Transposable elements are the most abundant components of plant genomes and can dramatically induce genetic changes and impact genome evolution. In the recently sequenced genome of tomato (Solanum lycopersicum), the estimated fraction of elements corresponding to retrotransposons is nearly 62%. Given that tomato is one of the most important vegetable crop cultivated and consumed worldwide, understanding retrotransposon dynamics can provide insight into its evolution and domestication processes. In this study, we performed a genome-wide in silico search of full-length LTR retroelements in the tomato nuclear genome and annotated 736 full-length Gypsy and Copia retroelements. The dispersion level across the 12 chromosomes, the diversity and tissue-specific expression of those elements were estimated. Phylogenetic analysis based on the retrotranscriptase region revealed the presence of 12 major lineages of LTR retroelements in the tomato genome. We identified 97 families, of which 77 and 20 belong to the superfamilies Copia and Gypsy, respectively. Each retroelement family was characterized according to their element size, relative frequencies and insertion time. These analyses represent a valuable resource for comparative genomics within the Solanaceae, transposon-tagging and for the design of cultivar-specific molecular markers in tomato.

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Interspecific hybridisation in tuber-bearing species of Solanum is a common phenomenon and represents an important source of variability, crucial for adaptation and speciation of potato species. In this regard, the effects of interspecific hybridisation on retrotransposon families present in the genomes, and their consequent effects on generation of genetic variability in wild tuber-bearing Solanum species, are poorly characterised. The aim of this study was to analyse the activity of retrotransposons in inter- and intraspecific hybrids between S. kurtzianum and S. microdontum, obtained by controlled crosses, and the effects on morphological, genetic and epigenetic variability. For genetic and epigenetic analysis, S-SAP (sequence-specific amplification polymorphism) and TMD (transposon methylation display) techniques were used, respectively, with specific primers for Tnt1 and Tto1 retrotransposon families (Order LTR, Superfamily Copia). The results indicate that at morphological level, interspecific hybrid genotypes differ from their parental species, whereas derived intraspecific hybrids do not. In both cases, we observed significant reductions in pollen grain viability, and a negative correlation with Tnt1 mobility. Both retrotransposons, Tto1 and Tnt1, were mobilised in the genotypes analysed, with mobility ranging from 0 to 7.8%. Furthermore, at the epigenetic level, demethylation was detected in the vicinity of Tnt1 and Tto1 in the hybrids compared with the parental genotypes. These patterns were positively correlated with the activity of the retrotransposons. The results suggest a possible mechanism through which hybridisation events generate genetic variability in tuber-bearing species of Solanum through retrotranposon activation.

This is the first report assessing epigenetic variation in garlic. High genetic and epigenetic polymorphism during in vitro culture was detected.Sequencing of MSAP fragments revealed homology with ESTs. Garlic (Allium sativum) is a worldwide crop of economic importance susceptible to viral infections that can cause significant yield losses. Meristem tissue culture is the most employed method to sanitize elite cultivars.Often the virus-free garlic plants obtained are multiplied in vitro (micro propagation). However, it was reported that micro-propagation frequently produces somaclonal variation at the phenotypic level, which is an undesirable trait when breeders are seeking to maintain varietal stability. We employed amplification fragment length polymorphism and methylation sensitive amplified polymorphism (MSAP) methodologies to assess genetic and epigenetic modifications in two culture systems: virus-free plants obtained by meristem culture followed by in vitro multiplication and field culture. Our results suggest that garlic exhibits genetic and epigenetic polymorphism under field growing conditions. However, during in vitro culture system both kinds of polymorphisms intensify indicating that this system induces somaclonal variation. Furthermore, while genetic changes accumulated along the time of in vitro culture, epigenetic polymorphism reached the major variation at 6 months and then stabilize, being demethylation and CG methylation the principal conversions.Cloning and sequencing differentially methylated MSAP fragments allowed us to identify coding and unknown sequences of A. sativum, including sequences belonging to LTR Gypsy retrotransposons. Together, our results highlight that main changes occur in the initial 6 months of micro propagation. For the best of our knowledge, this is the first report on epigenetic assessment in garlic.

Saline, alkaline and mixed saline-alkaline conditions frequently co-occur in soil. In this work, we compared these plant stress sources on the legume Lotus tenuis, regarding their effects on shoot growth and leaf and stem anatomy. In addition, we aimed to gain insight on the plant physiological status of stressed plants. We performed pot experiments with four treatments: control without salt (pH = 5.8; EC = 1.2 dSÁm À1 ) and three stress conditions, saline (100 mM NaCl, pH = 5.8; EC = 11.0 dSÁm À1 ), alkaline (10 mM NaHCO 3 , pH = 8.0, EC = 1.9 dSÁm À1 ) and mixed salt-alkaline (10 mM NaHCO 3 + 100 mM NaCl, pH = 8.0, EC = 11.0 dSÁm À1

The Flooding Pampa (FP) is the most important area for cattle breeding in Argentina. In this region, persistence and yield of typical forage legumes are strongly limited by soil salinity and alkalinity, which affect around 30% of the total area. Instead, naturalized Lotus tenuis is the main forage legume in this region. Rhizobial strains currently used for inoculating L. tenuis in the FP are exotic or native from non-saline soils of this region, their taxonomic identity being unknown. Assuming that rhizobia native from the most restrictive environments are well adapted to adverse conditions, the use of such isolates could improve the productivity of L. tenuis in the FP. Hence, the goal of this study was to evaluate the symbiotic efficiency of selected L. tenuis rhizobia native from the FP, as compared with strains currently used for field inoculation of this legume. Under non-stressing conditions, the symbiotic performance of native strains of FP exceeded those ones currently used for L. tenuis. Moreover, the symbiotic performance of the native strain ML103 was considerably high under salt stress, compared with strains currently used as inoculants. Analysis of 16S rRNA gene sequencing revealed that unclassified rhizobia currently used for field inoculation of L. tenuis and native strains grouped with the genus Mesorhizobium. As a whole, results obtained demonstrate that soils of the FP are a source of efficient and diverse rhizobia that could be used as a sustainable agronomic tool to formulate inoculants that improve forage yield of L. tenuis in this region.

A protocol for the production of transgenic plants was developed for Lotus tenuis via Agrobacteriummediated transformation of leaf segments. The explants were co-cultivated (for 3 days) with an A. tumefaciens strain harbouring either the binary vector pBi RD29A:oat arginine decarboxylase (ADC) or pBi RD29A:glucuronidase (GUS), which carries the neomycin phosphotransferase II (nptII) gene in the T-DNA region. Following co-cultivation, the explants were cultured in Murashige and Skoog medium supplemented with naphthalenacetic acid (NAA) and benzyladenine (BA) and containing kanamycin (30 lg ml -1 ) and cefotaxime (400 lg ml -1 ) for 45 days. The explants were subcultured several times (at 2-week intervals) to maintain the selection pressure during the entire period. About 40% of the explants inoculated with the pBiRD29:ADC strain produced eight to ten adventitious shoots per responsive explant through a direct system of regeneration, whereas 69% of the explants inoculated with the pBi RD29A:GUS strain produced 13-15 adventitious shoots per responsive explant. The selected transgenic lines were identified by PCR and Southern blot analysis. Three ADC transgenic lines were obtained from 30 infected explants, whereas 29 GUS transgenic lines were obtained from 160 explants, corresponding to a transformation efficiency of 10 and 18.1%, respectively.More than 90% of the in vitro plantlets were successfully transferred to the soil. The increase in the activity of arginine decarboxylase from stressed ADC-Lt19 lines was accompanied by a significant rise in the putrescine level. The GUS transgenic line driven by the RD29A promoter showed strong signals of osmotic stress in the leaves and stem tissues. All of the transgenic plants obtained exhibited the same phenotype as the untransformed controls under non-stress conditions, and the stability of the gene introduced into the cloned materials was established.

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