86 research outputs found
Tomato ionomic approach for food fortification and safety.
Food fortification is an issue of paramount of importance for people living both in developed
and in developing countries. Among substances listed as "nutriceuticals", essential minerals have
been recognised for their involvement in several healthy issues, involving all ages. In this frame,
food plants are playing a pivotal role since their capability to compartmentalise ions and proteinmetal
complexes in edible organs. Conversely, the accumulation of high metal levels in those
organs may lead to safety problems. In the recent years, thanks to the availability of new and
improved analytical apparatus in both ionic and genomic/transcrittomics areas, it is became feasible
to couple data coming from plant physiology and genetics. Ionomics is the discipline that studies
the cross-analysis of both data sets. Our group, in the frame of GenoPom project granted by MiUR,
is interested to study the ionomics of tomatoes cultivars derived by breeding programmes in which
wild relatives have been used to transfer several useful traits, such as resistance to biotic or abiotic
stresses, fruit composition and textiture, etc. The introgression of the wild genome into the
cultivated one produces new gene combinations. They might lead to the expression of some traits,
such as increased or reduced adsorption of some metals and their exclusion or loading into edible
organs, thus strongly involving the nutritional food value. Our final goal is to put together data
coming from ions homeostasis and gene expression analyses, thus obtaining an ionomic tomato
map related to ions absorption, translocation and accumulation in various plant organs, fruits
included. To follow our hypothesis, we are studying the ionome of Solanum lycopersicum cv. M82
along with 76 Introgression Lines (ILs) produced by interspecific crosses between this cultivar and
the wild species S. pennellii. These ILs are homozygous for small portions of the wild species
genome introgressed into the domesticated M82 one. They are used as a useful tool for mapping
QTL associated with many traits of interest. It is worthy to note that, until now, little information is
available on QTL for ions accumulation in tomato. Moreover, as our knowledge, effects of new
gene combinations in introgressed lines on ions uptake related to food safety have not been
extensively studied. In this presentation we show results coming from the ionome analysis, carried
out on S . lycopersicum M82 and several ILs. Plants were grown in pots in a greenhouse and
watered with deionised water Thirty day-old plants were left to grow for 15 days in the presence of
non-toxic concentration of Cd, Pb, As, Cr and Zn given combined. Leaves of all plants were then
harvested and stored at -80°C for ionome and gene expression analyses. Preliminary results of
ionome analysis of S. lycopersicum M82 and several ILs, carried out using an ICP-MS, showed that
traits correlated to toxic metals and micronutrients accumulation in apical leaves were significantly
modified in response to specific genetic backgrounds. Those results are perhaps due to the
introgression of traits linked to uptake, translocation and accumulation of useful and/or toxic metal
into plant apical leaves and to interactions of the wild type introgressed genomic regions with the
cultivated genome. Also, data are shown on the identification and isolation of Solanum gene
sequences related to ions uptake, translocation and accumulation, useful for further real-time gene
expression evaluation in both cultivated and ILs during the treatments with the above-mentioned
metals
Validation of an Eco-Friendly Automated Method for the Determination of Glucose and Fructose in Wines
Fermentable sugar dosage helps oenologists to establish a harvest’s moment and control
the fermentation process of the musts. The official analyses recommended for their determination
are long, laborious, and must be carried out by specialized personnel. On the contrary, instrumental
analysis automation limits human errors, increases precision, and reduces the time and cost of
the analyses. In the food production sector, to use methods other than those recommended by
supranational bodies in official reports, it is necessary to validate the analytical processes to establish
the conformity of the results between the new methods and the reference ones. This work validated
an automated enzymatic apparatus to determine the sum of glucose and fructose levels in wine
samples. The validation was carried out on wine samples (dry red wine, dry white wine, moderately
sweet wine, and sweet wine) containing different sugar concentrations by comparing data obtained
using the OIV-MA-AS311-02 method performed by a specialized operator (reference method) and
the same method performed by an automated apparatus. The difference between the results’ means
obtained with the two procedures was significant. Nevertheless, the automated procedure was
considered suitable for the intended use since the differences between the averages were lower than
the measurement uncertainty at the same concentration, and the repeatability results were better for
the automated procedure than the reference method
SOLANUM LYCOPERSICUM X S. PENNELLII INTROGRESSION LINES WERE USEFUL TO CHARACTERISE THE IONOME OF TOMATO FRUIT
In the frame of "GenoPom" PON-MIUR project, we have
began a study of tomato ionome in order to identify the contribution of specific chromosome and
part of them on the ionome. For this purpose, we have analysed by ICP-MS plants of an
introgression line (IL) population derived from the cross between Solanum lycopersicum cv. M82
and S. pennellii (Eshed and Zamir, 1995) grown under controlled environmental conditions.we report results derived from tomato whole fruits ionome analysis of 30 IL,
covering all 12 tomato chromosomes, along with the recurrent parent cv. M82. Among several
detected elements, the following ones have firstly been analysed: Ca, Fe, Cu, Zn and Se. Each
element concentration data were referred to cv. M82
Ion uptake and YSL1 gene identification in tomato
Tomato breeder are using wild tomato relatives, even non-cross compatibles ones, in order to
obtain cultivars with highly commercial values bearing new traits. However, the introgression of a
wild genome into the cultivated one produces a new gene combinations that may lead to the
expression of undeliverable traits, perhaps not so easy to recognise; even more, phenotypic
variations may escape during the selection procedure when minor genes or non-abnormal
phenotypes are involved. In the frame of the “GenoPom” project funded by MIUR, we have
focused our interest on the alteration of heavy metals uptake from the soil and their loading into
edible organs in commercial lines coming from Solanum interspecific crosses. Our final aim is to
put together data coming from ion homeostasis and gene expression analyses, thus obtaining a
ionomic map of tomato. To pursue our goal, we have started to study the cv M82 of Solanum
lycopersicon, the wild relative Solanum pennelli and their introgression lines IL. Regarding the
experiments on ion homeostasis, S. lycopersicon M82 and the introgression line IL 6-4-2 were
grown in hydroponics under controlled environmental conditions. Twenty day-old plants were left
to grow for 10 days in the presence of non-toxic concentration of Cd (10 mM), Pb (3 mM), Zn (100
mM) given separately or combined. Control and treated roots and leaves were then harvested and
stored at -80°C for ionic and gene expression analyses. Ions analysis of Solanum lycopersicon M82
and IL 6-4-2 showed that traits correlated to ionic homeostasis is significantly modified in response
to all metals and to the genotype. The analysis of ions data, obtained by ICP-MS, give a pictures of
the different responses performed both to different stress and to combined stress, probably
correlated to the up-regulation and/or down regulation of metal uptake proteins. Performed
experiments demonstrate that the introgression of the wild genome into the cultivated one produces
a new phenotype, perhaps due to the expression of traits linked to uptake, translocation and
accumulation of useful and/or toxic metal into plant tissues and organs.
Regarding the functional genomics approach for gaining insight into gene networks involved
in mineral-ion accumulation in tomato plants, in literature has been reported that at least 25 major
family genes are involved for metal homeostasis in plants. Among them, the genes ysl, hma, mtp,
znt, zrt have been already studied at least in the plant species Arabidopsis thaliana, A. halleri and
Thlaspi caerulescens. So far, no such genes have been reported to be cloned in Solanum species.
We have focused our study on the genes YSL1, ZNT1 and MTP1 responsible for uptake,
translocation and accumulation of metal such as zinc, cadmium, and iron into plant compartment.
For all of them, consensous sequences from nucleotide multialignment have been obtained. Then,
each of those were blasted in a Solanum EST collection databank and an assembled UniGene
sequence was obtained.. Finally, we have designed primers and performed PCR analysis on S.
lycopersicon and S. pennelli genomic DNA. So far, we have cloned a putative ysl1 sequence from
tomato, that has shown that a very high percentage of identity (92%) with whole ysl1 gene of
Nicotiana tabacum; the in silico translated sequence of this sequence has shown a 89% of identity
with the same tobacco protein
Ionome variations in tomato Introgressed Lines (Solanum Pennellii x S. Lycopersicum cv. M82) following metal treatements shed new light on food health.
A tomato introgression line population that combines single chromosomal segments introgressed from the wild, green fruited species Solanum pennelli in the background of the domesticated tomato, S. lycopersicum cv. M82, was used in this study. Results shed light both on the metal accumulation of ILs
tomato plants and on theirs ionome modifications
Wastewater phytoremediation: genomic analysis and screening of green microalgae species for extracellular laccase activity.
Phytoremediation deals with the use of plants, or other green photosynthetic organisms, to
reduce organic or inorganic pollutant in the environment, mainly waters and soils (Pivetz. B.E.
2001. EPA
/540/S-01/500). Our research group is involved in studies regarding the use of plants for soil
phytoremediation (Galante et al., 2005. Proc. SIGA Congress, L04) and, more recently, of
unicellular green algae species (green microalgae) for wastewater treatments. Some microalgae
species have been recently tested to degrade an array of pollutants such as phenols, polyphenolic
aromatic compounds (PAH) and even hormones (Pollio et.al., 1994. Phytochemistry, 37:1269-
1272;. Pinto et.al., 2003. Biotechnol Lett., 25:1657-1659). It is worthy to note that about 2,500
species belong from Chlorophyceae, seldom living in contrasting habitat under severe
environmental conditions. So far, a little has been done to exploit this genetic biodiversity bonanza;
thus, few reports have been published on enzymes implicated in their degradative action (Semple
et.al., 1996. Appl. Envr. Micr, 62:1265-1273).
Since a wide collection of green microalgae species are available at the University of Naples,
Department of Biological Science, recently we have started a research aimed to (a) find algae
species with extracellular phenoloxidase enzymatic activity; (b) identify extracellular enzymes able
to degrade xenobiotic like synthetic dyes and other PAHs; (c) clone and overexpress genes
producing phenoloxidases in homologous and in heterologous systems, in order to use these
enzymes primarily for phytoremediation of milling oil wastewaters. Among phenoloxidaes, we
focused our interest on laccases (EC 1.10.3.2) that are phenol-oxidoreductases able to catalyze the
oxidation of various aromatic compounds (particularly phenols) with the concomitant reduction of
oxygen to water.
Selected algae strains were grown in liquid culture at 22°C under continuous light conditions,
starting with an inoculum of 0.1 OD. After ten days, the algal growth was measured as optical
density at 600 nm. A screening was performed by detecting the laccase activity in the broth medium
culture, deprived of algae cells, in the presence of 2,2-azino-bis 3-ethybenz-thiazoline-6-sulfonic
acid (ABTS) at 420 nm. The laccase activity was referred to the polyphenol oxidase activity of
Trametes versicolor; thus, each positive strain was assayed on industrial azo-dye Remazol Brilliant
Blue R (RBBR) and on the natural phenol compound syringaldazine by kinetic analysis.
Preliminary results, obtained comparing different species, showed a wide variation both within the
same substrate and among the different microalgae.
Microalgae strains able to produce and secrete laccase enzymes were further chosen for more
detailed genetic studies. To clone phenoloxidase genes from those species, we have started a bioinformatics
approach, on the basis of highly conserved coding sequences of laccases already
isolated and sequenced from several higher plants. Primers drawn on the alignment of those
sequences have been used to amplify genomic DNA
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