the olive invasiveness remains to be investigated.     

Assessment of Genetic Diversity in <Emphasis Type="Italic">Trigonella foenum-graecum</Emphasis> Based on Nuclear Ribosomal DNA,Internal Transcribed Spacer and RAPD Analysis

Fenugreek (Trigonella foenum-graecum) is receiving global attention due to rare medicinal properties of significance to human health. Gene banks possess scanty germplasm and very little background information regarding its genetic variability that has hampered its improvement. We investigated the extent of variability among 17 Indian varieties of fenugreek using phenotypic and genetic markers. Multilocus genotyping by ten random amplified polymorphic DNA (RAPD) primers detected an average of intraspecific variations amounting to 64.7% polymorphism in banding patterns. Analysis of molecular variance indicated that a greater proportion of total genetic variation exists within population (91%) rather than among populations. Higher values of Nei’s gene diversity (h) and Shannon Information Index (i) and genetic distance analysis validate higher genetic diversity among Indian fenugreek varieties. SNPs at 14 sites of rDNA region revealed further lineages of distinct varieties with main RAPD clusters. The representative sequences of each subgroup and all distinct varieties have been submitted to NCBI database and assigned Gen Accession numbers HM 176640–176649. The measures of relative genetic distances among varieties of fenugreek did not completely correlate with the geographical distances of places of their development. The homogeneous phenotypic markers proved insufficient in exhibiting genetic divergence among fenugreek varieties studied. Eventually, the knowledge of their genetic relationships, DNA bar coding and phylogenies might contribute for the designing of intraspecific crosses between cultivars of this fenugreek collection with potential interest in seed spices breeding programme.     

The distinctive population structure of Colletotrichum species associated with olive anthracnose in the Algarve region of Portugal reflects a host–pathogen diversity hot spot

Anthracnose ( Colletotrichum spp.) is an important disease of olive fruits. Diversity and biogeographic relationships of the olive anthracnose pathogens in the Algarve (Portugal) were investigated, along with host association patterns and disease levels during 2004–2007, to test the hypothesis that this region is a host–pathogen diversity hot spot. Diverse Colletotrichum acutatum and Colletotrichum gloeosporioides populations were identified based on rRNA-internal transcribed spacer and partial β-tubulin 2 gene sequences of 95 isolates. Spatial and temporal variations in the occurrence of the eight genetic entities of the pathogens were linked to olive biogeography. Disease occurrence patterns suggest that C. acutatum populations are more stable pathogens, while C. gloeosporioides populations appear to be more influenced by favourable conditions. Three unique C. acutatum populations were identified, but none of the eight populations were dominant, with the most frequent type representing only 27%. Thus, the population structure of olive anthracnose pathogens in the Algarve is distinct from other parts of Portugal and other world locations, where only one or two genetic entities are dominant. This pattern and level of genetic diversity in a restricted area, where oleaster (wild olive tree), ancient landraces and modern cultivars of olive occur in close proximity, suggests the Algarve as a centre of diversity of the anthracnose pathogens and corroborates recent work suggesting western Mediterranean as an important centre of olive diversity and domestication.     

Fertilization and defoliation frequency affect genetic diversity of Festuca pratensis Huds. in permanent grasslands

Permanent pastures and meadows are species-rich vegetation systems that play an important role in the ecology and agriculture of temperate climates. Intensive management reduces species diversity and may also influence the genetic diversity within individual species and populations. The objective of this study was to assess genetic variability of meadow fescue, an important component of species-rich grasslands, and to determine whether fertilization and defoliation frequency influence genetic variability within natural populations. Genetic diversity of six natural populations and three cultivars of Festuca pratensis was investigated using randomly amplified polymorphic DNA (RAPD) markers and agronomic traits. Samples of natural populations were taken from two unrelated long-term experiments, where treatments had been applied for 11–38 years. RAPD analysis detected a clear genetic distinction of the cultivars from the natural populations. Genetic variability within cultivars was lower than within natural populations. Analysis of molecular variance ( AMOVA ) showed a significant effect of management on genetic variability. Fertilization and frequent defoliation led to a reduction in genetic variability within natural populations. Analysis of agronomic traits was only partially congruent with the results of RAPD analysis. This study shows that significant genetic variability exists within cultivars and natural populations of Festuca pratensis and can be reduced by intensive management.     

Genetic diversity and relationships of wild and cultivated olives in Turkey

In the present study, we proposed to determine the genetic diversity and relationships between local cultivars and wild olive trees from three important olive-growing regions, i.e., Marmara, Aegean, and Mediterranean, of Turkey. This is the first known large-scale molecular study to investigate the relationships between local cultivars and wild olives from the eastern Mediterranean basin. Two hundred and four oleaster trees and 27 cultivars were sampled to represent molecular diversity. We used 11 simple sequence repeat and 13 sequence-related amplified polymorphism markers to assess genetic variations and inter-relationships among the samples. The results of the analysis showed differences in the levels of allelic composition and heterozygosity between cultivated and wild olive trees. The observation of a high proportion of a certain wild-type genetic background in the cultivars may indicate the former use of local wild trees in olive domestication in Turkey, a possible autochthonal origin of cultivars. “Gemlik” was found to be the most common olive cultivar in the Marmara region and most of the wild olive samples from this region may be feral forms derived from cultivar seed spreading. The information obtained from this study can help to assist the management of an olive collection and sheds some light on the origin of Turkish olive cultivars.     

Genetic variation within and between winter wheat genotypes from Turkey, Kazakhstan, and Europe as determined by nucleotide-binding-site profiling

The genetic diversity within wheat breeding programs across Turkey and Kazakhstan was compared with a selection of European cultivars that represented the genetic diversity across eight European countries and six decades of wheat breeding. To focus the measure of genetic diversity on that relevant to disease-resistant phenotypes, nucleotide-binding-site (NBS) profiling was used to detect polymorphisms associated with the NBS motifs found within the NBS--leucine-rich repeat (LRR) class of resistance (R) genes. Cereal-specific NBS primers, designed specifically to the conserved NBS motifs found within cereal R-genes, provided distinct NBS profiles. Although the genetic diversity associated with NBS motifs was only slightly higher within the Eastern wheat genotypes, the NBS profiles produced by Eastern and European wheat lines differed considerably. Structure analysis divided the wheat genotypes into four groups, which compared well with the origin of the wheat genotypes. The highest levels of genetic diversity were seen for the wheat genotypes from the Genetic Resource Collection held in Ankara, Turkey, as wheat genotypes within breeding programs were genetically more similar. The wheat genotypes from Kazakhstan were the most similar to the European cultivars, reflecting the significant number of eastern European cultivars used in the breeding program in Kazakhstan. In general, the NBS profiles suggested that NBS-LRR R-gene usage in winter wheat breeding in Turkey and Kazakhstan differed from that deployed in European cultivars.     

The genetic structure of populations of the vulnerable aquatic macrophyte <Emphasis Type="Italic">Ranunculus nipponicus</Emphasis> (Ranunculaceae)

Ranunculus nipponicus (Makino) Nakai is a vulnerable aquatic macrophyte in the Kinki district, which is the southernmost distribution of this species in Japan. The genetic diversity and structure within and among eleven extant populations were assessed using the inter-simple sequence repeats (ISSR) polymerase chain reaction in association with combinations of propagation pattern (clonal and/or seeds) and genotypic geographical structure. In total, 53 bands were amplified, of which 18 (34%) were polymorphic. Analysis of the ISSR bands identified 46 genotypes among 81 individuals from one stream population and 72 distinct genotypes among 147 individuals in the Kinki district. An unweighted pair group method with arithmetic mean (UPGMA) dendrogram showed some unity among upstream and downstream subpopulations within one stream and eleven populations. The Shannon index of genetic diversity was 0.109 for one stream population and 0.313 for total genetic diversity, suggesting relatively high genetic diversity. Analysis of molecular variance (AMOVA) revealed that 84.1% of the total genetic diversity occurred among populations and the remaining diversity (15.9%) occurred within populations. Significant genetic differentiation occurred among populations in the Kinki district. These results suggest that conservation of each population is important for maintaining genetic diversity of R. nipponicus in this district. An erratum to this article can be found at