Browsing by Author "Muleo, Rosario"

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    Ethylene inhibitor silver nitrate enhances regeneration and genetic transformation of Prunus avium (L.) cv Stella
    (Springer Netherlands, 2014-08-22) Sgamma, Tiziana; Thomas, Brian; Muleo, Rosario
    The effects of the ethylene inhibitors, silver nitrate (AgNO3) and aminoethoxyvinylglycine (AVG), together with Agrobacterium co-cultivation were investigated for improving both regeneration and genetic transformation of leaf explants of the recalcitrant sweet cherry cultivar Stella. Co-cultivated young leaf explants with A. tumefaciens strain EHA105 harbouring a binary vector pROK containing both the phytochrome B of Arabidopsis thaliana and the nptII genes, were cultured on M2 MS-modified medium supplemented with either AVG (2.54 µM) or AgNO3 (296.08 µM) for 21 days in the dark, and then transferred onto ethylene-free inhibitors medium in the light. Efficiency, in terms of frequency on shoot organogenesis, was observed in explants subjected to co-cultivation with Agrobacterium and not co-cultivated in presence of each ethylene inhibitors. Ethylene inhibitors helped in the morphogenesis process and mass production of shoots. An increase in regeneration efficiency was observed in ethylene-inhibitor-free medium, which was almost four times greater than the control leaf explants. When ethylene inhibitors were added to the medium, a further regeneration efficiency enhancement for AVG (45%) and AgNO3 (63.3%) was observed. Whereas, in explants not co-cultivated with Agrobacterium, the resulting regeneration efficiency was 25% for AVG and 23.3%, for AgNO3. The regeneration frequency from leaf explants was also enhanced in treatments with AVG when combined with Agrobacterium co-cultivation, with 139 shoots regenerated from a total of 27 regenerating explants. The AgNO3 also contributed to control Agrobacterium growth in the post-co-cultivation phase. Only the lines originating from AgNO3-treated explants showed kanamycin resistance, 9 out of 141 regenerated lines, and polymerase chain reaction analysis confirmed both the insertion of the genes into the plant genome and the absence of bacterial contamination of the transgenic shoots.
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    In vitro plant culture system induces phase transition in fruit-bearing plants
    (Acta Horticulturae, 2016-02-25) Sgamma, Tiziana; Cirilli, M.; Caboni, E.; Maurizio, M.; Thomas, B.; Muleo, Rosario
    The juvenile to adult switch is the most important post-embryonic transition. In woody plants the juvenile phase can last many years with a great economic impact. In Arabidopsis, the small RNAs miR156 and miR172 play a crucial but opposite role in the regulation of this process. miR156 maintains juvenility, negatively regulating SPLs genes, while miR172 promotes adult transition, targeting the floral repressors AP2-like transcription factors. In this work, peach (Prunus persica L. Batch) orthologs of Arabidopsis epigenetic and genetic factors involved in the juvenility to adult phase transition were studied. In peach, higher levels of ppa-miR156 were detected in seedlings, in vitro and extra vitro plants than in adult plants. Also, PpSPLs were more expressed in adult plants, confirming a possible role for the miR156-SPL pathway in promoting juvenile-like characteristics. ppa-miR172 expression level was low in seedlings and in vitro plants but an increase was observed in the adult donor plant, corresponding to lower expression of PpAP2-like genes. In Arabidopsis, flower induction is also promoted by activation of the FLOWERING LOCUS T (FT) gene. In peach leaf tissue, low levels of PpFT-like expression in rejuvenated plants and seedlings were detected. We propose that, in peach, conserved key genes present in herbaceous plants and woody species are involved in juvenile to adult and adult to juvenile-like phase transitions.
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    Monochromic Radiations Provided by Light Emitted Diode (LED) Modulate Infection and Defense Response to Fire Blight in Pear Trees
    (MDPI, 2021-09-12) Sgamma, Tiziana; Forgione, Ivano; Luziatelli, Francesca; Iacona, Calogero; Mancinelli, Roberto; Thomas, Brian; Ruzzi, Maurizio; Muleo, Rosario
    Pathogenesis-related (PR) proteins are part of the systemic signaling network that perceives pathogens and activates defenses in the plant. Eukaryotic and bacterial species have a 24-h ‘body clock’ known as the circadian rhythm. This rhythm regulates an organism’s life, modulating the activity of the phytochromes (phys) and cryptochromes (crys) and the accumulation of the corresponding mRNAs, which results in the synchronization of the internal clock and works as zeitgeber molecules. Salicylic acid accumulation is also under light control and upregulates the PR genes expression, increasing plants’ resistance to pathogens. Erwinia amylovora causes fire blight disease in pear trees. In this work, four bacterial transcripts (erw1-4), expressed in asymptomatic E. amylovora-infected pear plantlets, were isolated. The research aimed to understand how the circadian clock, light quality, and related photoreceptors regulate PR and erw genes expression using transgenic pear lines overexpressing PHYB and CRY1 as a model system. Plantlets were exposed to different circadian conditions, and continuous monochromic radiations (Blue, Red, and Far-Red) were provided by light-emitting diodes (LED). Results showed a circadian oscillation of PR10 gene expression, while PR1 was expressed without clear evidence of circadian regulation. Bacterial growth was regulated by monochromatic light: the growth of bacteria exposed to Far-Red did not differ from that detected in darkness; instead, it was mildly stimulated under Red, while it was significantly inhibited under Blue. In this regulatory framework, the active form of phytochrome enhances the expression of PR1 five to 15 fold. An ultradian rhythm was observed fitting the zeitgeber role played by CRY1. These results also highlight a regulating role of photoreceptors on the expression of PRs genes in non-infected and infected plantlets, which influenced the expression of erw genes. Data are discussed concerning the regulatory role of photoreceptors during photoperiod and pathogen attacks.