Polyamine patterns in haploid and diploid tobacco tissues and in vitro cultures

The aim of this work was to determine PAs levels in pith tissues and callus cultures from haploid and diploid tobacco plants, explanted from the apical and basal regions of the stem. These explants were cultured in an RM-64 medium supplied with IAA and kinetin, under light or in the dark, during successive subcultures. PAs levels followed a basipetal decrease in diploid and an increase in haploid, pith tissues. A similar pattern of total PAs (free + conjugated) was observed for the callus of diploid and haploid plants maintained in the light, and for the haploid callus in the dark, whereas the diploid callus in the dark showed a constant increase in total PAs levels until the end of culture. The PA increase in the diploid callus in the dark was related to free Put levels increase. The ploidy status of the plants could express different PA gradients together with the plant pith and in vitro callus cultures.O objetivo deste trabalho foi determinar os níveis de PAs em tecidos de medula e cultura de calos de plantas haplóides e diplóides de tabaco, obtidas da região apical e basal do caule. Estes explantes foram cultivados em meio RM-64 suplementado com AIA e cinetina, na luz e no escuro, durante vários subcultivos. Nos tecidos medulares, os níveis de PAs apresentam um decréscimo basípeto em diplóides e um aumento em haplóides.Um padrão similar nos níveis de PAs totais (livres+ conjugadas) foi observado em calos haplóides e diplóides mantidos na luz, e haplóides no escuro, enquanto os diplóides cultivados no escuro mostraram um aumento constante até o final do cultivo. O aumento no conteúdo de PAs nos calos diplóides no escuro, foi devido ao aumento do conteúdo de Put livre. Foi observado que a ploidia da planta pode expressar diferentes gradientes de PA ao longo do tecido medular e nas culturas de calos in vitro.(FAPESP) São Paulo Research Foundation(CNPq) National Council for Scientific and Technological Developmen

Cloning and expression of embryogenesis-regulating genes in Araucaria angustifolia (Bert.) O. Kuntze (Brazilian Pine)

Angiosperm and gymnosperm plants evolved from a common ancestor about 300 million years ago. Apart from morphological and structural differences in embryogenesis and seed origin, a set of embryogenesis-regulating genes and the molecular mechanisms involved in embryo development seem to have been conserved alike in both taxa. Few studies have covered molecular aspects of embryogenesis in the Brazilian pine, the only economically important native conifer in Brazil. Thus eight embryogenesis-regulating genes, viz., ARGONAUTE 1, CUP-SHAPED COTYLEDON 1, WUSCHEL-related WOX, S-LOCUS LECTIN PROTEIN KINASE, SCARECROW-like, VICILIN 7S, LEAFY COTYLEDON 1, and REVERSIBLE GLYCOSYLATED POLYPEPTIDE 1, were analyzed through semi-quantitative RT-PCR during embryo development and germination. All the eight were found to be differentially expressed in the various developmental stages of zygotic embryos, seeds and seedling tissues. To our knowledge, this is the first report on embryogenesis-regulating gene expression in members of the Araucariaceae family, as well as in plants with recalcitrant seeds

Producción de metabolitos secundarios en cultivo de raíces in vitro y suspensiones celulares de Ipomoea carnea spp. carnea Jacq.

Ipomoea carnea spp. carnea Jacq. es un arbusto dominante en el bosque estacionalmente seco del Perú y Ecuador. Los cultivos de raíces fueron establecidos en medio MS con diferentes concentra- ciones de AIB, sacarosa y combinaciones de AIB-sacarosa. Diver- sos explantes obtenidos de plántulas in vitro y plantas silvestres se cultivaron en diferentes combinaciones de 2,4-D, AIA, ANA y BAP, para inducir callos friables, dependiendo del tipo de explante y los reguladores de crecimiento. Suspensiones celulares se establecie- ron a partir de callos y estudiaron en su crecimiento y acumulación de metabolitos secundarios. Los metabolitos secundarios identifi- cados en cultivo de raíces y suspensiones celulares fueron feno- les, diversos flavonoides y la cumarina escopoletina

Dynamics of physiological and biochemical changes\ud during somatic embryogenesis of Acca sellowiana

Feijoa (Acca sellowiana [O. Berg] Burret [Myrtaceae]) is a native fruit species of southern Brazil and northern Uruguay. This species is amenable to somatic embryogenesis and therefore suitable as a model system for comparative studies of zygotic and somatic embryo development. In seed plants, embryogenesis involves three main steps, which are regulated by many factors, such as hormones, proteins, polyamines, and transcription factors. In the present work, the dynamics of protein, sugar, starch, amino acid, and polyamine accumulation were assayed during somatic embryogenesis of A. sellowiana. Protein, starch, amino acid, and polyamine levels accumulated unevenly during the induction phase of somatic embryogenesis, while the sugar content remained stable. Throughout the different developmental stages of somatic embryogenesis, synthesis and accumulation of proteins and amino acids showed patterns similar to those reported previously during the development of zygotic embryos of this same species. Differential patterns of polyamine accumulation were observed. This is important because these compounds affect the synthesis of other endogenous growth regulators, such as auxinindole-3-acetic acid that is mainly involved in the establishment of embryo polarity. Taken together, the present work brings new insights to the physiological and biochemical dynamics that occur during somatic embryogenesis of A. sellowiana

Ecophysiological and biochemical parameters for assessing Cr+6 stress conditions in Pterogyne nitens Tul. : new and usual methods for the management and restoration of degraded areas.

Heavy metals, such as Cr (chromium), have received attention during recent years due to their disposal in soil and water. Brave peanut (Pterogyne nitens Tul.), a tropical tree, was cultivated in 500 ?mol L-1 Cr+6 to observe how this species responds to this stress, with a view to its possible application in the restoration of degraded areas. In the present study, we report the use of a new method, photoacoustic spectroscopy, to evaluate the emission of CO2 and ethylene and the use of other standard techniques for assessing stress (chlorophyll a fluorescence, photosynthetic pigment determination, leaf relative water, specific leaf area, NO (nitric oxide) and polyamines. Photoacoustic spectroscopy appears to provide an innovative and efficient technique for detecting stress induced by heavy metals soon after their contact with plants. Polyamine content, as well as ethylene and NO, were seen to be sensitive to Cr+6; polyamines were decreased in leaves and increased in roots, ethylene was increased in the whole plant and NO was increased in the roots. Our results demonstrating alterations in ethylene in response to Cr+6 are, to date, unreported and the technique used for these measurements is novel. With regard to the polyamines, modulations in their concentrations may have protected the photosystem II, since no photosynthetic alterations were observed for pigments and chlorophyll a fluorescence

Morphological and polyamine content changes in embryogenic and non-embryogenic callus of sugarcane

Differences in competence acquisition and subsequent embryo maturation in embryogenic and non-embryogenic callus of sugarcane var. SP79-1011 were evaluated using histomorphological analysis, growth curves, numbers of somatic embryos, and polyamine contents. Embryogenic callus was formed by cells with embryogenic characteristics such as a rounded shape, prominent nuclei, a high nucleus: cytoplasm ratio, small vacuoles and organized globular structures. However, non-embryogenic callus presented dispersed, elongated and vacuolated cells with a low nucleus: cytoplasm ratio; these characteristics did not allow for the development of somatic embryos even upon exposure to a maturation stimulus. These results suggest that non-embryogenic callus does not acquire embryogenic competence during induction and that maturation treatment is not sufficient to promote somatic embryo differentiation. The use of activated charcoal (AC; 1.5 g L−1) resulted in a higher somatic embryo maturation rate in embryogenic callus but did not yield success in non-embryogenic callus. Embryogenic callus incubated with control (10 μM 2,4-dichlorophenoxyacetic acid) and maturation (1.5 g L−1 AC) treatments for 28 days showed similar patterns of total free polyamines; these results differed from the results observed with non-embryogenic callus, suggesting that embryogenic callus already exhibits a characteristic pattern of endogenous polyamine levels. At 28 days of culture with maturation treatment, embryogenic callus exhibited significantly higher levels of free Spm than embryogenic callus incubated with control treatment and non-embryogenic callus incubated with both treatments. This result suggests that Spm could be important for the acquisition of embryogenic competence and somatic embryo maturation in sugarcane var. SP79-1011.Funding for this work was provided by the CNPq (403015/2008-1 and 480142/2010-6) and FAPERJ (E-26/101.513/2010) to VS. AMV thanks CAPES for her fellowship

The transition of proembryogenic masses to somatic embryos in Araucaria angustifolia (Bertol.) Kuntze is related to the endogenous contents of IAA, ABA and polyamines

In somatic embryogenesis (SE) of conifers, the inability of many embryogenic cell lines to form well-developed somatic embryos may results from failure and constraints during the transition of proembryogenic masses (PEMs) to early somatic embryos. In the present work, we propose the inclusion of a preculture and prematuration steps looking at enhancing PEM III-to-early somatic embryos transition. It was further hypothesized that these results would correlate with the contents of endogenous indole-3-acetic acid (IAA), abscisic acid (ABA) and polyamines (PA). To test these hypotheses, the embryogenic culture was subjected to preculture with fluridone (FLD) and prematuration treatments with different combinations of carbon source and polyethylene glycol (PEG). The frequency of PEM III was increased after FLD preculture and the contents of IAA and ABA decreased, while the contents of PA increased. Putrescine (Put) was the most abundant PA present at this stage, followed by spermidine (Spd) and spermine (Spm). In early embryogenesis, prematuration treatments supplemented with maltose or lactose plus PEG enhanced the PEM III-to-early somatic embryos transition. IAA and ABA contents increased at this stage, while a decrease of the total free PA levels was observed. Put was the most abundant PA, followed by Spd and Spm, mainly in the treatment supplemented with PEG. This resulted in a decrease of PA ratio (Put/Spd + Spm) and, hence, PEM III-to-early somatic embryos transition. It was concluded that the preculture with FLD and prematuration treatments promote the PEM III-to-early somatic embryos transition throughout the whole early developmental process in Araucaria angustifolia.This study was supported by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq, Brazil), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brazil) and Fundação de Apoio à Pesquisa Cientifica e Inovação Tecnológica do Estado de Santa Catarina (FAPESC). This study is part of the Ph.D. thesis of the first author