Retrotransposon-based S-SAP as a platform for the analysis of genetic variation and linkage in globe artichoke.

A high copy number of retrotransposon sequences are present and widely dispersed in plant genomes. Their activity generates a considerable degree of sequence polymorphism. Here, we report the cloning of CYRE-5, a long-terminal repeat carrying retrotransposon-like sequence in Cynara cardunculus L., and its exploitation to develop a DNA fingerprinting assay across 22 accessions, including both cultivated (globe artichoke and cultivated cardoon) and wild (wild cardoon) types. The effectiveness of the sequence-specific amplified polymorphism (S-SAP) platform is compared with that of amplified fragment length polymorphism (AFLP). A genetic linkage analysis, based on a hybrid population between 2 globe artichoke varietal types, resulted in the inclusion of 29 S-SAP loci in the core genetic map, confirming their dispersed distribution across the globe artichoke genome.     

M‐AFLP‐based protocol for microsatellite loci isolation in Cynara cardunculus L. (Asteraceae)

Nine microsatellite markers for Cynara cardunculus L. were developed using a two‐step ‘primer extension’ procedure, based on microsatellite‐amplified fragment length polymorphism (M‐AFLP) technique. In the first step, highly enriched SSR gel profiles were produced and, from the derived sequences of selected bands, forward primers directed towards the microsatellite motif were designed. In the second step, the opposite microsatellite flanking sequence was isolated using a nested approach on a restricted‐ligated genomic fraction. Polymorphism was explored in 24 plants of wild cardoon (Cynara cardunculus L. var. sylvestris) as well as two accessions of both globe artichoke (Cynara cardunculus L. var. scolymus), and cultivated cardoon (Cynara cardunculus L. var. altilis).     

Isolation of microsatellite loci in artichoke (Cynara cardunculus L. var. scolymus)

We report the development of nine microsatellite markers in globe artichoke (Cynara cardunculus L. var. scolymus). Four markers were obtained from sequences available in GenBank and five were isolated using a biotin/streptavidin capture technique for (CA)_n and (CT)_n motifs directly from artichoke genomic DNA. Polymorphism was explored in 15 artichoke accessions that represent the genetic variation within cultivated varietal types. Inter‐specific amplification was tested using cultivated cardoon (C. cardunculus L. var. altilis DC.) and wild cardoon (C. cardunculus L. var. sylvestris Lam.). Primers and conditions for polymerase chain reaction (PCR) amplification of detected loci are described.     

Genetic mapping and annotation of genomic microsatellites isolated from globe artichoke

Cynara cardunculus includes three taxa, the globe artichoke (subsp. scolymus L. Hegi), the cultivated cardoon (var. altilis) and their progenitor, the wild cardoon (var. sylvestris). Globe artichoke is an important component of the Mediterranean rural economy, but its improvement through breeding has been rather limited and its genome organization remains largely unexplored. Here, we report the isolation of 61 new microsatellite loci which amplified a total of 208 alleles in a panel of 22 C. cardunculus genotypes. Of these, 51 were informative for linkage analysis and 39 were used to increase marker density in the available globe artichoke genetic maps. Sequence analysis of the 22 loci associated with genes showed that 9 are located within coding sequence, with the repetitive domain probably being involved in DNA binding or in protein–protein interactions. The expression of the genes associated with 9 of the 22 microsatellite loci was demonstrated by RT-PCR.     

Analysis of molecular genetic diversity of cardoon (Cynara cardunculus L.) in Tunisia

The objective of this study is to investigate the genetic diversity, the relationships among six Tunisian wild cardoon populations (Cynara cardunculus var. sylvestris) and a Tunisian's population of cultivated cardoon (Cynara cardunculus var. altilis DC) in seven different geographical locations (Tiurif, Bahra, Zriba, Bouficha, Enfidha, Beja and Wad mliz) from semi-arid and wet regions of Tunisia. Twenty-three selected microsatellite markers are used for a sample of 98 cardoon genotypes. The total of 243 alleles is detected in the studied populations and the number of alleles per locus ranged from six to 23. The dendrogram based on Nei's (1972) UPGMA method divides the seven studied populations to five clusters. These preliminary results show that microsatellites are effective tools for plant species characterization and the analysed populations have a high genetic variability and will be suitable as genetic stocks for conservation and sustainable utilization programs of Cynara cardunculus L. in Tunisia.     

Population structure of Cynara cardunculus complex and the origin of the conspecific crops artichoke and cardoon

Background and Aims

Globe artichoke and leafy cardoon, two crops within the same species Cynara cardunculus, are traditionally cultivated in the Mediterranean region and play a significant role in the agricultural economy of this area. The two cultigens have different reproductive systems: artichoke is generally vegetatively propagated, while leafy cardoon is seed propagated. The domestication events underlying the origin of both artichoke and cultivated cardoon from their wild relative and the area of occurrence are not yet fully understood. The aim of this study was to investigate population structure in wild cardoon, globe artichoke and leafy cardoon material and infer domestication events.

Methods

Thirty-five microsatellite (simple sequence repeat) markers, distributed in the C. cardunculus genome, and a large geographical and numerical sampling in southern Europe and North Africa were used to assess population structure and diversity.

Key Results

The results suggest the presence of two distinct domestication events for artichoke and leafy cardoon, and also suggest a new possible scenario, with western wild cardoon having originated from cultivated cardoon escaped from cultivation. Evidence was found for a demographic bottleneck in the past history of globe artichoke.

Conclusions

The results shed new light on the relationships between the three taxa of C. cardunculus and highlight relevant aspects on the evolution of domestication of two crops with a different reproductive system within the same species. It is proposed that the probable centre of origin of artichoke is located in southern Italy, probably Sicily.     

More than multiple introductions: Multiple taxa contribute to the genesis of the invasive California’s wild artichoke thistle

The history of some invasive species is so complex that their origins can be difficult to determine. One example of such invasive species is the California invasive known as “wild artichoke thistle” (Cynara cardunculus var. sylvestris), found in natural and disturbed ecosystems. Wild artichoke thistle is a Mediterranean native and the progenitor of two domesticated horticultural taxa, artichoke and cardoon. Different hypotheses regarding the origins of California plants have included introductions by 19th century Italian immigrants and the de-domestication (evolutionary reversion to wild-type morphology) of feral (escaped, free-living) cultivars. Using microsatellite markers, we compared the genetic constitutions of 12 artichoke thistle populations in California with possible progenitor populations: 17 Spanish and Italian wild populations and eight different artichoke and cardoon cultivars. Each California population was compared with its putative progenitors using STRUCTURE analysis. Our results suggest that California's artichoke thistle populations are polyphyletic. Surprisingly, two-thirds of California's populations closely matched populations from the Iberian Peninsula. Three populations matched domesticated artichoke. One population appears to have wild and cultivar hybrid ancestry. Alleles specific to Italian populations were found at low frequencies in some California plants, suggesting that Italian wild plants may have been in California, but have left a trivial genetic legacy. Given that the de-domesticated plants in this study appear to be as invasive as the wild taxon, we conclude with a discussion of the role that ferality and de-domestication may have in plant invasions.     

Graft union formation in artichoke grafting onto wild and cultivated cardoon: An anatomical study

In order to develop a non-chemical method such as grafting effective against well-known artichoke soil borne diseases, an anatomical study of union formation in artichoke grafted onto selected wild and cultivated cardoon rootstocks, both resistant to Verticillium wilt, was performed. The cardoon accessions Belgio (cultivated cardoon) and Sardo (wild cardoon) were selected as rootstocks for grafting combinations with the artichoke cv. Romolo. Grafting experiments were carried out in the autumn and spring. The anatomical investigation of grafting union formation was conducted by scanning electron microscopy (SEM) on the grafting portions at the 3rd, 6th, 10th, 12th day after grafting. For the autumn experiment only, SEM analysis was also performed at 30 d after grafting.     

Genetic diversity of globe artichoke landraces from Sicilian small-holdings: implications for evolution and domestication of the species

Globe artichoke (Cynara cardunculus var. scolymus) is native to the Mediterranean Basin, where it grows in close proximity with its ancestor wild cardoon (C. cardunculus var. sylvestris); its commercial production is mainly based on vegetatively propagated clones which guarantee high yields of marketable product (i.e. immature inflorescence or capitula). A collection of 24 landraces of globe artichoke was made from small-holdings in Sicily, which is assumed to be one of the possible centres of its domestication. These landraces have been cultivated for centuries by local farmers, mainly due to their culinary uniqueness. The collection was characterised for a combination of morphological traits and AFLP, gSSR and cpSSr markers. Molecular analyses included genotypes of wild cardoon collected from different sites in Sicily as well as accessions of the most widely grown Sicilian varietal types: the spiny ‘Spinoso di Palermo’ and the non-spiny ‘Violetto di Sicilia’. The landraces follow a gradient of ‘ennoblement’ towards either the domesticated spiny or the non-spiny types. ‘Cimiciusa di Mazzarino’ was an outlier, in that it resembled the cultivated forms with respect to its AFLP fingerprint, but was more closely related to the wild cardoon on the basis of SSR profile. This particular landrace presents an example of an intermediary form in the domestication process, although it could also have derived from introgression from sympatric wild cardoon, followed by farmer selection. The abundant genetic variation present demonstrates the key role of farmers’ practice in the maintenance of genetic diversity, which should be preserved because of its potential value for plant breeders.     

Amplified fragment length polymorphism for genetic diversity assessment in globe artichoke

Globe artichoke (Cynara cardunculus L. var. scolymus L.) is a diploid (2n=2x=34), predominantly cross-pollinated plant native to the Mediterranean basin, and Italy contains the richest primary cultivated gene pool. Commercial production is mainly based on perennial cultivation of vegetatively propagated clones that are highly heterozygous and segregate widely when progeny-tested. Analysis of the artichoke genome by means of molecular markers has been limited to a few studies; here we report on the genetic relatedness among 118 artichoke accessions, including clones belonging to the same varietal type, two accessions of cultivated cardoon (C. cardunculus L. var. altilis DC.) and four accessions of wild cardoon [C. cardunculus L. var. sylvestris (Lamk) Fiori] as measured by amplified fragment length polymorphism (AFLP). Eight primer combinations yielded a total of 667 bands, of which 519 were polymorphic. Genetic similarities among accessions were calculated according to Jaccards Similarity Index and used to construct a dendrogram based on the unweighted pair group method using arithmetic averages. Our results demonstrate that AFLP markers can be useful in evaluating Cynara cardunculus genetic diversity and in classifying accessions to phylogenetic groups based on their genetic similarity values. Genetic variation among artichoke clones belonging to the same varietal type was in some cases higher than that found among accessions differently named and coming from different areas. The lowest Jaccards Similarity Index found within a varietal type can be considered as a threshold for the identification of accessions which share an analogous genetic background. This will enable the selection of representatives in order to develop and manage a germplasm core collection as well as the identification of suitable material for future artichoke breeding efforts.Communicated by J.S. Heslop-Harrison     

Genetic map of artichoke × wild cardoon: toward a consensus map for <Emphasis Type="Italic">Cynara cardunculus</Emphasis>

An integrated consensus linkage map is proposed for globe artichoke. Maternal and paternal genetic maps were constructed on the basis of an F₁ progeny derived from crossing an artichoke genotype (Mola) with its progenitor, the wild cardoon (Tolfa), using EST-derived SSRs, genomic SSRs, AFLPs, ten genes, and two morphological traits. For most genes, mainly belonging to the chlorogenic acid pathway, new markers were developed. Five of these were SNP markers analyzed through high-resolution melt technology. From the maternal (Mola) and paternal (Tolfa) maps, an integrated map was obtained, containing 337 molecular and one morphological markers ordered in 17 linkage groups (LGs), linked between Mola and Tolfa. The integrated map covers 1,488.8 cM, with an average distance of 4.4 cM between markers. The map was aligned with already existing maps for artichoke, and 12 LGs were linked via 31 bridge markers. LG numbering has been proposed. A total of 124 EST-SSRs and two genes were mapped here for the first time, providing a framework for the construction of a functional map in artichoke. The establishment of a consensus map represents a necessary condition to plan a complete sequencing of the globe artichoke genome.     

Identification of unique expression signatures and therapeutic targets in esophageal squamous cell carcinoma

Background

The Asteraceae species Cynara cardunculus (2n?=?2x?=?34) includes the two fully cross-compatible domesticated taxa globe artichoke (var. scolymus L.) and cultivated cardoon (var. altilis DC). As both are out-pollinators and suffer from marked inbreeding depression, linkage analysis has focussed on the use of a two way pseudo-test cross approach.

Results

A set of 172 microsatellite (SSR) loci derived from expressed sequence tag DNA sequence were integrated into the reference C. cardunculus genetic maps, based on segregation among the F₁ progeny of a cross between a globe artichoke and a cultivated cardoon. The resulting maps each detected 17 major linkage groups, corresponding to the species?? haploid chromosome number. A consensus map based on 66 co-dominant shared loci (64 SSRs and two SNPs) assembled 694 loci, with a mean inter-marker spacing of 2.5?cM. When the maps were used to elucidate the pattern of inheritance of head production earliness, a key commercial trait, seven regions were shown to harbour relevant quantitative trait loci (QTL). Together, these QTL accounted for up to 74% of the overall phenotypic variance.

Conclusion

The newly developed consensus as well as the parental genetic maps can accelerate the process of tagging and eventually isolating the genes underlying earliness in both the domesticated C. cardunculus forms. The largest single effect mapped to the same linkage group in each parental maps, and explained about one half of the phenotypic variance, thus representing a good candidate for marker assisted selection.     

Construction of a reference molecular linkage map of globe artichoke (<Emphasis Type="Italic">Cynara cardunculus</Emphasis> var. <Emphasis Type="Italic">scolymus</Emphasis>)

The genome organization of globe artichoke (Cynara cardunculus var. scolymus), unlike other species belonging to Asteraceae (=Compositae) family (i.e. sunflower, lettuce and chicory), remains largely unexplored. The species is highly heterozygous and suffers marked inbreeding depression when forced to self-fertilize. Thus a two-way pseudo-testcross represents the optimal strategy for linkage analysis. Here, we report linkage maps based on the progeny of a cross between globe artichoke (C. cardunculus var. scolymus) and cultivated cardoon (C. cardunculus var. altilis). The population was genotyped using a variety of PCR-based marker platforms, resulting in the identification of 708 testcross markers suitable for map construction. The male map consisted of 177 loci arranged in 17 major linkage groups, spanning 1,015.5 cM, while female map was built with 326 loci arranged into 20 major linkage groups, spanning 1,486.8 cM. The presence of 84 loci shared between these maps and those previously developed from a cross within globe artichoke allowed for map alignment and the definition of 17 homologous linkage groups, corresponding to the haploid number of the species. This will provide a favourable property for QTL scanning; furthermore, as 25 mapped markers (8%) correspond to coding regions, it has an additional value as functional map and might represent an important genetic tool for candidate gene studies in globe artichoke.     

Isolation and functional characterization of a cDNA coding a hydroxycinnamoyltransferase involved in phenylpropanoid biosynthesis in <Emphasis Type="Italic">Cynara cardunculus</Emphasis> L

Background  

Cynara cardunculus L. is an edible plant of pharmaceutical interest, in particular with respect to the polyphenolic content of its leaves. It includes three taxa: globe artichoke, cultivated cardoon, and wild cardoon. The dominating phenolics are the di-caffeoylquinic acids (such as cynarin), which are largely restricted to Cynara species, along with their precursor, chlorogenic acid (CGA). The scope of this study is to better understand CGA synthesis in this plant.     

The genus Cynara L. (Asteraceae-Cardueae)

WIKLUND, A., 1992. The genus Cynara L. (Asteraceae-Cardueae). This study includes a taxonomic revision of the genus Cynara. Eight species and four subspecies are recognized, viz. C. algarbiensis, C. auranitica, C. baetica including subsp. baetica and subsp. maroccana (formerly known as C. hystrix), C. cardunculus including subsp. cardunculus and subsp. flavescens, C. cornigera, C. cyrenaica, C. humilis (formerly sometimes in the genus Bourgaea) and C. syriaca. The cultivated artichoke (formerly C. scolymus) and cardoon are both included in C. cardunculus. One species, C. tournefortii , is excluded from Cynara. A cladistic study of the genus is also undertaken and its morphology, anatomy and phytogeography are discussed.     

Mapping the genomic regions encoding biomass-related traits in <Emphasis Type="Italic">Cynara cardunculus</Emphasis> L

Cynara cardunculus, a member of the Asteraceae family, comprises the three taxa var. scolymus (globe artichoke), var. altilis (cultivated cardoon) and the ancestral var. sylvestris (wild cardoon). The substantial quantities of lignocellulosic biomass produced by these plants (up to 30.0 t/ha year^?1 dry matter by the cultivated cardoon) can be used either as a source of bioenergy and/or as raw material for paper pulp production. Here, genotyping-by-sequencing to an F₁ population derived from a cross between a globe artichoke (C3) and a cultivated cardoon (ALT) genotypes has been used to perform a genome-wide linkage analysis, leading to the elaboration of a pair of highly dense genetic maps, each derived from one of the two highly heterozygous parental genotypes. In both maps, the number of linkage groups (17) matched the species’ haploid chromosome number. The F₁ population was phenotyped over two seasons with respect to plant height, stem number, capitulum number, leaf and stem fresh weight, and the dry weight of the whole plant, the leaves, the stems, the capitula and the achenes. The phenotypic data were combined with the linkage maps to identify 81 quantitative trait loci, of which 50 were placed on the C3 map and 31 on the ALT map. The loci were scattered over 13 linkage groups, and were clustered within 27 genomic regions, 22 of which harboured two or more QTL. Ten of these regions were specific to the C3 map and six to the ALT map, while the other 11 were represented on both maps. The 27 regions harboured in all 1960 genes, 83% of which could be functionally annotated. An enrichment for certain gene ontology terms was noted for the gene content of the genomic regions harbouring loci influencing seed yield and the number/weight of stems.     

The isolation and mapping of a novel hydroxycinnamoyltransferase in the globe artichoke chlorogenic acid pathway

Background  

The leaves of globe artichoke and cultivated cardoon (Cynara cardunculus L.) have significant pharmaceutical properties, which mainly result from their high content of polyphenolic compounds such as monocaffeoylquinic and dicaffeoylquinic acid (DCQ), and a range of flavonoid compounds.     

Characterization and utilization of microsatellites in the <Emphasis Type="Italic">Coffea canephora</Emphasis> genome to assess genetic association between wild species in Kenya and cultivated coffee

Coffee is an important beverage crop in the world and has a significant contribution to Kenya’s economy. Here, we analyzed the genome-wide distribution of microsatellites in the Coffea canephora genome. A total of 159,041 SSRs were identified, with an overall density of 308 SSRs per Mb. Tetra-nucleotide repeats are the most abundant, accounting for 32 % of the total SSRs. AT-rich motifs are dominant across all SSR repeat units, while GC-rich motifs were generally rare. A set of 100 SSRs was selected to amplify 96 coffee accessions, including 10 wild accessions collected from Mt. Marsabit (Kenya). Of these SSRs, 33 % generated clear polymorphic bands among all tested accessions, with an average of 3.9 alleles per SSR locus. Wild coffee species from Mt. Marsabit showed a close genetic similarity with cultivated accessions in Kenya, suggesting that the wild species in Mt. Marsabit played an important role in the domestication of cultivated coffee in Kenya. Significantly low pairwise genetic divergence was observed between cultivated and wild accessions in Kenya, suggesting a relatively narrow level of genetic basis among coffee germplasm in Kenya. In addition, cultivated and wild coffee accessions in Kenya show a great divergence from those in other countries. Our results not only provide molecular tools for genetic studies in coffee but are also helpful for conservation and coffee breeding programs in Kenya.     

Mapping yield-associated QTL in globe artichoke

Globe artichoke (Cynara cardunculus var. scolymus), whose immature inflorescences (capitula) are consumed as a vegetable all over the world, contributes significantly to the agricultural economy of the Mediterranean basin. Here, we describe a QTL (quantitative trait loci) analysis aimed at elucidating the mode of inheritance of seven main and first-order capitulum traits. Mapping was carried out in an F₁ population obtained by crossing a globe artichoke with a cultivated cardoon (C. cardunculus var. altilis). A total of 100 QTL associated with the seven capitulum traits were mapped to 23 chromosomal regions, scattered over 12 of the 17 linkage groups. Among these, 73 were expressed in both growing seasons, while the others were only detected in one season. Up to nine QTL per trait were identified, and major QTL, responsible for some 20 % of the phenotypic variation, were detected for capitulum length, diameter, shape index and fresh weight. The QTL for correlated traits frequently co-localized, most likely due to pleiotropy. This study represents the first report on yield traits QTL in globe artichoke. The QTL identified, along with linked markers, particularly those located in four hot-spot QTL regions are of practical interest for crop improvement based on marker-assisted breeding.     

New genetic maps for globe artichoke and wild cardoon and their alignment with an SSR-based consensus map

An F1 mapping population was bred by crossing an accession of wild cardoon with a single Argentinian globe artichoke plant of the variety Estrella del Sur FCA with a view to generating new Cynara cardunculus linkage maps. Genotyping was conducting using a set of 553 SRAP, SSR, AFLP and SNP markers. The 1,465.5 cM map based on the segregation of alleles present in the wild cardoon parent comprised 214 loci distributed across 16 linkage groups (LGs), while the 910.1 cM globe artichoke-based map featured 141 loci falling into 12 LGs covering the total length. Three of the morphological traits (head spininess, leaf spininess and head color) for which the parents contrasted were inherited monogenically, and the genes conditioning them were mapped. A set of 48 co-dominant loci was used to align the LGs with those derived from a reference SSR-based consensus map of the species.