Selection of internal control genes for quantitative real-time RT-PCR studies during tomato development process

<p>Abstract</p> <p>Background</p> <p>The elucidation of gene expression patterns leads to a better understanding of biological processes. Real-time quantitative RT-PCR has become the standard method for in-depth studies of gene expression. A biologically meaningful reporting of target mRNA quantities requires accurate and reliable normalization in order to identify real gene-specific variation. The purpose of normalization is to control several variables such as different amounts and quality of starting material, variable enzymatic efficiencies of retrotranscription from RNA to cDNA, or differences between tissues or cells in overall transcriptional activity. The validity of a housekeeping gene as endogenous control relies on the stability of its expression level across the sample panel being analysed. In the present report we describe the first systematic evaluation of potential internal controls during tomato development process to identify which are the most reliable for transcript quantification by real-time RT-PCR.</p> <p>Results</p> <p>In this study, we assess the expression stability of 7 traditional and 4 novel housekeeping genes in a set of 27 samples representing different tissues and organs of tomato plants at different developmental stages. First, we designed, tested and optimized amplification primers for real-time RT-PCR. Then, expression data from each candidate gene were evaluated with three complementary approaches based on different statistical procedures. Our analysis suggests that SGN-U314153 (<it>CAC</it>), SGN-U321250 (<it>TIP41</it>), SGN-U346908 ("<it>Expressed</it>") and SGN-U316474 (<it>SAND</it>) genes provide superior transcript normalization in tomato development studies. We recommend different combinations of these exceptionally stable housekeeping genes for suited normalization of different developmental series, including the complete tomato development process.</p> <p>Conclusion</p> <p>This work constitutes the first effort for the selection of optimal endogenous controls for quantitative real-time RT-PCR studies of gene expression during tomato development process. From our study a tool-kit of control genes emerges that outperform the traditional genes in terms of expression stability.</p

Molecular analysis of menadione-induced resistance against biotic stress in Arabidopsis

19 páginas, 6 figuras, 2 tablas.Menadione sodium bisulphite (MSB) is a water-soluble derivative of vitamin K3, or menadione, and has been previously demonstrated to function as a plant defence activator against several pathogens in several plant species. However, there are no reports of the role of this vitamin in the induction of resistance in the plant model Arabidopsis thaliana. In the current study, we demonstrate that MSB induces resistance by priming in Arabidopsis against the virulent strain Pseudomonas syringae pv. tomato DC3000 (Pto) without inducing necrosis or visible damage. Changes in gene expression in response to 0.2 mm MSB were analysed in Arabidopsis at 3, 6 and 24 h post-treatment using microarray technology. In general, the treatment with MSB does not correlate with other publicly available data, thus MSB produces a unique molecular footprint. We observed 158 differentially regulated genes among all the possible trends. More up-regulated genes are included in categories such as 'response to stress' than the background, and the behaviour of these genes in different treatments confirms their role in response to biotic and abiotic stress. In addition, there is an over-representation of the G-box in their promoters. Some interesting functions are represented among the individual up-regulated genes, such as glutathione S-transferases, transcription factors (including putative regulators of the G-box) and cytochrome P450s. This work provides a wide insight into the molecular cues underlying the effect of MSB as a plant resistance inducer.This work was partially funded by an INVESCAN, S.L. grant (No.OTT2001438) to the CSIC and by a BIO2006-02168 grant of MICINN to PT. The microarrays were funded in part by the “Genome España” Foundation. MER was supported by a research contract (ID-TF-06/002) from the Consejería de Industria, Comercio y Nuevas Tecnologías (Gobierno de Canarias). The authors thank CajaCanarias for their research support. We also thank Lorena Perales for her help in performing the bacterial growth curves, Dr. Héctor Cabrera for his useful advice on writing the manuscript, the English translation service of the Universidad Politécnica de Valencia and Mrs. Pauline Agnew whose endeavoured to edit the English translation of this paper.Peer reviewe

Uso de menadiona para aumentar la tolerancia al estrés salino de las plantas

Uso de menadiona para aumentar la tolerancia al estrés salino de las plantas. Esta invención está relacionada con el uso de composiciones que contienen menadiona (Vitamina K3 o Provitamina K) y/o algunos de sus derivados hidrosolubles para la mejora de la tolerancia al estrés salino de las plantas afectadas. El tratamiento se lleva a cabo mediante la aplicación de las soluciones que contengan los citados compuestos por cualquier vía que ponga en contacto las composiciones de la invención con cualquier parte de la planta y/o semillas.Peer reviewedB1 Patente con informe sobre el estado de la ténic

Bioinformatics Discovery of Vertebrate Cathelicidins from the Mining of Available Genomes

Due to the worrying increase in antimicrobial resistance to conventional antibiotics, the search for alternatives is becoming increasingly important. Antimicrobial peptides (AMPs), originating from natural resources, have been recognised as a novel class of antibiotics. An advantage of peptides over antibiotics is that the resistance is more difficult to attain than for conventional antibiotics. With the increasing number of genomes sequenced and available in the public domain, one alternative methodology to obtain novel AMPs is to analyse genes and proteins from genomic databases to predict and identify amino acid sequences that share similarities and molecular features with natural bioactive antimicrobial peptides. In this chapter, we summarise some of our recent results on the production of antimicrobial peptides, particularly, how we managed to identify a family of antimicrobial peptides: cathelicidins, through bioinformatics tools, from the genomes of two lower vertebrates (a reptile and a bird) available in public databases. We hope that our preliminary investigation with these novel peptides could be useful for the design of future strategies that pursue the production of antimicrobial peptides through biotechnology

Gestión de restricciones físicas a través de indicadores logísticos

In this article is proposed one proceeding to management physical restraints in the logistic systems, follow-up the general philosophy of the Theory of Constraints through logistic indicators like main instrument. In order to evidence its pertinence a case of study in a trading enterprise in Cuba shows up. A group of indicators of logistic management that measure the performance of the entity was obtained, and in terms of these were determined the restrictive resources that affect its effectiveness. From its identification, the physical constraint of logistic system was able to rise.En este artículo se propone un proceder para la gestión de las restricciones físicas en los sistemas logísticos en empresas comercializadoras, que sigue la filosofía general de la Teoría de las Restricciones y la utilización de indicadores logísticos como principal herramienta. Para evidenciar su pertinencia se presenta un caso de aplicación en una entidad comercializadora en Cuba. Se obtuvo un conjunto de indicadores de gestión logística que miden el desempeño de la entidad, y en función de estos se determinaron los recursos restrictivos que afectan su efectividad. A partir de su identificación se logró elevar la restricción física del sistema logístico

Effect of Enerplant® doses on the development and nutrient use by cacao (Theobroma cacao L.) seedlings

Objective: To evaluate the effect of the application of doses of a biostimulant on the development, absorption and efficiency of nitrogen, phosphorus and potassium utilization by cocoa seedlings. Design / methodology / approach. In a completely randomized design, 4 doses of Enerplant® (0; 1; 2; 3 and 4 mL ha-1.) applied to cocoa seedlings grown under saran mesh were studied. The biostimulant was applied monthly via foliar. To determine the absorption of nutrients, 3 plants of each treatment were divided into leaves, stems and roots, dried for 72 hours at 65 ° C; they were ground, packaged, labeled, and sent to the laboratory, where the macronutrients of each component were determined. Results. The cocoa seedlings responded positively to the application of the biostimulant. As an average of the two years of study, Enerplant® in doses of 1 mL ha-1 significantly increased height (24%), dry mass (30%); the quality index (58%) and the foliar area (56%) of the cocoa seedlings. Doses higher than 1 - 2 mL ha-1 decreased plant growth. The pattern of nutrient absorption by cocoa in the nursery phase behaved in the order potassium&gt; nitrogen&gt; phosphorus Conclusions. The efficiency of nutrient utilization, regardless of the Enerplant® dose applied, was higher for phosphorus followed by nitrogen and finally potassium The application of the biostimulant in doses of 2 ml ha-1 guaranteed the highest absorption and efficiency values of utilization of macronutrients by cocoa seedlingsObjective: To evaluate the effect of the application of biostimulant doses on the development, absorption, and use efficiency of nitrogen, phosphorus, and potassium by cacao seedlings. Design/Methodology/Approach: Five doses of Enerplant® (0, 1, 2, 3, and 4 mL ha-1) were applied to cacao seedlings grown under a saran mesh. The study was carried out using a completely randomized design. The foliar area received a monthly application of the biostimulant. In order to determine the nutrient absorption, 3 plants per treatment were divided into leaves, stems, and roots; they were subsequently dried for 72 h at 65 °C. Finally, they were ground, packaged, labeled, and sent to the laboratory, where the macronutrients of each component were analyzed. Results: The cacao seedlings had a positive response to the biostimulant application. In average, the results of our two-year study were: 1 mL ha-1 doses of Enerplant® significantly increased height (24%), dry mass (30%), quality index (58%), and foliar area (56%) of the cacao seedlings. Doses higher than 1- and 2-mL ha-1 decreased plant growth. The nutrient absorption by cacao in the nursery stage followed the following pattern: K &gt; N &gt; P. Findings/Conclusions: The efficiency of nutrient use —regardless of the Enerplant® dose applied— was higher for P, followed by N and, finally, K. Applying 2 mL ha-1 doses of the biostimulant guaranteed the highest absorption and use efficiency values of macronutrients by cacao seedlings

Risk of suicide attempt repetition after an index attempt: A systematic review and meta-analysis

Objectives To estimate the risk of suicide attempt repetition among individuals with an index attempt. It also aims to study the role of risk factors and prevention programme in repetition. Methods This systematic review and meta-analysis was conducted in keeping with the PRISMA 2020 guidelines. Studies on attempt repetition (both cohort studies and intervention studies) were searched from inception to 2022. Results A total of 110 studies comprising 248,829 attempters was reviewed. The overall repetition rate was 0.20 (0.17, 0.22). Repetition risk linearly increased over time. A higher risk of attempt repetition was associated with female sex and index attempts in which self-cutting methods were used. Moreover, a mental disorder diagnosis was associated with an increasing repetition risk (OR = 2.02, p < .01). The delivery of a preventive programme reduced the repetition risk, OR = 0.76, p < .05; however, this effect was significant for psychotherapy interventions, OR = 0.38, p < .01. Conclusion One in five suicide attempters will engage in a new suicide attempt. An elevated repetition risk is associated with being female, more severe index methods and psychiatric disorder diagnosis. Preventive programmes, particularly psychotherapy, may contribute to reducing repetition risk and eventually save lives.This study was supported by the Instituto de Salud Carlos III-FIS research grants (PI16/00187, PI19/00236, PI19/00569, PI19/00685, PI19/00941, PI19/00954, PI19/01027, PI19/01256, PI19/01484, PI20/00229), co-funded by the European Regional Development Fund (ERDF) “A Way to Build Europe”; the Government of the Principality of Asturias (grant ref.: PCTI-2018-2022 IDI/2018/235); Secretaria d'Universitats i Recerca from the Departament d'Economia i Coneixement (ref.: 2017SGR1365 and 2017SGR134), and Generalitat de Catalunya (Government of Catalonia), CERCA Programme

Propuestas en las ruinas del Convento de San Antón. Castrojeriz

Propuestas de intervención en las ruinas del Convento de San Antón de Castrojeriz.Departamento de Teoría de la Arquitectura y Proyectos ArquitectónicosDepartamento de Teoría de la Arquitectura y Proyectos ArquitectónicosAsociación Hospital de Peregrinos San AntónDemarcación de Burgos del COACYL

Treatment with tocilizumab or corticosteroids for COVID-19 patients with hyperinflammatory state: a multicentre cohort study (SAM-COVID-19)

Objectives: The objective of this study was to estimate the association between tocilizumab or corticosteroids and the risk of intubation or death in patients with coronavirus disease 19 (COVID-19) with a hyperinflammatory state according to clinical and laboratory parameters. Methods: A cohort study was performed in 60 Spanish hospitals including 778 patients with COVID-19 and clinical and laboratory data indicative of a hyperinflammatory state. Treatment was mainly with tocilizumab, an intermediate-high dose of corticosteroids (IHDC), a pulse dose of corticosteroids (PDC), combination therapy, or no treatment. Primary outcome was intubation or death; follow-up was 21 days. Propensity score-adjusted estimations using Cox regression (logistic regression if needed) were calculated. Propensity scores were used as confounders, matching variables and for the inverse probability of treatment weights (IPTWs). Results: In all, 88, 117, 78 and 151 patients treated with tocilizumab, IHDC, PDC, and combination therapy, respectively, were compared with 344 untreated patients. The primary endpoint occurred in 10 (11.4%), 27 (23.1%), 12 (15.4%), 40 (25.6%) and 69 (21.1%), respectively. The IPTW-based hazard ratios (odds ratio for combination therapy) for the primary endpoint were 0.32 (95%CI 0.22-0.47; p < 0.001) for tocilizumab, 0.82 (0.71-1.30; p 0.82) for IHDC, 0.61 (0.43-0.86; p 0.006) for PDC, and 1.17 (0.86-1.58; p 0.30) for combination therapy. Other applications of the propensity score provided similar results, but were not significant for PDC. Tocilizumab was also associated with lower hazard of death alone in IPTW analysis (0.07; 0.02-0.17; p < 0.001). Conclusions: Tocilizumab might be useful in COVID-19 patients with a hyperinflammatory state and should be prioritized for randomized trials in this situatio

Post-Franco Theatre

In the multiple realms and layers that comprise the contemporary Spanish theatrical landscape, “crisis” would seem to be the word that most often lingers in the air, as though it were a common mantra, ready to roll off the tongue of so many theatre professionals with such enormous ease, and even enthusiasm, that one is prompted to wonder whether it might indeed be a miracle that the contemporary technological revolution – coupled with perpetual quandaries concerning public and private funding for the arts – had not by now brought an end to the evolution of the oldest of live arts, or, at the very least, an end to drama as we know it