Down-regulation of a pectin acetylesterase gene modifies strawberry fruit cell wall pectin stracture and increases fruit firmness

Antisense-mediated down-regulation of several fruit-specific genes has previously demonstrated how the cell wall disassembly in strawberry fruit is mediated by a series of enzymes that act sequentially (Posé et al. 2011). An interesting example, the silencing of the polygalacturonase gene FaPG1, was also related with a significant increase of the post-harvest strawberry fruit firmness (Posé et al. 2013). Our research group has isolated a pectin acetylesterase gene, FaPAE1, which expression is enhanced during strawberry ripening. The main goal of this work was to elucidate the role of the degree of acetylation in cell wall integrity and fruit firmness through the antisense-mediated down-regulation of FaPAE1 in strawberry plants. Several transgenics lines were generated and 5 of them produced fruits 5-15% firmer than controls. Cell wall from ripe fruits was isolated from two independent transgenic lines and a control line, and sequentially extracted with different solvents (PAW, H2O, CDTA, Na2CO3). Modifications in fraction yield, its sugar composition and the degree of acetylation in each fraction were determined. Higher amounts of CDTA and Na2CO3 fractions were obtained in transgenic fruits, suggesting a decreased pectin solubilization as results of FaPAE1 silencing. Accordingly, the degree of acetylation of the Na2CO3-soluble pectins was greater in the transgenic lines than the control, but the opposite result was found in pectins from the CDTA fraction. These results suggest that PAE is preferentially active in pectis that are tightly bound to the cellulose-hemicellulose network and its activity could reduce the complexity of the cell wall structure, allowing that other hydrolytic enzymes could access the pectin chains. Thus, the increased fruit firmness observed in the transgenic FaPAE1 lines could be attributed to the direct effect of the silencing of the PAE enzyme and also to the indirect effect that the increase of the degree of acetylation of pectins has on the activity of other enzymes involved in the cell wall degradation. * Posé et al. (2011). Genes, Genomes and Genomics, 5 (Special Issue 1):40-48 * Posé et al. (2013). Plant Physiology, 150: 1022-1032 We acknowledge support from the Spanish Ministry of Economy and competitivity and Feder EU Funds (grant reference AGL2011-24814), FPI fellowships support for SP (BES-2006-13626) and CP (BES-2009027985), and grant "Ramón y Cajal" support for AJMA (RYC-2011-08839).Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Islanding detection in grid-connected power converters using harmonics due to the non-ideal behavior of the inverter

This paper analyzes the use of the voltage distortions in PWM voltage-source-inverters (VSIs) caused by the non-ideal behavior of the inverter for islanding detection purposes. The non-ideal characteristic of the inverters, mainly due to the dead-time needed to have safe commutations, produces fundamental frequency dependent harmonics (-5th, 7th...) in the output voltage. Although these harmonics are in principle an unwanted effect, since they reduce the power quality, they can potentially be used for islanding detection purposes. The physical principles of the method would be the same as for high frequency signal injection methods that have already been proposed but without the need of injecting a high frequency signa

Sensorless control of doubly-fed induction generators based on stator high frequency signal injection

High frequency signal injection based methods have been widely investigated for sensorless position/speed control of induction machines (IMs), permanent magnet synchronous machines (PMSMs) and more recently for doubly fed induction generators (DFIGs). When used with IMs and PMSMs, the high frequency signal is injected in the stator windings, an asymmetric (salient) rotor being required for this case. Contrary to this, both stator and rotor terminals are accessible and sensored in DFIGs, being therefore possible to inject the high frequency signal either in the stator or the rotor terminals. As consequence of this, the method can be used even if the machine is non-salient. In the implementation of the method with DFIGs, the high frequency voltage signal is typically injected in the rotor, the high frequency components (voltages of currents) induced in the stator being used for rotor position estimation. A drawback of this alternative is that the method is sensitive to the grid impedance in the stator side, which will be affected by the grid configuration, and is normally unknown. This paper proposes the sensorless control a DFIG injecting the high frequency voltage in the stator side, and using a high frequency current cancellation strategy in the rotor side. The main advantage of the proposed strategy is that the estimated position is independent of the grid characteristic

Coordinated operation of parallel-connected inverters for active islanding detection using high frequency signal injection

The high frequency impedance measured at the terminals of inverters connected in a microgrid by means of the injection of a small magnitude, high frequency voltage, has been shown to be a reliable metric to detect islanding. While the implementation of this method is simple when only an inverter injects the high frequency signal, this case is of limited applicability. On the other hand, several concerns arise when multiple inverters work in parallel, primarily due to risk interference among inverters. Islanding detection using high frequency signal injection in microgrids with multiple parallel-connected inverters is studied in this paper. A strategy for the coordinated operation of the inverters, without the need of communications or pre-established roles is proposed. Simulation and experimental results will be provided to demonstrate the viability of the concep

Islanding detection in three-phase and single-phase systems using pulsating high frequency signal injection

This paper analyzes the use of pulsating high frequency signal injection for islanding detection purposes. Active islanding detection using high frequency signal injection is an appealing option due to its reduced non-detection zone, reduced cost and ease of implementation. The use of a rotating high frequency signal has been reported and analyzed. However, this method can only be applied to three-phase systems. In this paper, the use of a pulsating high frequency signal injection is proposed. While it uses the same principles as rotating signal injection, it can be applied to both threephase and single-phase system

Rhamnogalacturonase lyase gene downregulation in strawberry and its potential on mechanical fruit properties

Strawberry softening is one of the main factors that reduces fruit quality and leads to economically important losses. Textural changes during fruit ripening are mainly due to the dissolution of middle lamellae, a reduction in cell-to-cell adhesion and the weakening of parenchyma cell walls as a result of the action of cell wall modifying enzymes. Functional studies of genes encoding pectinase enzymes (polygalacturonase, pectate lyase and -galactosidase) support a key role of pectin disassembly in strawberry softening. Evidence that RG-I may play an important role in strawberry texture has been obtained from the transient silencing of a RG-lyase gene. Pectins are major components of fruit cell walls and highly dynamic polysaccharides, but due to their heterogeneity the precise relation between the structures and functions is incomplete. In this work, stable transgenic strawberry lines with a rhamnogalacturonate lyase gene (FaRGLyase1) down-regulated have been analyzed. Several transgenic lines showing more than 95% silencing of FaRGLyase1 displayed fruit firmness values higher than control. Cell walls from these lines were extracted and analyzed by ELISA and Epitope Detection Chromatography (EDC). This last technique is based on the detection of specific cell wall oligosaccharide epitopes and provides information on sub-populations of pectins containing homogalacturonan and RG-I domains, but also reveals potential links with other cell wall polysaccharides such as xyloglucan. The results obtained indicate that the silencing of FaRGLyase1 reduces degradation of RG-I backbones, but also homogalacturonan, in cell walls, especially in pectin fractions covalently bound to the cell wall. These changes contribute to the increased firmness of transgenic fruits.This research was supported by FEDER EU Funds and the Ministerio de Economía y Competitividad of Spain (grant reference AGL2014-55784-C2), a Marie Curie IEF within the 7th European Community Framework Programme (reference: PIEF-2013-625270) for SP and a FPI fellowship (BES-2015-073616) to support PR-V. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Implementacion y análisis de métodos numéricos basados en flujo potencial con aplicación en turbomáquinas

Este artículo aborda la resolución de flujos no viscosos y en particular su aplicación al estudio de cascada de álabes. Para ello se analizarán modelos de flujo potencial resolviendo la ecuación de Laplace mediante diferentes variaciones del método de los Paneles. Se ha realizado una validación de los resultados obtenidos para perfiles aislados con valores experimentales obtenidos por la NACA. Además, se ha realizado un estudio comparativo entre los distintos resultados obtenidos con los diferentes métodos de los paneles implementados(Source, Vortex y Source-Vortex). Una vez comprobada la bondad de los distintas variantes del método de los paneles para perfiles aislados se ha ampliado el método a cascadas de álabes y se ha realizado también una validación con valores experimentales obtenidos de la bibliografía.In this work an analysis of different panel methods applied to turbomachinery has been presented, within the conditions of potential flow as well its behaviour with different shapes and its advantadges and inconvenients applied to aerodynamics profiles. Finally one of these methods have been extended and corrected to the analysis of compressor cascade

Context-aware Assessment Using QR-codes

In this paper we present the implementation of a general mechanism to deliver tests based on mobile devices and matrix codes. The system is an extension of Siette, and has not been specifically developed for any subject matter. To evaluate the performance of the system and show some of its capabilities, we have developed a test for a second-year college course on Botany at the School of Forestry Engineering. Students were equipped with iPads and took an outdoor test on plant species identification. All students were able to take and complete the test in a reasonable time. Opinions expressed anonymously by the students in a survey about the usability of the system and the usefulness of the test were very favorable. We think that the application presented in this paper can broaden the applicability of automatic assessment techniques.The presentation of this work has been co-founded by the Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Efecto del silenciamiento de genes que codifican poligalacturonasas sobre el reblandecimiento del fruto de fresa asociado a la maduración

La degradación de las pectinas de la pared celular mediada por poligalacturonasas juega un papel clave en el reblandecimiento de la fresa. Así, el silenciamiento del gen FaPG1 incrementa la firmeza del fruto maduro y alarga su vida postcosecha. Además de FaPG1, en fresa se ha descrito otro gen que codifica una poligalacturonasa específica de maduración, FaPG2. Con el fin de profundizar en el papel de estos genes, se han obtenido plantas transgénicas con el gen FaPG2 silenciado (líneas BPG), así como plantas con FaPG1 y FaPG2 silenciados (líneas ABPG), obtenidas mediante retransformación de una línea antiFaPG1 (APG29) que mostraba un fuerte silenciamiento del gen y un incremento en la firmeza de fruto. Se obtuvieron 24 líneas BPG y 15 ABPG. Estas plantas, junto con la línea APG29 y controles sin transformar, fueron analizadas durante 3 años consecutivos. El 50% de las líneas BPG mostraron mayor firmeza de fruto rojo que el control sin transformar, aunque el incremento en firmeza fue similar al obtenido en la línea APG29. Todas las líneas dobles transformantes dieron frutos de mayor firmeza que el control, siendo los valores ligeramente superiores a los de la línea APG29 en alguna de ellas. A nivel de expresión, las líneas BPG seleccionadas mostraron un silenciamiento del gen FaPG2 que varió entre el 60-70%, e inesperadamente, un silenciamiento significativo de FaPG1, a pesar de la baja homología entre ambos genes. El silenciamiento de FaPG1 en las líneas ABPG fue superior al 95%; sin embargo, el silenciamiento de FaPG2 fue similar al obtenido en las plantas BPG. Estos resultados confirman el papel clave de las poligalacturonasas en el reblandecimiento de la fresa y sugieren la existencia de una regulación compleja en la expresión de ambos genes. Este trabajo ha sido financiado por el proyecto AGL2011-24814 y Fondos FEDERUniversidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Expresión del gen chit42 de Trichoderma harzianum en plantas de fresa: obtención de plantas de fresa con mayor resistencia a la infección por Colletrotrichum acutatum

La antracnosis, causada por diferentes especies del hongo Colletotrichum, origina importantes pérdidas de producción en el cultivo de la fresa, un cultivo de gran interés agrícola en Andalucía, que aporta el 92.01% de la producción nacional y que, a su vez, genera más del 10% de la producción mundial total. C. acutatum está considerado un organismo de cuarentena en Europa (Directiva EC 77/93, Real Decreto 2071/1993). No se conocen variedades de fresa totalmente resistentes a C. acutatum y pese a la importancia agronómica de este cultivo en el mundo, existen muy pocos estudios moleculares de los mecanismos relacionados con la defensa de la planta de fresa a Colletotrichum spp. Esta falta de conocimiento molecular afecta negativamente a programas de mejora genética de la resistencia de la fresa a patógenos que, ya en sí, se ven notablemente mermados por el hecho de que la fresa es un cultivo de propagación clonal. Nuestro grupo PAI CVI-278 lleva más de 10 años investigando en fresa a nivel de biología molecular los mecanismos relacionados con diversos procesos bioquímicos, como la maduración del fruto y la defensa y resistencia de la planta a patógenos. Para incrementar la resistencia de la planta a C. acutatum, nuestro grupo está produciendo líneas transgénicas de fresa que expresan genes, procedentes de especies del hongo Trichoderma, que codifican enzimas que actúan sobre las paredes celulares de hongos ascomicetos y basidiomicetos, y que tienen, sustancialmente, una mayor actividad antifúngica que las propias enzimas de la planta y son activas frente a un rango más amplio de fitopatógeno