Plant tissue cultures from four Tuscan globe artichoke cultivars

The aim of the study was to examine the possibility of propagating in vitro four of the most common cultivars in Tuscany (central Italy): Terom, Violetto di Toscana, Chiusure and Empolese. The first three belong to the “Violetti” group, while cv Empolese belongs to the “Romaneschi” group. Explants were cultured on an induction medium (IM), which is a modified MS medium consisting of nitrate concentrations reduced by one quarter, 0.8 mg L⁻¹ 6-benzylaminopurine (BA) and 0.2 mg L⁻¹ 3-indole butyric acid (IBA). Explants were then transferred to a proliferation medium (PM) consisting of the same basal medium together with 0.03 mg L⁻¹ BA and 0.05 mg L⁻¹ gibberellic acid (GA₃). A rooting double-phase was then established. The pre-rooting medium (PRM), consisting of a basal MS medium with half strength nitrate concentrations, 0.5 mg L⁻¹ indole-3-acetic acid (IAA) and 1 mg L⁻¹ paclobutrazol (PBZ) was used for two weeks. Over the next four weeks, a rooting medium (MR) was used, consisting of a basal MS medium with 2 mg L⁻¹ β-cyclodextrin and 2 mg L⁻¹ α-naphthaleneacetic acid sodium salt (NAA). The cv Empolese provided the highest number of proliferated explants and rooted plantlets using the method described.     

Ontology annotation: mapping genomic regions to biological function

With numerous whole genomes now in hand, and experimental data about genes and biological pathways on the increase, a systems approach to biological research is becoming essential. Ontologies provide a formal representation of knowledge that is amenable to computational as well as human analysis, an obvious underpinning of systems biology. Mapping function to gene products in the genome consists of two, somewhat intertwined enterprises: ontology building and ontology annotation. Ontology building is the formal representation of a domain of knowledge; ontology annotation is association of specific genomic regions (which we refer to simply as 'genes', including genes and their regulatory elements and products such as proteins and functional RNAs) to parts of the ontology. We consider two complementary representations of gene function: the Gene Ontology (GO) and pathway ontologies. GO represents function from the gene's eye view, in relation to a large and growing context of biological knowledge at all levels. Pathway ontologies represent function from the point of view of biochemical reactions and interactions, which are ordered into networks and causal cascades. The more mature GO provides an example of ontology annotation: how conclusions from the scientific literature and from evolutionary relationships are converted into formal statements about gene function. Annotations are made using a variety of different types of evidence, which can be used to estimate the relative reliability of different annotations.     

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.     

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.     

The phenology of seed‐propagated globe artichoke

The ability of globe artichoke to produce inflorescences (capitula) during the autumn when the market price is highest is lost when plants are propagated from seed, as are most F₁ hybrid cultivars. To gain an understanding of the phenology of seed‐propagated globe artichoke, both vernalised and non‐vernalised seedlings grown from open pollinated progeny of ‘Spinoso sardo’ plants were transplanted into the field at two monthly intervals covering a whole year. Final leaf number and the date of unfolding of each leaf were used to calculate the phyllochrons. The average final leaf number increased from 24 to 88 in response to increasing daylength above an approximately 11 h threshold during the first 30 days after emergence, and this variate explained about 90% of the variation in the thermal time taken to reach the bolting stage. The average phyllochron ranged from 56 to 96°Cd among the six emergence dates and was the major underlying physiological cause of the loss of early flowering shown by seed‐propagated plants. Genotypic variation for phyllochron within an emergence date was strongly associated with variation for the length of the emergence to bolting stage period. The limited effect of vernalisation on development indicated that the late flowering tendency of seed‐propagated plants cannot be attributed to an unfulfilled cold requirement.     

Production and fingerprinting of virus-free clones in a reflowering globe artichoke

Most of the 24 viruses which infect globe artichoke are detrimental to the crop’s performance and hamper the development of a nursery activity in the respect of current EU legislation. We describe a procedure to sanitize globe artichoke “Brindisino” from Artichoke Italian latent virus (AILV) and Artichoke latent virus (ArLV), while preserving its valuable early flowering trait. ArLV was successfully eliminated by meristem-tip culture, while AILV was removed when two rounds of meristem-tip culture were spaced out with in vitro thermotherapy. In vivo thermotherapy, followed by meristem-tip culture, was also successful in producing virus-free material but was less efficient in terms of the number of plants recovered post treatment. Due to the multi-clonal composition of the populations at present in cultivation, the selected and sanitised clones were fingerprinted by applying microsatellite and AFLP (amplified fragment length polymorphism) markers. One AFLP primer combination produced 28 informative fragments used to evaluate genetic relatedness among the clones in study. Our results demonstrates that AFLP-based molecular fingerprinting enables to verify the true to clone correspondence in nurseries, ensure the effective correspondence between the real and the declared identity of a clone, so that to avoid commercial frauds, and might represents a valuable tool for assessing somaclonal variation occuring during ‘in vitro’ propagation.     

Structural families of genomic microsatellites

We present an analysis of tandem repeats of short sequence motifs (microsatellites) in twelve eukaryotes for which a large part of the genome has been sequenced and assembled. The pattern of motif abundance varies significantly in different species, but it is very similar in different chromosomes of the same species. The most abundant repeats can be classified in two main families. The first family has a rigid conformation, with purines in one strand and pyrimidines in the complementary strand, mainly A(n)/T(n) and (AG)(n)/(CT)(n). The second family has alternating, flexible sequences, such as (AT)(n), (AC)(n) and related sequences. In the pluricellular organisms the relative frequency of both families is rather constant. These observations indicate that microsatellites have structural information and may be involved in the organization of chromatin fibers and in chromosome architecture in general. An additional intriguing finding is the absence of microsatellites with sequences which appear to be forbidden, such as (AATT)(n).     

Characterization and mapping of canine microsatellites isolated from BAC clones harbouring DNA sequences homologous to seven human genes

Human primers specific for the genes LEP, HBB, PAX3, ESR2, TPH1, ABCA4 and ATP2A2 were used to identify clones in a canine BAC library. Subcloning of the positive BACs in plasmids, screening with microsatellite motifs and subsequent sequencing allowed for the identification of eight novel microsatellites. The presence of the gene of interest was confirmed by sequencing the polymerase chain reaction (PCR) products amplified in the positive BACs. Fluorescent in situ hybridization (FISH) using the positive BACs as probes allowed for the chromosomal localization of the insert DNAs in two canid species, dog (Canis familiaris) and red fox (Vulpes vulpes). The use of gene-associated microsatellites may accelerate the identification of candidate genes for phenotypic traits in linkage studies.     

Characterisation and pathogenicity of bacteria from shoot tips of the globe artichoke (Cynara scolymus L.)

Bacteria have been isolated from shoot tips of symptomless globe artichoke plants. These were identified as Pseudomonas fluorescens, Pseudomonas marginalis, Pseudomonas spp., Serratia liquefaciens, Enterobacter agglomerans/Erwinia, Agrobacterium radiobacter, an unidentified member of Rhizobiaceae and another classified in the “corynebacteria” group. The most frequently isolated species was P. fluorescens, biovars II and III. The endogenous character of these bacteria was studied in plants growing in vitro and in the open field. P. fluorescens, P. marginalis, S. liquefaciens and E. agglomerans/Erwinia caused symptoms in plants growing in vitro, but only P. fluorescens biovar II and P. marginalis produced symptoms in plants growing in open fields. Differences in pathogenicity were observed on inoculated plants growing in vitro or in the open field. This suggests that several endophytic bacterial species may be responsible for the high levels of contaminants found during the micropropagation of globe artichoke.     

2-D DIGE analysis of UV-C radiation-responsive proteins in globe artichoke leaves

Plants respond to ultraviolet stress inducing a self-defence through the regulation of specific gene family members. The UV acclimation is the result of biochemical and physiological processes, such as enhancement of the antioxidant enzymatic system and accumulation of UV-absorbing phenolic compounds (e.g. flavonoids). Globe artichoke is an attractive species for studying the protein network involved in UV stress response, being characterized by remarkable levels of inducible antioxidants. Proteomic tools can assist the evaluation of the expression patterns of UV-responsive proteins and we applied the difference in-gel electrophoresis (DIGE) technology for monitoring the globe artichoke proteome variation at four time points following an acute UV-C exposure. A total of 145 UV-C-modulated proteins were observed and 119 were identified by LC-MS/MS using a ～144,000 customized Compositae protein database, which included about 19,000 globe artichoke unigenes. Proteins were Gene Ontology (GO) categorized, visualized on their pathways and their behaviour was discussed. A predicted protein interaction network was produced and highly connected hub-like proteins were highlighted. Most of the proteins differentially modulated were chloroplast located, involved in photosynthesis, sugar metabolisms, protein folding and abiotic stress. The identification of UV-C-responsive proteins may contribute to shed light on the molecular mechanisms underlying plant responses to UV stress.     

Genetic differentiation within and between isolated Algerian subpopulations of Barbary macaques (Macaca sylvanus): evidence from microsatellites

This study of wild-living Algerian Barbary macaques ( Macaca sylvanus ) was designed to examine genetic variability in subpopulations isolated in residual forest patches, in an attempt to obtain data on the effects of habitat fragmentation. The wild population of this species (estimated at a maximum of 15 000) is vulnerable and this study therefore has direct relevance for conservation measures. Data from five microsatellite loci were analysed for 159 individuals from nine different groups living in four isolates in Algeria. Genetic polymorphism was found to be relatively high (4–12 alleles per locus) compared with other genetic markers used in previous studies of this species; mean expected heterozygosity was 65%. The four isolates are all genetically distinct ( F _ST = 0; P < 0.001). Indeed, the results suggest that dispersal is limited even between some social groups within a single isolate. Genetic distances based on models not assuming stepwise mutation ( F _ST and chord distance) gave very similar results and are highly correlated with geographical distances within one isolate but not between isolates. This may indicate that isolation by distance exerts a significant influence within an isolate but that genetic drift prevails between the four isolates. After allowing for variation in sample size, we found no evidence of reduced allelic diversity within small isolates that may have been separated for 250 years or more. The surviving population of Algerian Barbary macaques taken as a whole still shows marked variability in microsatellite alleles, but maintenance of genetic variability over the long term will surely require effective protection of all isolates.     

The domestication of artichoke and cardoon: from Roman times to the genomic age

BACKGROUND: The history of domestication of artichoke and leafy cardoon is not yet fully understood and when and where it occurred remains unknown. Evidence supports the hypothesis that wild cardoon is the wild progenitor of both these crops. Selection for large, non-spiny heads resulted in artichoke and selection for non-spiny, large stalked tender leaves resulted in leafy cardoon. The two crops differ in their reproductive system: artichoke is mostly vegetatively propagated and perennial, while leafy cardoon is seed propagated and mostly grown as an annual plant. Here, new trends in artichoke cultivation are analysed, while the consequences of these tendencies on the conservation of artichoke genetic resources are highlighted. SCOPE: The historical and artistic records, together with recent literature on genetics and biosystematics, are examined with the aim of achieving a better understanding of the present-day knowledge on the domestication of these two crops. CONCLUSIONS: Historical, linguistic and artistic records are consistent with genetic and biosystematic data and indicate that the domestication of artichoke and cardoon diverged at different times and in different places. Apparently, artichoke was domesticated in Roman times, possibly in Sicily, and spread by the Arabs during early Middle Ages. The cardoon was probably domesticated in the western Mediterranean in a later period.     

Barley microsatellites: allele variation and mapping

Microsatellites have developed into a powerful tool for mapping mammalian genomes and first reports about their use in plants have been published. A database search of 228 barley sequences from GenBank and EMBL was made to determine which simple sequence repeat (SSR) motif prevails in barley. Nearly all types of SSRs were found. The (A)n and (T)n SSRs occurred more often than (C)n and (G)n for n10. Among the dinucleotide repeats, the (CG)n SSRs occurred least often. Trinucleotide repeats did not occur with n>7 and there is no correlation between the GC content in the trinucleotide motifs and the number of observed SSRs. Analysing 15 different microsatellites with 11 barleys yielded 2.1 alleles per microsatellite. Sequencing 25 putative microsatellites showed that the resolution capacity of highquality agarose gels was sufficient to determine differences of only three base paris. Five microsatellites were mapped on three different chromosomes of a barley RFLP map.     

Direct characterization of bovine microsatellites from cosmids: polymorphism and synteny mapping

A (TG)₈ oligonucleotide probe was used to screen 186 cosmids from a commercial bovine cosmid library. Of the 56 positive discovered, 7 were sequenced in the region of the microsatellite and analysed for polymorphism. These microsatellites, IDVGA-2, -3, -7, -8, -9, -10, -11 showed the following number of alleles and polymorphism information content (PIC) values (7/0.616, 8/0.693, 6/0.641, 5/0.643, 2/0.239, 10/0.844, 6/0.720). The microsatellites were also assigned to synteny groups as follows: IDVGA-2/U17, IDVGA-3/U16, IDVGA-7/U7, IDVGA-8/U29, IDVGA-9/U3, IDVGA-10/U19, IDVGA-11/A (probably U18).     

Polymorphic microsatellites isolated from the cave fish Astyanax fasciatus

Six polymorphic microsatellite loci from a library of the tetra Astyanax fasciatus from Mexico were isolated. Amplification and heterozygosity were tested in four cave and four surface populations. These loci were developed for population genetic study to detect gene flow between cave and surface populations.