The effect of CuSO4 for establishing in vitro culture,and the role nitrogen and iron sources in in vitro multiplication of Corylus avellana L. cv. Tonda Gentile Romana

Abstract

In the present work, we have used copper sulphate (CuSO₄·5H₂O) enriched medium for effective control of visible and latent contamination. Among the different concentrations used, 1.25–2.5?mg/L resulted the most appropriate. In addition, the role of different nitrogen source and concentrations (NH₄NO₃ and KNO₃), as different iron source (FeEDTA and FeEDDHA) has been investigated in the proliferation and rooting phases of European hazelnut (cv. Tonda Gentile Romana). The normal concentration of nitrogen present in Murashige and Skoog medium is too high for hazelnut micropropagation cv. Tonda Gentile Romana. A reduction of total nitrogen, accompanied by a reduction of ammonium forms, resulted in a better quality of the shoots. Similar results have been obtained when the common iron source FeEDTA has been replaced by the same concentration of FeEDDHA. An increase in rooting occurs when the amount of nitrogen was reduced in the rooting medium, particularly when the NH₄NO₃ was not present.     

Olive (<Emphasis Type="Italic">Olea europaea</Emphasis> L.) plants transgenic for tobacco osmotin gene are less sensitive to in vitro-induced drought stress

Olive is one of the most important tree crops in the Mediterranean region, because of its ability to grow and produce acceptable yields under limited water availability. In this study, the drought tolerance of an olive cultivar Canino was compared to the performance of its derived transgenic line expressing osmotin gene from tobacco, obtained by Agrobacterium-mediated transformation of Canino cultivar. Shoot cultures of both wild-type (wt) and transgenic lines were exposed to drought stress over a 28-day period, and their differential responses to in vitro-drought stress were investigated. After exposure to PEG, most of the shoots from wt plants resulted in damage and exhibited decreased levels of chlorophyll, while those of transgenic line did not show injuries and showed a normal growth even when exposed to the highest PEG concentration (4%). After preliminary evaluation we characterized Canino AT17-1, by measuring several physiological parameters, including the activities of the antioxidant enzymes (POD and CAT), and the content of malondialdehyde (MDA). Both the activity of catalase and the proline content were higher in the leaves of the transgenic shoots compared to wt plants. Consequently, it was observed that the transgenic line accumulated less MDA indicating that the presence of the osmotin gene protected the cell membrane from damage by lipid peroxidation. Together, these results could suggest that the transgenic line Canino AT17-1 was more efficient in the activation of defense responses against oxidative stress with respect to the Canino wt. The further finding that the transgenic shoots also showed higher proline accumulation supported the hypothesis that the osmotin gene conferred to transgenic shoots increased tolerance to drought stress compared with the wt.     

Alu Exonization Events Reveal Features Required for Precise Recognition of Exons by the Splicing Machinery

Despite decades of research, the question of how the mRNA splicing machinery precisely identifies short exonic islands within the vast intronic oceans remains to a large extent obscure. In this study, we analyzed Alu exonization events, aiming to understand the requirements for correct selection of exons. Comparison of exonizing Alus to their non-exonizing counterparts is informative because Alus in these two groups have retained high sequence similarity but are perceived differently by the splicing machinery. We identified and characterized numerous features used by the splicing machinery to discriminate between Alu exons and their non-exonizing counterparts. Of these, the most novel is secondary structure: Alu exons in general and their 5′ splice sites (5′ss) in particular are characterized by decreased stability of local secondary structures with respect to their non-exonizing counterparts. We detected numerous further differences between Alu exons and their non-exonizing counterparts, among others in terms of exon–intron architecture and strength of splicing signals, enhancers, and silencers. Support vector machine analysis revealed that these features allow a high level of discrimination (AUC=0.91) between exonizing and non-exonizing Alus. Moreover, the computationally derived probabilities of exonization significantly correlated with the biological inclusion level of the Alu exons, and the model could also be extended to general datasets of constitutive and alternative exons. This indicates that the features detected and explored in this study provide the basis not only for precise exon selection but also for the fine-tuned regulation thereof, manifested in cases of alternative splicing.     

In vitro system for studying the interaction between Erwinia amylovora and genotypes of pear

A new in vitro system is described for studying an interaction between Erwinia amylovoraand Pyrus communis (L.). The system uses single shoots placed onto the solid medium, and it enables to detect changes in pH of the medium and differential appearance of shoot necrosis. Shoots of susceptible cultivar (Williams) and tolerant cultivar (Harrow Sweet) were compared measuring the necrosis rate along the in vitroshoots and the pH variation following proton extrusion of both plant and pathogen. Shoots acidified differentially the culture medium depending on the presence of the pathogen, cultivar susceptibility and shoot inoculation methods. Differences in the tolerance level against pathogen among the cultivars were distinguishable only when the shoots were inoculated at the basal end. In susceptible cultivar, the necrosis appeared after 48 h of inoculation, while in tolerant cultivars after 72 h. This system is repeatable and more reliable than already known methods, such as in vitroleaf explants or in vivoplants; it can be used all around the year to test the gene expression and products essential to characterize the genes involved in the pathogenesis. This system showed the effects of E. amylovoraon the photosystem dependent system of host cells, confirmed by the effects of pathogen attack on the variation of chlorophyll a and chlorophyll b ratios and positive effects of light on the appearance of the first disease symptoms.     

Micropropagation of <Emphasis Type="Italic">Teucrium fruticans</Emphasis> L., an ornamental and medicinal plant

An efficient protocol for in vitro propagation of the valuable ornamental and medicinal plant Bush germander (Teucrium fruticans L.) was developed through axillary shoot proliferation. A Murashige and Skoog agar medium supplemented with benzylaminopurine (6.6 μM), α-naphthaleneacetic acid (0.053 μM), and sucrose (3%) significantly improved the production of multiple shoots directly from nodal segment explants, resulting in an average of 2.8 shoots per segment with an average of 6.8 nodes per shoot that would be potential newly formed explants. The new shoots were developed without a marked decrease in the average height of the shoots. Shoots treated with 2.5 μM indole-3-butyric acid showed the highest average root number (7.9) and the highest percentage of rooting (94%). Plantlets were hardened off and transferred to jiffy pots for acclimatization under greenhouse conditions, resulting in a 100% survival rate.