AFLP Markers Research Articles

QTL analysis and the development of closely linked markers for days to flowering in spring oilseed rape (Brassica napus L.)

Days to flowering (DTF) is an important trait impacting cultivar performance in oilseed rape (Brassica napus L.), but the interaction of all loci controlling this trait in spring-type oilseed rape is not fully understood. We identified quantitative trait loci (QTL) for variation in DTF in a doubled haploid (DH) population from the Qinghai–Tibet Plateau that includes 217 lines derived from a cross between spring-type oilseed rape (B. napus L.) line No. 5246 and line No. 4512, the latter of which is responsive to the effective accumulated temperature (EAT). A linkage map was constructed for the DH population, using 202 SSR and 293 AFLP markers. At least 22 DTF QTL were found in multiple environments. Four major QTL were located on linkage groups A7, C2, C8 and C8. Among these QTL, cqDTFA7a and cqDTFC2a were identified in five environments and individually explained 10.4 and 23.0 % of the trait variation, respectively. cqDTFC8, a major QTL observed in spring environments, and a unique winter environment QTL, qDTFC8-3, were identified; these QTL explained 10.0 and 46.5 % of the phenotypic variation, respectively. Minor QTL (for example, cqDTFC2c) and epistatic interactions seemed evident in this population. Two closely linked SSR markers for cqDTFA7a and cqDTFC8 were developed (G1803 and S034). BnAP1, a B. napus gene with homology to Arabidopsis thaliana that was identified as a cqDTFA7a candidate gene, played a major role in this study. The allelic effects of the major and minor QTL on DTF were further validated in the DH population and in 93 breeding genotypes.

Genetic diversity in Oryza glumaepatula wild rice populations in Costa Rica and possible gene flow from O. sativa.

Wild crop relatives are an important source of genetic diversity for crop improvement. Diversity estimates are generally lacking for many wild crop relatives. The objective of the present study was to analyze how genetic diversity is distributed within and among populations of the wild rice species Oryza glumaepatula in Costa Rica. We also evaluated the likelihood of gene flow between wild and commercial rice species because the latter is commonly sympatric with wild rice populations. Introgression may change wild species by incorporating alleles from domesticated species, increasing the risk of losing original variation. Specimens from all known O. glumaepatula populations in Costa Rica were analyzed with 444 AFLP markers to characterize genetic diversity and structure. We also compared genetic diversity estimates between O. glumaepatula specimens and O. sativa commercial rice. Our results showed that O. glumaepatula populations in Costa Rica have moderately high levels of genetic diversity, comparable to those found in South American populations. Despite the restricted distribution of this species in Costa Rica, populations are fairly large, reducing the effects of drift on genetic diversity. We found a dismissible but significant structure (θ = 0.02 ± 0.001) among populations. A Bayesian structure analysis suggested that some individuals share a significant proportion of their genomes with O. sativa. These results suggest that gene flow from cultivated O. sativa populations may have occurred in the recent past. These results expose an important biohazard: recurrent hybridization may reduce the genetic diversity of this wild rice species. Introgression may transfer commercial traits into O. glumaepatula, which in turn could alter genetic diversity and increase the likelihood of local extinction. These results have important implications for in situ conservation strategies of the only wild populations of O. glumaepatula in Costa Rica.

Genetic differentiation within multiple common grassland plants supports seed transfer zones for ecological restoration

Summary Ecological restoration of grasslands is increasingly based on regional seeds derived from predefined seed transfer zones. However, the degree and spatial pattern of genetic differentiation among provenances of different seed transfer zones is largely unknown. We assessed the genetic differentiation among eight out of 22 German seed transfer zones for seven common grassland species (Arrhenatherum elatius, Centaurea jacea, Daucus carota, Galium album, Hypochaeris radicata, Knautia arvensis and Lychnis flos‐cuculi) using AFLP markers. We analysed genetic population structure with AMOVA and Bayesian cluster analysis and tested for isolation by distance and isolation by environment. In all of the investigated species, almost all pairs of provenances were genetically differentiated. Bayesian cluster analysis revealed species‐specific numbers and spatial patterns of gene pools, with between two (Arrhenatherum) and eight clusters (Lychnis). Most investigated seed transfer zones represented a unique gene pool in the majority of the species. We found isolation by distance in four species, isolation by environment, driven by climatic seasonality, in three species, and a lack of both in three species. Thus, the observed genetic differentiation appears to be caused by both neutral and adaptive processes. Synthesis and applications. Our study shows that grassland plants are indeed strongly genetically differentiated across Germany supporting the strategy of seed transfer zones for ecological restoration. Although the predefined seed transfer zones are unlikely to match the exact genetic structure of many species, they serve their purpose by capturing a substantial amount of intraspecific genetic variation across species.

Morpho-agronomic and AFLP characterization to explore guar (Cyamopsis tetragonoloba L.) genotypes for the Mediterranean environment

Morpho-productive evaluation in combination with molecular studies has received great attention for their synergetic action in the characterization of plant species. Guar (Cyamopsis tetragonoloba (L.) Taub.) is an annual legume crop that in the last century gained great interest as an industrial crop due to the galactomannan content of its grain. Guar genotypes adapted to India and the United States have been selected, while genotypes suitable for the Mediterranean environment able to maximize seed production, are missing. In the present study valuable morpho-agronomic traits such as growth habit, pods per plant and seeds per pod were assessed and genetic traits were investigated as selection criterion for breeding purposes. Pods per plant resulted negatively associated with seeds per pod and with 1000 seed weight. Present data underline that morpho-agronomic evaluation together with molecular analysis through AFLP markers could be an appropriate approach to individuate elite lines for future breeding programs of the species. Among studied genotypes, Monument seems to show the best morpho-productive traits for cultivation in the Mediterranean environment such as a short crop cycle, an elevated height of the first fertile node, a lack of branches and a good seed production. Lewis also seems interesting, even though a little less early and with a greater number of branches, but also with a higher seed production.

Assessing the taxonomic status ofOsmoderma cristinae(Coleoptera: Scarabaeidae), endemic to Sicily, by genetic, morphological and pheromonal analyses

Resolving complexes of closely related and cryptic insect species can be challenging, especially when dealing with rare and protected taxa that are difficult to collect for genetic and morphological analyses. Until recently, populations of the genus Osmoderma (Scarabaeidae), widespread in Europe, were treated as a single species O. eremita (Scopoli, 1763) in spite of observed geographic variation in morphology. A previous survey using sequence data from the mtDNA cytochrome C oxidase I gene (COI) revealed the occurrence of at least two distinct lineages within this species complex: O. eremita in the west and O. barnabita Motschulsky, 1845, in the east. Interestingly, beetles confined to Sicily have been described as a distinct species, O. cristinae Sparacio, 1994, based on morphological traits. Only few Sicilian specimens were included in the former genetic analysis, and the results led to a still questionable taxonomic rank for these populations. To explore the robustness of the previous taxonomic arrangement for O. cristinae, a combination of genetic, morphological and pheromonal analyses was used. A 617-bp fragment of the COI gene, aligned with O. cristinae and O. eremita sequences already available in GenBank, showed a clear genetic divergence between the two species (interspecific mean distance = 6.6%). Moreover, results from AFLP markers sustained the distinction of the two species. In addition, geometric morphometric analyses of the shape of male genitalia revealed a clear differentiation between the two species. Via scent analysis and field trapping, we demonstrated the production of the sex pheromone (R)-(+)-γ-decalactone by males of O. cristinae, the attraction by conspecific individuals (mostly females) to this compound, and a lack of antagonistic effect of (S)-(-)-γ-decalactone. The fact that O. eremita and O. eremita use the same compound for mate finding suggests that this sex pheromone has not undergone a differentiation and probably the allopatry of these two species compensates for the absence of a mechanism to avoid cross-attraction. Our genetic and morphological data support the divergence of the two species and confirm the species status for O. cristinae, while sex pheromones are confirmed to be invariant among different species of the genus Osmoderma.

Development of a Genomic Resource and Quantitative Trait Loci Mapping of Male Calling Traits in the Lesser Wax Moth, Achroia grisella.

In the study of sexual selection among insects, the Lesser Waxmoth, Achroia grisella (Lepidoptera: Pyralidae), has been one of the more intensively studied species over the past 20 years. Studies have focused on how the male calling song functions in pair formation and on the quantitative genetics of male song characters and female preference for the song. Recent QTL studies have attempted to elucidate the genetic architecture of male song and female preference traits using AFLP markers. We continued these QTL studies using SNP markers derived from an EST library that allowed us to measure both DNA sequence variation and map loci with respect to the lepidopteran genome. We report that the level of sequence variation within A. grisella is typical among other Lepidoptera that have been examined, and that comparison with the Bombyx mori genome shows that macrosynteny is conserved. Our QTL map shows that a QTL for a male song trait, pulse-pair rate, is situated on the Z chromosome, a prediction for sexually selected traits in Lepidoptera. Our findings will be useful for future studies of genetic architecture of this model species and may help identify the genetics associated with the evolution of its novel acoustic communication.

Identification of QTLs for sex expression in dioecious and monoecious hemp (Cannabis sativa L.)

Hemp (Cannabis sativa L.) is a diploid species including both dioecious and monoecious cultivars with hetero- and homomorphic sex chromosomes, respectively. It displays a high plasticity of sex expression, i.e., the ratio of female and male flowers. In this study, we investigated the role of sex chromosomes in the genetic determinism of sex expression in dioecious and monoecious hemp. The experimental materials were three F1 segregating populations, two dioecious (C1 and C2: ‘Carmagnola’ ♀ × ‘Carmagnola’ ♂), and one monoecious (UF: ‘Uso 31’ × ‘Fedora 17’). A ‘sex’ phenotypic marker was mapped in C1 and C2. In total, 23, 42, and 26 AFLP markers (71 markers in total) were mapped to three, nine, and three co-segregation groups putatively located on sex chromosomes in C1, C2, and UF, respectively. Recombination rates with sex ranged from 0 to 0.5. Five sex-linked markers were detected in UF, revealing homologies between the X chromosomes of monoecious hemp and the X and Y chromosomes of dioecious hemp. Five QTLs associated with quantitative variations in sex expression were identified in each map. Four markers associated with variations in sex expression in UF segregated with sex or accounted for a putative QTL in C1 or C2. Two QTLs and three of these markers were mapped in UF in a region homologous to the sex-locus region of the dioecious maps. Given these results, conducting further research on the genetic determinism of sex expression in hemp using a quantitative approach appears relevant.

Natural epigenetic variation contributes to heritable flowering divergence in a widespread asexual dandelion lineage.

Epigenetic variation has been proposed to contribute to the success of asexual plants, either as a contributor to phenotypic plasticity or by enabling transient adaptation via selection on transgenerationally stable, but reversible, epialleles. While recent studies in experimental plant populations have shown the potential for epigenetic mechanisms to contribute to adaptive phenotypes, it remains unknown whether heritable variation in ecologically relevant traits is at least partially epigenetically determined in natural populations. Here, we tested the hypothesis that DNA methylation variation contributes to heritable differences in flowering time within a single widespread apomictic clonal lineage of the common dandelion (Taraxacum officinale s. lat.). Apomictic clone members of the same apomictic lineage collected from different field sites showed heritable differences in flowering time, which was correlated with inherited differences in methylation-sensitive AFLP marker profiles. Differences in flowering between apomictic clone members were significantly reduced after invivo demethylation using the DNA methyltransferase inhibitor zebularine. This synchronization of flowering times suggests that flowering time divergence within an apomictic lineage was mediated by differences in DNA methylation. While the underlying basis of the methylation polymorphism at functional flowering time-affecting loci remains to be demonstrated, our study shows that epigenetic variation contributes to heritable phenotypic divergence in ecologically relevant traits in natural plant populations. This result also suggests that epigenetic mechanisms can facilitate adaptive divergence within genetically uniform asexual lineages.

Strigolactone pathway genes and plant architecture: association analysis and QTL detection for horticultural traits in chrysanthemum.

Chrysanthemums are important ornamental plants with abundant phenotypic diversity. Especially in cut-flower breeding, shoot branching is important for the success of new varieties. To assess the genetic regulation of shoot branching and other horticultural important traits, we phenotyped and genotyped two types of chrysanthemum populations: a genotype collection of 86 varieties and a biparental F1-population (MK11/3) of 160 individuals. Using two different statistical approaches, a genome-wide association analysis and a single marker ANOVA, with AFLP marker data and candidate gene markers for shoot branching, we tried to identify markers correlated to the traits of interest. As expected for the outcrossing hexasomic chrysanthemums most of the phenotypic traits showed a continuous variation in both populations. With the candidate gene approach we identified 11 significantly associated marker alleles for all 4 strigolactone pathway genes BRC1, CCD7, CCD8 and MAX2 regulating shoot branching in the genotype collection. In the MK11/3 we detected seven markers for all candidate genes except MAX2 explaining a large proportion of the variation. Using anonymous AFLP markers in the GWA with the 86 genotypes and the single locus analysis with the F1-population we could detect 15 and 17 additional marker-trait associations, respectively. Our analyses indicate a polygenic inheritance of the shoot branching in the chrysanthemum, with a fundamental role of the strigolactone pathway genes BRC1, CCD7, CCD8 and MAX2 and we identified 50 associated markers to all traits under study. These markers could be used in the selection of the parental plants for breeding chrysanthemums to enrich them for positive alleles influencing plant architecture traits.

Genomewide prediction of tropical maize single-crosses

Genomic selection studies usually use few environmental models to predict progenies from biparental crosses. However, in a maize breeding program, numerous crosses need to be evaluated in multiple field trials to identify single-crosses with greater yield potential. In this study, 614 AFLP marker effects estimated in 250 maize single-crosses evaluated in 13 environments were used to assess the influence of training population size (N), number of markers (NM), genotype-by-environment interaction (G × E) and population structure on the prediction accuracy (rMG) for grain yield using RR-BLUP model. Cross validation analysis were performed within and across environments, and within and across groups of related single-crosses. Then, genomic selection was compared to phenotypic selection in identifying the best single-crosses in a maize breeding program scheme. In general, increasing the training population size and the number of markers did not led to higher accuracy estimates. Predicted accuracies from cross validation analysis within environments were significantly higher than between environments, indicating that the effect of G × E interaction was important. Accuracy estimates were also higher when training and validation sets were composed of related single-crosses. In all scenarios, wide intervals of accuracy were found, meaning that genomic prediction may not be effective depending on the single-crosses sampled. The use of genomic prediction in maize breeding programs was discussed emphasizing the need of a training set evaluated in multiple environments and designing a genomic selection experiment according to the population structure so as to reduce sample problems and maximize the accuracy and the success of prediction.

Development of specific SCAR markers identifying recessive genic male sterile gene in sesame (Sesamum indicum L.)

To develop a simple and convenient marker for genetic linkage of sesame male sterile gene,based on an AFLP marker,7 SCAR( sequence characterized amplified region) markers were developed using recessive genic male sterile two- type line 95ms- 5AB as fertile and sterile gene bulks. These 7 markers could detect bands from fertile bulk with 806 bp to 1 638 bp in size,and could not from sterile bulk. They were designated as PMC- 1to PMC- 7. These markers were used to detect 1 184 fertile and sterile plants of 95ms- 5AB with 0. 25% exchange rate. 4 F2 populations obtained from 95ms- 5A and different genotypes and 2 recessive genic male sterile lines were identified using these 7 SCAR markers. Result indicated that the SCAR markers were specific for 95ms- 5.

Genetic relationships among the genera Cicer L., Lathyrus L., Lens Mill., and Vicia L., together with similarity of Lens taxa based on morphological and AFLP markers

The molecular characterization of the seven taxa in the genus Lens Mill. with Vicia montbretii has not been performed, and species of the genera Vicia L., Lathyrus L., and Cicer L. using AFLP markers have not been elucidated so far. The objectives of the present study were the assessment of (i) genetic relationships among the genera Lens, Vicia, Lathyrus, and Cicer based on AFLP markers, (ii) relationships within the genus Lens including the accessions of L. lamottei Czefr., L. tomentosus Ladiz., and L. odemensis Ladiz., based on morphological and AFLP data, and (iii) position of the species Vicia montbretii (Fisch. and Mey.) Davis and Plitmann based on molecular markers. The genetic material consisted of seven Lens taxa comprising each three accessions of cultivated lentils (L. culinaris Medik.), L. orientalis (Boiss.) Ponert, L. tomentosus, L. odemensis, L. ervoides (Brign.) Grande, and L. nigricans (M.Bieb.) Godr., plus two L. lamottei accessions. Two Vicia montbretii accessions were also included to assess their relationship with the genus Lens Mill. As morphological markers the number of toothed stipules, stipule angle, aristae on peduncle, and pod pubescence were evaluated regarding lentil species. Of the 414 AFLP markers, 280 entailing the most informative and polymorphic were used for the molecular analysis. The genus Lens was genetically closer to the genera Vicia and Lathyrus than to the genus Cicer. V. montbretti was well separated from the genus Lens. The results indicated the following genetic relationship: Lens culinaris subsp. culinaris, Lens culinaris subsp. orientalis, Lens culinaris subsp. tomentosus, Lens culinaris subsp. odemensis, L. ervoides, L. lamottei, and L. nigricans.

Accessing genetic diversity levels of Brazilian genotypes of castor with AFLP and ISSR markers

Castor ( Ricinus communis L.), an important oilseed crop for the production of biofuels and high quality lubricants, has a long history of cultivation worldwide. However, very little is known about the genetic diversity of cultivars adapted to the semi-arid conditions of Northeastern Brazil, a strategic area for castor cultivation. Aiming to provide new approaches to select dominant markers for genetic diversity analysis among castor accessions from a Brazilian germplasm bank, we have tested 21 AFLP primer combinations and 16 ISSR primers with 27 genotypes from a Brazilian germplasm bank. PIC, MI and RP values were calculated for both marker systems and a strong correlation was observed among these parameters. Therefore, due to a higher degree of variation, primers could be better selected taking into account the RP values, which ranged from 0.231 to 18.231 (AFLP) and from 1.630 to 9.556 (ISSR). Dissimilarity values ranged from 0.189 to 0.504 in the combined analysis (AFLP + ISSR), with genotypes grouping into four main clusters, although without any evidence of genetic structuration (F st = 0.108). The successful association between genotypes and speci c marks highlighted the great potential of AFLP and ISSR markers to access the variability within the species. Additionally, by describing new polymorphic markers for genotype differentiation, we provide an important contribution to guide crosses between accessions with traits of interest.

Sequence Characterized Amplified Regions Linked with Apomictic Mode of Reproduction in Four Different Apomictic <i>Cenchrus</i> Species

The genus Cenchrus comprises tropical and sub-tropical forage grasses, many of which reproduce by aposporous apomixis. This mode of reproduction hampers genetic improvement of the species through conventional breeding; however, it may facilitate the fixing of heterosis in a hybrid. Apomixis can be deployed in plant breeding to maintain hybrid vigour over the generations, thereby accelerating the breeding process. Markers for apospory have been identified in grasses, but we have identified conserved genomic regions and apomixis-specific AFLP markers in eight Cenchrus spp. and converted them into sequence characterised amplified regions (SCARs). Seventeen of 94 identified polymorphic AFLP markers were located in the apospory-specific genomic region and were converted into SCAR markers. Only four (23.5%) were successfully converted into SCARs linked with an apomictic mode of reproduction in Cenchrus spp. The SCAR markers were validated in an F 2 population of C. ciliaris consisting of 48 apomictic and 38 sexual individuals. The SCARs were conserved across the four apomictic Cenchrus spp. with >98% sequence homology. In silico mapping of the SCARs based on sequence homology indicated synteny with segments of chromosome 12 of rice and chromosome 5 of maize. These markers would be very useful for genetic/molecular analyses of apomixis, comparative mapping studies and marker-assisted breeding of Cenchrus .

Development of a transposon-based marker system for mutation breeding in sorghum (Sorghum bicolor L.).

Under certain circumstances, transposable elements (TE) can create or reverse mutations and alter the genome size of a cell. Sorghum (Sorghum bicolor L.) is promising for plant transposon tagging due to its small genome size and its low content of repetitive DNA. We developed a marker system based on targeted region amplification polymorphisms (TE-TRAP) that uses the terminal inverted repeats (TIRs) of transposons. A total of 3816 class 2 transposons belonging to the PIF/Harbinger family were identified from the whole sorghum genome that produced five primers, including eight types of TIRs. To define the applicability and utilization of TE-TRAP, we used 21 individuals that had been bred after ɤ-ray irradiation. In total, 31 TE-TRAP, 16 TD, and 21 AFLP primer combinations generated 1133, 223, and 555 amplicons, respectively. The percent polymorphic marker was 62.8, 51.1, and 59.3% for the TE-TRAP, TD, and AFLP markers, respectively. Phylogenetic and principal component analyses revealed that TE-TRAP divided the 21 individuals into three groups. Analysis of molecular variance suggested that TE-TRAP had a higher level of genetic diversity than the other two marker systems. After verifying the efficiency of TE-TRAP, 189 sorghum individuals were used to investigate the associations between the markers and the ɤ-ray doses. Two significant associations were found among the polymorphic markers. This TE-based method provides a useful marker resource for mutation breeding research.

Development of new restorer lines for CMS ogura system with the use of resynthesized oilseed rape (Brassica napus L.).

Resynthesized (RS) oilseed rape (Brassica napus L.) is potentially of great interest for hybrid breeding. However, a major problem with the direct use of RS B. napus is the quality of seed oil (high level of erucic acid) and seed meal (high glucosinolate content), which does not comply with double-low quality oilseed rape. Thus, additional developments are needed before RS B. napus can be introduced into breeding practice. In this study, RS oilseed rape was obtained through crosses between B. rapa ssp. chinensis var. chinensis and B. oleracea ssp. acephala var. sabellica. RS plant was then crossed with double-low (00) winter oilseed rape lines containing the Rfo gene for Ogura cytoplasmic male sterility (CMS ogu) system. Populations of doubled haploids (DH) were developed from these F1 hybrids using the microspore in vitro culture method. The seeds of semi-RS DH lines were analyzed for erucic acid and glucosinolate content. Among the populations of semi-RS DHs four 00-quality lines with the Rfo gene were selected. Using 344 AFLP markers to estimate genetic relatedness, we showed that the RS lines and semi-RS lines formed clusters that were clearly distinct from 96 winter oilseed rape parental lines of F1 hybrids.

Morphological and AFLP-Based Genetic Diversity in Rosa platyacantha Population in Eastern Tianshan Mountains of Northwestern China

Morphological and AFLP-based genetic analyses of six natural populations of Rosa platyacantha in the Eastern Tianshan Mountains of Xinjiang in Northwestern China were performed. Nested analysis of variance models (ANOVA) showed that there were significant differences (P > 0.05) in morphological traits among and within the populations of R. platyacantha. The phenotypic variation coefficient (CV) of the traits varied from 9.78% to 37.71%, confirming the abundant phenotypic variations in the species. The average value of the phenotypic differentiation coefficient (Vst) was 27.50%, indicating a lower phenotypic diversity among populations than within populations. Principal component analysis (PCA) showed that the two decisive traits of ANH and TAW contributed to the proportion of 97.19% in the 1st principal component (PC), suggesting that the phenotypic variations of the traits primarily originated from the achene characters. The percentages of polymorphic loci (PPL), Nei's genetic diversity index (h) and Shannon information index (I) were highly similar among populations, with average values of 63.96%, 0.2361 and 0.3506, respectively. Moreover, the gene differentiation (Gst) and estimated value of gene flow (Nm) were 8.10% and 5.6766, respectively, indicating very low genetic diversity among populations. Clustering analysis based on morphological traits and AFLP markers showed that the populations were mainly grouped with similar altitudes and geographic distances, respectively. The Mantel test revealed that there was no significant correlation between the phenotypic and genetic diversity among populations, implying a possible influence of environments on the genotypes of R. platyacantha.

Association mapping for identification of ISSR and AFLP markers linked with resin yield in Himalayan Chir Pine (Pinus roxburghii Sarg

AbstractIn Pinus roxburghii, marker-trait associations were studied for resin yield using a set of 238 DNA based markers derived from 9 ISSR and 5 AFLP primer pairs. A total of 240 genotypes of P. roxburghii from a natural population in Chakrata division (Tiunee range), Uttarakhand (India) were evaluated for resin yield. Based on stable resin production in consecutive years, 53 genotypes that were best representatives of the variation in resin yield (0.25 to 8.0 kg/tree/year) were used for genotyping and association analysis. For carrying out the association studies, two approaches were used. First, using simple linear regression, resin yield was regressed on all 238 available polymorphic markers (148 ISSR markers and 90 AFLP markers) and second, evidence for association was examined applying population structure information using STRUCTURE and STRAT to avoid any spurious associations between a candidate marker and phenotype. In regression analysis, 16 ISSR and 12 AFLP markers showed significant association with resin yield. In the second approach, 18 ISSR and 18 AFLP loci were found to be associated with resin yield in the absence of population structure out of which 2 ISSR and 3 AFLP loci were common with those associated in regression analysis. After accounting for population structure, 38 ISSR, and 3 AFLP loci revealed association out of which one ISSR and one AFLP locus was found to be common with both simple linear regression and test of association in the absence of population stratification. There was no obvious correlation between population structure and resin yield.

Genetic and cytogenetic structure of wild lemon grass (Elionurus muticus ) populations

Abstract Elionurus muticus is a native aromatic grass from the Pampa biome that produces an essential oil that is rich in citral. Despite the importance of citral, few studies have examined this species. The aims of this work were to evaluate the genetic structure and to characterize cytogenetically natural populations collected from Brazil. Genetic characterization was performed using AFLP markers, and cytogenetics assessed the chromosome number, karyotype and meiosis. The studied populations had genetic variability, especially within populations, indicating the possibility of selecting plants with relevant characters. High variability also suggests the preferential occurrence of outcrossing in natural populations. Regular meiosis was observed in the cytogenetic analysis with chromosome number 2n=20. The karyotype of the species is presented for the first time, with the karyotype formula 3sm + 4a + 1saSAT.

Genetic diversity and pathogenicity of Sphaceloma rosarum (teleomorph Elsinoë rosarum) causing spot anthracnose on roses

Roses produced or grown in the field, as well as pot‐grown and cut roses, are attacked by different fungal pathogens causing leaf spot diseases. The incorrect identification and scoring of these pathogens and the lack of information about their genetic and pathotype diversity hamper resistance breeding. This is especially true for the hemibiotrophic ascomycete Sphaceloma rosarum, which is often confused with other fungi. Here for the first time, the genetic variability between isolates at both the molecular and morphological level is analysed. Eighty leaf spot samples were collected from different rose genotypes at five different locations, and 15 single conidial isolates established. All of the samples showed high morphological similarities to the reference isolate CBS 213.33 that was obtained from a public repository. By sequencing a part of the large subunit (LSU) of the 28S ribosomal RNA and phylogenetic analysis, high sequence similarities were shown to other Sphaceloma species for 13 of the isolates and the CBS reference. One of the isolates clustered with Septoria species and another clustered with Seimatosporium species. UPGMA clustering with 145 polymorphic AFLP markers resulted in five distinct groups in the majority rule consensus tree for the 14 S. rosarum isolates, including the CBS reference. Jaccard similarities ranged from 0·31 to 0·91. A detached leaf assay using a differential set of five rose genotypes led to the classification of the five tested isolates as five distinct pathotypes. Therefore, grouping depending on the avirulence gene diversity was clearly different from clustering using selectively neutral AFLP markers that were evenly distributed throughout the genome.