43 research outputs found
A systems biology approach to shed light on apple fruit development
No full textThe research carried out and discussed in the present dissertation is positioned within the “TranscrApple” project (www.transcrapple.com), funded by the Provincia Autonoma di Trento (PAT) within the call “Grandi Progetti 2012”. The general objectives of this project, a significant part of which overlap with the present thesis, deal with the characterization, as wide as possible with the currently available technologies, of the transcriptional events, including those related to small RNAs (not discussed in the present dissertation), the metabolic changes, on a subset of primary and secondary metabolites, and hormones’ cross-talk, through a hormone profiling approach, occurring during apple fruit development.
The present thesis is organized in different chapters, mirroring the experimental and temporal rationale effectively pursued to develop the research herein described. The main objective of the present work deals not only with providing an overview of transcripts, metabolites and hormones and their variations during fruit development, but also with the setting up of technical and experimental solutions aimed at using the achieved information within an integrative platform, according to a “systems biology view”. In model species, all this kind of studies are extremely easier, thanks to the availability of ready-to-use bioinformatics tools that are not flexible enough to be used in other species. However, especially in tree crops, this approach is still far from being defined and standardized.
Chapter 1 introduces the theme “apple fruit development”, discussing the adoption of apple as a model system that, in the last decade, acquired great importance in terms of research among the fruit species thanks to the availability of its sequenced genome. After a brief introduction on the fundamental information available about the apple fruit growth, taking into account the technological and scientific points of view, few difficulties and gaps that hamper the achievement of a complete overview of the regulatory events coordinating the development and growth of the apple, are discussed in relation to the main quantitative and qualitative parameters characterizing the apple fruit production.
Chapter 2 enters into the apple fruit research area; the preliminary phase and the multiple validations (concerning different cultivars and seasons) of transcriptional markers during the main apple developmental stages were shown to be fundamental for choosing, on the base of the expression profiles of these genes, the most representative samples, among those collected also in different seasons. Several markers have been identified, validated and
employed, among those available from literature, allowing the selection of samples of cv Golden Delicious (herein considered as model) to be used for the subsequent transcriptional and metabolomic characterization carried out in the present research.
Chapter 3 deals with the hormonal profiling survey carried out along the apple fruit development for the first time in this species. The results have allowed not only the achievement of brand new data related to the major hormonal classes, to be employed for further researches, but also the clarification and/or confirmation of new hormonal interactions connected to the fruit development stage or the transition between stages. Moreover, the relevance of this study consists in having achieved, for the first time in apple, quantitative data of an important set of hormones concurrently on the same samples.\ud
Chapter 4 concerns the survey on the metabolites and their variations during the apple fruit development. A complete overview of the changes of the different classes of metabolites (mainly sugars, organic acids, aminoacids and polyphenols) is given during the apple developmental cycle. The acquired data have been derived from the same samples already analyzed in the previous chapters, and will be integrated with data of diverse nature, such as the RNAseq.
Chapter 5 of the present thesis comes into the “system biology” area, initially among several technical difficulties, then partially solved, and gives an example of an alternative interpretation of the hormonal data put within a correlative network along with the RNAseq results achieved on the same samples
Thermographic imaging to identify abscising apple fruitlets after a thinning treatment
No full textMetamitron is a widely used active ingredient in pome fruit thinning. Its inhibitory action on photosynthesis determines a nutritional stress that in turn stimulates the weakest fruitlets to abscise. However, a successful thinning treatment with metamitron, commercially known as Brevis®, relies upon several interacting factors, both endogenous and exogenous, and its effects are difficult to predict under certain specific conditions. For this reason, new predictors are desirable to assist the growers in applying the treatment at optimal time and dosage. Moreover, the physiology of metamitron action is in part still unknown, especially at the fruitlet level. In the present research, a thermographic assessment was carried out to verify the hypothesis that high abscisic acid levels together with photosynthesis block in the fruitlet tissues cause the temperature to raise to higher levels in treated fruitlets than in the untreated ones. A specific trial was carried out in ‘Golden Delicious’, pointing out that both leaves and fruitlets of thinned trees show higher temperatures, which can be used to identify abscising fruitlets. A working model is also proposed that summarizes the way of action of Brevis® on both the considered organ
Profili trascrizionali dei geni coinvolti nella biosintesi e trasduzione del segnale etilenico durante l’abscissione dei frutticini in melo
No full textL’abscissione dei frutticini di melo è preceduta da una stimolazione della biosintesi dell’etilene e un aumento della sensibilità della zona di abscissione (AZ) agli stimoli promossi dall’ormone. Recentemente, è stato ipotizzato il ruolo dell’etilene come “carrier” del segnale abscissione che, generato nella cortex mediante l’instaurazione di un cross-talk con ABA, zuccheri e ROS, viene trasferito al seme nelle fasi che precedono l’attivazione dell’AZ. Rimangono tuttavia ancora molti aspetti da chiarire. A tal proposito, è stata condotta un’approfondita indagine a livello trascrizionale dei geni coinvolti nella biosintesi e trasduzione del segnale etilenico in cortex e seme di frutticini di melo trattati con benziladenina (BA). Sono stati identificati nella sequenza genomica 80 geni codificanti elementi della biosintesi (ACO e ACS), percezione (ETR, ERS, TPR e RTE) e trasduzione del segnale (CTR, EIN2 e EIN3/EIL), regolatori coinvolti nell’ubiquitinizzazione e degradazione mediante proteasoma (EBF e ETP) e altri elementi noti (EIN5 e RAN1). Inoltre, 13 geni codificanti chinasi (MAPK e MAPKK), coinvolte nella presunta via trasduttiva del segnale coordinata da CTR, sono stati presi in esame in questo lavoro. Un elevato livello di ridondanza è stato riscontrato per la maggior parte dei geni, probabilmente dovuto all’origine del genoma del melo e al recente evento di duplicazione a cui è andato incontro. In seguito, sono stati analizzati i profili trascrizionali dei geni identificati in cortex e seme di frutticini caratterizzati da diversi potenziali di abscissione. I risultati preliminari hanno evidenziato una notevole tessuto-specificità di espressione dei geni analizzati e differenze trascrizionali che potrebbero coordinare l’attivazione del segnale primario specifico dell’abscissione. Questo studio è finanziato dal Progetto AGER, grant n° 2010-2119
GENDER SPECIFIC VARIATION OF TWO PHENOLIC GLYCOSIDES (POPULIN AND SALICIN) IN POPULUS CILIATA AND IDENTIFICATION OF A NEW COMPOUND (CINNAMOYL-SALICIN)
No full textObjective: To observe gender specific seasonal variation of two phenolic glycosides (PG\u27s) (populin and salicin) in Populus ciliata male and female trees.Methods: Plant material (bark) was collected from male and female trees throughout the year. The content of salicin and populin was measured using HPLC. Because of the lack of populin in the market, the standard compound was synthesized in the laboratory from salicin following standard procedure. Confirmation and characterization of synthesized populin were done using 1H NMR and 13C NMR. TLC and LC-MS of methanolic extract were performed to observe the presence of populin and salicin in the plant bark.Results: TLC showed the presence of populin and salicin in crude plant extract at Rf value 0.84 and 0.52, respectively. The results of monthly variation showed a consistent pattern of two PG\u27s for both the sexes. However, salicin content was observed highest compared to populin content. Whereas female trees were observed with low content of PG\u27s compared to male trees. The maximum content of salicin and populin was observed in the flowering season. LC-MS of bark methanolic extract confirmed the presence of a significantly larger peak, which was identified as a cinnamoyl-salicin peak at 463.Conclusion: The study confirms the highest content of PG\u27s in the flowering season. Additionally, LC-MS study concluded a new compound cinnamoyl-salicin (M+at 463) which is reported first time to the best of author\u27s knowledge. It seems that it could be the denaturation product of 2\u27-O-cinnamoyl salicortin and can be further explored for biological applications
Apple (Malus domesticaL. Borkh) as an emerging model for fruit development
No full textApple research has undergone great improvements in the last years, in both quantitative and qualitative terms. Huge amount of data are now available, especially as far as the early development and the ripening phase are concerned. Moreover, the recent release of the apple genome sequence is significantly speeding up research, allowing on one hand to shed light on the most critical aspects of fruit development with almost immediate practical implications and, on the other hand, to identify new molecular markers that will improve the future breeding programs. In this context, apple is being increasingly considered as a model for fruit development studies, although many gaps still exist in apple research. These gaps are being filled by coupling the next generation high-throughput technologies with new physiological approaches, aimed at achieving both new basic knowledge and innovative tools to improve the final quality of the fruit. In this review, the available information on the regulatory aspects of apple fruit development will be reported and discussed in the light of the future perspectives of apple research
Identification of culturable bacterial endophyte community isolated from tissues of Vitis vinifera“Glera”
No full textEndophytes are micro-organisms that colonize the internal tissues of plants without inducing signs of negative effects and that can provide benefits to plant health and yield. In the present work, the culturable bacterial endophyte community, colonizing vegetative organs of grapevine, was isolated from surface-sterilized plant tissues and characterized by molecular methods. From roots, shoots and leaves of Vitis vinifera “Glera”, located in six different vineyards throughout the Conegliano-Valdobbiadene DOCG area (Veneto, Italy), 381 culturable strains were successfully isolated; amplified ribosomal DNA restriction analysis and nucleotide sequencing showed that approximately 30% of the endophyte community belonged to the genus Bacillus, which was the most represented; other genera such as Staphylococcus, Microbacterium, Paenibacillus,
Curtobacterium, Stenotrophomonas, Variovorax, Micrococcus and Agrococcus were identified. Endophyte community composition within each vine was different in respect to other endophyte populations living in grapevine plants coming from different vineyards; moreover, the bacterial composition changed depending on the season of sampling. The above data highlight the great diversity of culturable bacterial species inhabiting Glera grapevines and open the way for a characterization and selection of strains that could potentially be used to improve the vineyard management for plant growth and yield, plant responses to stresses, biocontrol and biofertilization
Isoenzymes of the flavonoid and phenylpropanoid pathways show organ-specific regulation during apple fruit development
No full textElucidating the molecular mechanisms controlling fruit development is a primary target for the improvement of new apple (Malus × domestica Borkh.) cultivars. The first two weeks of development following pollination are crucial to determine fruit characteristics. During this period, a lot of changes take place in apple fruit, going from rapid cell division to the production of important metabolites. In this work, attention was focused on the phenylpropanoid and flavonoid pathways responsible for the production of numerous compounds contributing to fruit quality, such as flavonols, catechins, dihydrochalcones and anthocyanins. A total of 17 isoenzymes were identified, belonging to seven classes of the phenylpropanoid and flavonoid pathways that, despite showing more than 80% sequence identity, showed differential expression regulation during the first two weeks of apple fruit development. This feature seems to be quite common for most of the enzymes of both pathways. Differential regulation of isoenzymes was shown to be present in both 'Golden Delicious' and a wild relative (Malus mandshurica), even though differences were also present. Each isoenzyme showed a specific pattern of expression in the flower and fruit organs, suggesting that genes coding for enzymes with the same function may control different aspects of plant biology. Finally, promoter analysis was performed in order to highlight differences in the number and type of regulatory motifs. Overall, our results indicate that the control of the expression of genes involved in the phenylpropanoid and flavonoid pathways may be very complex as not only enzymes belonging to the same class, but even putative isoenzymes, can have different roles for the plant. Such genes may represent an important regulatory mechanism, as they would allow the plant to fine-tune the processing of metabolic intermediates towards different branches of the pathway, for example, in an organ-specific way
Alla ricerca della mela ipoallergenica: strategie e prospettive future
No full textLo studio delle allergie sta assumendo sempre maggiore importanza nell’ambito della ricerca agroalimentare. La mela è uno dei frutti che è più frequentemente coinvolto in reazioni allergiche. Per questo motivo, numerosi progetti nazionali e internazionali (es. progetti europei SAFE e ISAFRUIT), sono stati dedicati allo studio delle allergie a questo frutto, chiarendone alcuni aspetti sia dal punto di vista clinico-allergologico, sia dal punto di vista biochimico-molecolare. Tali progetti hanno contribuito alla formazione di una prima rete di ricerca nazionale e internazionale su questo importante argomento. Nell’ambito di queste iniziative, si colloca il progetto AGER-Melo che sta affrontando alcuni aspetti critici dell’allergia alla mela, nell’ottica del miglioramento della qualità e della salubrità del frutto, coinvolgendo competenze mediche, biochimiche, genetico-molecolari e agro-biotecnologiche. Nel melo sono state identificate 4 famiglie di principali allergeni: Mal d 1, 2, 3 e 4, ognuna composta da numerose isoforme che aumentano la complessità delle ricerche. Nell’AGER-Melo sono state previste diverse linee di ricerca allo scopo di valutare il potenziale allergenico della mela e i fattori endogeni ed esogeni che lo determinano. Tali valutazioni sono attualmente in corso mediante l’impiego di diverse tecniche quali l’utilizzo di sieri di pazienti allergici, di anticorpi policlonali specifici per ogni famiglia di allergeni, di analisi della espressione di ogni singolo allergene e dell’intero trascrittoma dei genotipi più promettenti. L’identificazione dei determinanti dell’allergenicità della mela permetterà di ottenere varietà ipoallergeniche anche mediante la progettazione di programmi di miglioramento genetico dedicati a questo scopo con l’obiettivo finale di consentire anche a persone allergiche di consumare questo frutto particolarmente noto anche per il suo valore nutrizionale. Questo studio è finanziato dal Progetto AGER-Melo, grant n° 2010-2119
A multifaced approach sheds light on the molecular details underlaying the mechanism preventing enzymatic browning in ‘Majda’ apple cultivar (Malus domestica Borkh.)
No full textFruit flesh browning is a natural oxidative phenomenon occurring after cutting or processing. This event, manifesting dark-brown coloration, can seriously limit the production and marketability of fresh cut fruit products. Antioxidant compounds can partially prevent this phenomenon, but in turn they can affect the quality and the organoleptic properties of fresh products. In this work we have investigated, through a multidisciplinary approach, the regulation of a natural non-browning phenotype of a particular apple cultivar ‘Majda’ in comparison with the browning reference apple cultivar ‘Golden Delicious’. The results highlight that the non- browning phenotype of ‘Majda’ was controlled by multiple mechanisms, particularly distinguished by a different concentration of chlorogenic acid and the expression of the polyphenol oxidase (MdPPO) gene. The metabolite assessment and gene expression profiling also disclosed the involvement of organic acids and glutathione in the prevention of oxidative browning phenomenon. The peculiar non-browning behavior of ‘Majda’ was also verified by a DPPH•kinetic approach that demonstrated the higher antioxidant activity of this apple cultivar. The results presented in this work proposed ‘Majda’ as a potential parental candidate to be considered for breeding activity oriented to select novel non-browning accessions addressed to ameliorate the sustainability of pre-cut fresh products
