Search

"Many research works have unveiled how grapevine cultivars respond to environmental conditions; however, the molecular mechanisms underlying the interplay between clones (vegetatively propagated lines of selected mother plants) and environment need to be elucidated. This study was aimed to explore the complexity of the clone x environment interaction by investigating changes controlled by clone, vineyard or developmental stage. The transcriptome of berries collected over ripening in different vineyards from three 'Nebbiolo' clones was analyzed by integrating RNA-seq data with analysis of candidate genes, quantification of secondary metabolites and agronomical parameters. Transcripts associated to sugar transport and anthocyanin biosynthesis were differently modulated among clones, while genes involved in secondary metabolism and defence, such as stilbene synthase genes, were affected by vineyard, consistently with stilbenoid accumulation. This attests that clone-specific responses exert a role in shaping agronomic performances of a grape variety in different environments, thus providing indications for orienting viticultural practices in light of both cultivation area and clone choice."

The role of plant tyrosyl-DNA phosphodiesterase 1? in genome stability is studied using a Medicago truncatula MtTdp1?-depleted line. Lack of MtTdp1? results in a 39% reduction of methylated cytosines as compared to control. RNA-Seq analyses revealed that 11 DNA transposons and 22 retrotransposons were differentially expressed in the Tdp1?-2a line. And among them all DNA transposons (MuDR, hAT, DNA3-11_Mad) and seven retrotransposons (LTR/Gipsy, LTR/Copia, LTR and NonLTR/L1) were down-regulated, while remaining 15 retrotransposons were up-regulated. Results suggest that the occurrence of stress-responsive cis-elements as well as changes in the methylation pattern at the LTR promoters might be responsible for the enhanced retrotransposon transcription.

Grapevine (Vitis vinifera L.) is one of the major fruit crop worldwide with thousands of cultivated varieties. Among the important Italian cultivars, 'Nebbiolo' is characterized by a wide genetic variability expressed by a high number of clones (i.e. vegetatively propagated lines of selected mother plants) registered in the National Catalogue. However, little morphological differences exist between clones and often it is very difficult to visually distinguish the different clones, and the Simple Sequence Repeats (SSRs) extensively used as markers, showed low or no success in discriminating clones. The aim of this on-going study is to identify a set of single nucleotide polymorphisms (SNPs) able to discriminate the registered 'Nebbiolo' clones, setting up an effective and rapid SNP genotyping protocol based on the TaqMan® probes technology. Three 'Nebbiolo' clones were sequenced by Illumina technology (CVT 71, CVT 185 and CVT 423) and processed reads were mapped against the PN40024 reference genome. SNPs were first detected using the GATK-UnifiedGenotyper tool with default parameters followed by filtering and by a calibration step. SNPs were validated as true positive by Sanger sequencing, and nine of the 11 SNPs confirmed to date were successfully used to perform TaqMan® genotyping assays in genomic DNA from 37 'Nebbiolo' clones. Combining results obtained from each single SNP assay, we classified the 'Nebbiolo' clones in six main groups linked to the geographical origin of accessions. Furthermore, these SNPs were tested in other grapevine cultivars, and interestingly in some case, these markers are polymorphic also in clones of 'Brachetto' and 'Nebbiolo rosè'. This work is in progress and the methodology will be applied in the vine supply chain to ensure a traceability from grape nurseries up to wine industry, providing a product of sure origin to the consumers. This research was funded by Fondazione Cassa di Risparmio di Cuneo: "Nebbiolo Genomics"; Fondazione Cassa di Risparmio di Torino: "Track W/Vine"; and Regione Piemonte: "MicroGenVine".

The success of the grape and wine-related agribusiness depends on yield and quality of the grapes produced, which is heavily influenced by genetic diversity (cultivars and clones) and environment (terroir). Among the important Italian varieties, 'Nebbiolo' is one of the best examples of this connection. The behavior of three 'Nebbiolo' clones (CVT71, CVT185, CVT423) grafted on the same rootstock (Kober 5BB) and grown in three different experimental vineyards in the typical growing areas of Piedmont (Italy), was analyzed in relation to the different soil and environmental conditions. In two years (2013 and 2014) the agronomical and biochemical performances of those vines were monitored from winter pruning to harvest, recording vigor, yield, juice composition and anthocyanin profile. RNA-Seq analyses were performed on berries collected in three different developmental stages during the vegetative season: berry pea size stage, veraison and at harvest (24 °Brix). RNA-seq data were analyzed through a data mining process with the aim of investigating the relationships between time course of gene expression profiles and environmental variables, and of quantifying how much these variables affect the gene expression. The most important changes in the berry transcriptome are related to the different developmental stages. However, also vineyard and genotype induce significant modulation of several genes. For example, in a vineyard with a less favorable exposure we observed the activation of many genes related to stress responses. Another example is given by the clone CVT423, which showed different agronomic and metabolic performances with respect to the other two clones as well as different modulation of several genes. This research was funded by Fondazione Cassa di Risparmio di Cuneo, Project "Nebbiolo Genomics".

Grapevine (Vitis vinifera L.) is one of the major fruit crop worldwide with thousands of cultivated varieties. The economical success of the grape and wine related agribusiness depends on the quantity and quality of the grapes produced, which are heavily influenced by genotype and environment. Among the important Italian varieties, 'Nebbiolo' is a valuable cultivar from Piedmont renowned for its use in producing high quality red wines (Barolo, Barbaresco and others). 'Nebbiolo' is characterized by a wide genetic variability expressed by a high number of clones (i.e. vegetatively propagated lines of selected mother plants) registered in the National Catalogue. However, little morphological differences exist between clones and often it is very difficult to visually distinguish the different clones. Furthermore, an important point is to ensure traceability of the vine supply chain, from grape nurseries up to wine industry, in order to provide a product of sure origin to the consumers. To study clonal diversity, simple sequence repeats (SSRs) have been used as markers, however with low success rates in discriminating clones. Recently, studies reported that allelic variations within the genome of a plant species could be identified by using single nucleotide polymorphisms (SNPs) as high-throughput sequence-based markers. The aim of this on-going study is to identify a set of SNPs able to discriminate the registered 'Nebbiolo' clones, setting up an effective and rapid SNP genotyping protocol based on the TaqMan® probes technology. Three 'Nebbiolo' clones were sequenced by Illumina technology (CVT 71, CVT 185 and CVT 423) and processed reads were mapped against the PN40024 reference genome. SNPs were first detected using the GATK-UnifiedGenotyper tool with default parameters followed by filtering and by a calibration step. Moreover, for the allelic variations identified in coding regions, prediction of changes in the amino acid sequence was performed using SNPeff tool. Then, SNPs were validated as true positive by Sanger sequencing and divided in: i) clone specific SNPs, ii) clones common SNPs, iii) SNPs in protein coding region by affecting genes. To date, Sanger sequencing confirmed 11 SNPs (29%), and nine of these were successfully used to perform TaqMan® genotyping assays, amplifying and detecting specific SNP alleles in genomic DNA from 37 'Nebbiolo' clones. Combining results obtained from each single SNP assay, we classified the 'Nebbiolo' clones in six main groups linked to the geographical origin of accessions. This work is in progress to implement the set of SNPs and to improve 'Nebbiolo' discrimination from other grape cultivars. Furthermore, SNPs validation of allelic variations in coding regions involved in relevant biochemical pathways (i.e., flavonoid biosynthesis) is in progress as well.

Nebbiolo is a valuable Vitis vinifera cultivar from Piedmont, north-west Italy, and it is renowned for its use in producing high quality red wines. Very little information is available so far for this cultivar at molecular genetics or genomics level. In this work we present the assembly and the functional characterization of the genome of three clonal selections of the Nebbiolo variety. Genomic DNA libraries from Nebbiolo Michet (clone 71), Nebbiolo Picotener (clone 423) and Nebbiolo Lampia (clone 185) were deep sequenced using an Illumina HiSeq 1000 sequencer. A genomic coverage of 60x was achieved for all libraries. The three genomes were assembled using a bioinformatic pipeline which combines a reference-guided iterative mapping of the sequenced reads with a de novo assembly. Computational analysis showed differences ranging from 1.78 to 1.91 x 106 single nucleotide polymorphisms (SNPs) between the Nebbiolo clones and the Vitis vinifera PN40024 reference genome. Transposable elements, already found in genomes of other plants as well as of other V. vinifera, cover a fraction of more than 34% of the Nebbiolo genomes. A pool of RNA isolated from Nebbiolo plant organs at different developmental stages was deep sequenced and the obtained reads were used to re-annotate the three genomes. The results of this work will open a path for a better understanding of the Nebbiolo genetic variability and the interaction between environmental factors and genotype, along with the possible identification of molecular markers able to distinguish different 'Nebbiolo' clones. This research was funded by Fondazione Cassa di Risparmio di Cuneo, Project "NEBBIOLO GENOMICS: genomica strutturale e funzionale su aspetti patologici e qualitativi"

Here, for the first time, a comprehensive transcriptomics study is presented of leaf senescence in the legume model Medicago truncatula, providing a broad overview of differentially expressed transcripts involved in this process. The cDNA-amplification fragment length polymorphism (AFLP) technique was used to identify > 500 genes, which were cloned and sorted into functional categories according to their gene ontology annotation. Comparison between the datasets of Arabidopsis and M. truncatula leaf senescence reveals common physiological events but differences in the nitrogen metabolism and in transcriptional regulation. In addition, it was observed that a minority of the genes regulated during leaf senescence were equally involved in other processes leading to programmed cell death, such as nodule senescence and nitric oxide signalling. This study provides a wide transcriptional profile for the comprehension of key events of leaf senescence in M. truncatula and highlights a possible regulative role for MADS box transcription factors in the terminal phases of the process

With approximately 450 species, spiny Solanum species constitute the largest monophyletic group in the Solanaceae family, but a high-quality genome assembly from this group is presently missing. We obtained a chromosome-anchored genome assembly of eggplant (Solanum melongena), containing 34,916 genes, confirming that the diploid gene number in the Solanaceae is around 35,000. Comparative genomic studies with tomato (S. lycopersicum), potato (S. tuberosum) and pepper (Capsicum annuum) highlighted the rapid evolution of miRNA:mRNA regulatory pairs and R-type defense genes in the Solanaceae, and provided a genomic basis for the lack of steroidal glycoalkaloid compounds in the Capsicum genus. Using parsimony methods, we reconstructed the putative chromosomal complements of the key founders of the main Solanaceae clades and the rearrangements that led to the karyotypes of extant species and their ancestors. From 10% to 15% of the genes present in the four genomes were syntenic paralogs (ohnologs) generated by the pre-γ, γ and T paleopolyploidy events, and were enriched in transcription factors. Our data suggest that the basic gene network controlling fruit ripening is conserved in different Solanaceae clades, and that climacteric fruit ripening involves a differential regulation of relatively few components of this network, including CNR and ethylene biosynthetic genes.

Fruits stored at low temperature can exhibit different types of chilling injury. In apple, one of the most serious physiological disorders is superficial scald, which is characterized by discoloration and brown necrotic patches on the fruit exocarp. Although this phenomenon is widely ascribed to the oxidation of α-farnesene, its physiology is not yet fully understood. To elucidate the mechanism of superficial scald development and possible means of prevention, we performed an integrated metabolite screen, including an analysis of volatiles, phenols and lipids, together with a large-scale transcriptome study. We also determined that prevention of superficial scald, through the use of an ethylene action inhibitor, is associated with the triggering of cold acclimation-related processes. Specifically, the inhibition of ethylene perception stimulated the production of antioxidant compounds to scavenge reactive oxygen species, the synthesis of fatty acids to stabilize plastid and vacuole membranes against cold temperature, and the accumulation of the sorbitol, which can act as a cryoprotectant. The pattern of sorbitol accumulation was consistent with the expression profile of a sorbitol 6-phosphate dehydrogenase, MdS6PDH, the overexpression of which in transgenic Arabidopsis thaliana plants confirmed its involvement in the cold acclimation and freezing tolerance.

Pathogen receptor proteins such as receptor-like protein (RLP), receptor-like kinase (RLK), and nucleotide-binding leucine-rich repeat (NLR) play a leading role in plant immunity activation. The genome architecture of such genes has been extensively investigated in several plant species. However, we still know little about their elaborate reorganization that arose during the plant speciation process. Using recently released pepper and eggplant genome sequences, we were able to identify 1097 pathogen recognition genes (PRGs) in the cultivated pepper Zunla-1 and 775 in the eggplant line Nakate-Shinkuro. The retrieved genes were analysed for their tendency to cluster, using different methods to infer the means of grouping. Orthologous relationships among clustering loci were found, and interesting reshuffling within given loci was observed for each analysed species. The information obtained was integrated into a comparative map to highlight the evolutionary dynamics in which the PRG loci were involved. Diversification of 14 selected PRG-rich regions was also explored using a DNA target-enrichment approach. A large number of gene variants were found as well as rearrangements of sequences encoding single protein domain and changes in chromosome gene order among species. Gene duplication and transposition activity have clearly influenced plant genome R-gene architecture and diversification. Our findings contribute to addressing several biological questions concerning the parallel evolution that occurred between genomes of the family Solanaceae. Moreover, the integration of different methods proved a powerful approach to reconstruct the evolutionary history in plant families and to transfer important biology findings among plant genomes.

Botaccini, Francesca et al., Bifidobacteria are known as anaerobic/microaerophilic and fermentative microorganisms, which commonly inhabit the gastrointestinal tract of various animals and insects. Analysis of the 2,167,301 bp genome of Bifidobacterium asteroides PRL2011, a strain isolated from the hindgut of Apis mellifera var. ligustica, commonly known as the honey bee, revealed its predicted capability for respiratory metabolism. Conservation of the latter gene clusters in various B. asteroides strains enforces the notion that respiration is a common metabolic feature of this ancient bifidobacterial species, which has been lost in currently known mammal-derived Bifidobacterium species. In fact, phylogenomic based analyses suggested an ancient origin of B. asteroides and indicates it as an ancestor of the genus Bifidobacterium. Furthermore, the B. asteroides PRL2011 genome encodes various enzymes for coping with toxic products that arise as a result of oxygen-mediated respiration. © 2012 Bottacini et al., This work was financially supported by the Cariparma Bank Foundation to MV. This work was also financially supported by a Federation European Microbiology society (FEMS) Advanced Fellowship 2011 and an Irish Research Council for Science, Engineering & Technology (IRCSET) Embark postdoctoral fellowship to FT. DV is a member of The Alimentary Pharmabiotic Centre and the Alimentary Glycoscience Research Cluster, both funded by Science Foundation Ireland (SFI), through the Irish Government's National Development Plan (grant nos. 07/CE/B1368 and 08/SRC/B1393, respectively).

Turroni, Francesca et al., The human intestine is densely populated by a microbial consortium whose metabolic activities are influenced by, among others, bifidobacteria. However, the genetic basis of adaptation of bifidobacteria to the human gut is poorly understood. Analysis of the 2,214,650-bp genome of Bifidobacterium bifidum PRL2010, a strain isolated from infant stool, revealed a nutrient-acquisition strategy that targets host-derived glycans, such as those present in mucin. Proteome and transcriptome profiling revealed a set of chromosomal loci responsible for mucin metabolism that appear to be under common transcriptional control and with predicted functions that allow degradation of various O-linked glycans in mucin. Conservation of the latter gene clusters in various B. bifidum strains supports the notion that host-derived glycan catabolism is an important colonization factor for B. bifidum with concomitant impact on intestinal microbiota ecology., This work was supported by National Institutes of Health–National Institute of Child Health and Human Development Grant 1R01HD061923 (to D.M.). D.v.S., A.Z., and F.B. are members of the Alimentary Pharmabiotic Centre, a Center for Science and Technology funded by Science Foundation Ireland, through the Irish Government’s National Development Plan Grant 07/CE/B1368.

Background Investigation of tomato genetic resources is a crucial issue for better straight evolution and genetic studies as well as tomato breeding strategies. Traditional Vesuviano and San Marzano varieties grown in Campania region (Southern Italy) are famous for their remarkable fruit quality. Owing to their economic and social importance is crucial to understand the genetic basis of their unique traits. Results Here, we present the draft genome sequences of tomato Vesuviano and San Marzano genome. A 40x genome coverage was obtained from a hybrid Illumina paired-end reads assembling that combines de novo assembly with iterative mapping to the reference S. lycopersicum genome (SL2.40). Insertions, deletions and SNP variants were carefully measured. When assessed on the basis of the reference annotation, 30% of protein-coding genes are predicted to have variants in both varieties. Copy genes number and gene location were assessed by mRNA transcripts mapping, showing a closer relationship of San Marzano with reference genome. Distinctive variations in key genes and transcription/regulation factors related to fruit quality have been revealed for both cultivars. Conclusions The effort performed highlighted varieties relationships and important variants in fruit key processes useful to dissect the path from sequence variant to phenotype.