RESUMO
Reverse Transcription quantitative PCR (RT-qPCR) is a technique for gene expression profiling with high sensibility and reproducibility. However, to obtain accurate results, it depends on data normalization by using endogenous reference genes whose expression is constitutive or invariable. Although the technique is widely used in plant stress analyzes, the stability of reference genes for iron toxicity in rice (Oryza sativa L.) has not been thoroughly investigated. Here, we tested a set of candidate reference genes for use in rice under this stressful condition. The test was performed using four distinct methods: NormFinder, BestKeeper, geNorm and the comparative ΔCt. To achieve reproducible and reliable results, Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines were followed. Valid reference genes were found for shoot (P2, OsGAPDH and OsNABP), root (OsEF-1a, P8 and OsGAPDH) and root+shoot (OsNABP, OsGAPDH and P8) enabling us to perform further reliable studies for iron toxicity in both indica and japonica subspecies. The importance of the study of other than the traditional endogenous genes for use as normalizers is also shown here.
Assuntos
Ferro/toxicidade , Oryza/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/genética , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Oryza/genética , Fator 1 de Elongação de Peptídeos/genética , Fator 1 de Elongação de Peptídeos/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Brotos de Planta/efeitos dos fármacos , Brotos de Planta/genética , RNA de Plantas/isolamento & purificação , RNA de Plantas/metabolismo , Reação em Cadeia da Polimerase em Tempo Real/normas , Padrões de Referência , Plântula/efeitos dos fármacos , Plântula/genética , Transcrição Gênica/efeitos dos fármacosRESUMO
Iron is a well-known metal. Used by humankind since ancient times in many different ways, this element is present in all living organisms, where, unfortunately, it represents a two-way problem. Being an essential block in the composition of different proteins and metabolic pathways, iron is a vital component for animals and plants. That is why iron deficiency has a severe impact on the lives of different organisms, including humans, becoming a major concern, especially in developing countries where access to adequate nutrition is still difficult. On the other hand, this metal is also capable of causing damage when present in excess, becoming toxic to cells and affecting the whole organism. Because of its importance, iron absorption, transport and storage mechanisms have been extensively investigated in order to design alternatives that may solve this problem. As the understanding of the strategies that plants use to control iron homeostasis is an important step in the generation of improved plants that meet both human agricultural and nutritional needs, here we discuss some of the most important points about this topic.
RESUMO
Abstract Iron is a well-known metal. Used by humankind since ancient times in many different ways, this element is present in all living organisms, where, unfortunately, it represents a two-way problem. Being an essential block in the composition of different proteins and metabolic pathways, iron is a vital component for animals and plants. That is why iron deficiency has a severe impact on the lives of different organisms, including humans, becoming a major concern, especially in developing countries where access to adequate nutrition is still difficult. On the other hand, this metal is also capable of causing damage when present in excess, becoming toxic to cells and affecting the whole organism. Because of its importance, iron absorption, transport and storage mechanisms have been extensively investigated in order to design alternatives that may solve this problem. As the understanding of the strategies that plants use to control iron homeostasis is an important step in the generation of improved plants that meet both human agricultural and nutritional needs, here we discuss some of the most important points about this topic.
RESUMO
Foram estudadas alternativas para a redução da adstringência (destanização) e conservação de frutos de jambolão (Syzygium cumini, L.). Para isso, no primeiro experimento, os frutos foram colhidos no estádio meio-maduro, mantidos sob temperatura ambiente e submetidos aos seguintes tratamentos: controle (atmosfera normal), 98kPa CO2, 20kPa CO2, 98kPa N2, etileno (100ppm), 1-MCP (1ppm) seguido de etileno (100ppm) e 3,85mL de etanol absoluto L-1. Foram avaliados os teores de acetaldeído e de taninos solúveis, a firmeza e a adstringência dos frutos. No segundo experimento, os frutos foram colhidos no estádio maduro e armazenados em atmosfera modificada passiva, sob temperatura ambiente (AMPA, 23±1°C) e atmosfera modificada passiva refrigerada (AMPR, 4±1°C), pelo período de cinco, 10 e 15 dias. Frutos provenientes dos tratamentos com N2, etileno e etanol apresentaram os melhores resultados quanto à destanização, com aumento nos teores de acetaldeído e diminuição dos taninos solúveis e da adstringência; entretanto, os tratamentos com etileno e etanol causaram amolecimento excessivo nos frutos. O armazenamento em AMPR conservou melhor a coloração roxo-azulada dos frutos e, além de prevenir a incidência de fungos, auxiliou na manutenção dos teores de fenóis totais e de ácido ascórbico e na atividade antioxidante dos frutos.
Alternative methods were investigated for the reduction of astringency (detannization) and conservation of Jambolan fruit (Syzygium cumini, L.). In the first experiment, fruits were harvested at an intermediate ripening stage, maintained at room temperature and subjected to the following treatments: control (normal atmosphere), 98kPa CO2, 20kPa CO2, 98kPa N2, ethylene (100ppm), 1-MCP (1ppm) followed by treatment with ethylene (100ppm) and 3,85mL absolute ethanol L-1. Acetaldehyde, soluble tannin, fruit firmness and astringency were evaluated. In the second experiment, fruits were harvested ripe and stored under passive modified atmosphere at room temperature (PMART, 23±1°C) and passive modified atmosphere under cold storage (PMACS, 4±1°C) for 5, 10 and 15 days. Fruits treated with N2, ethylene and ethanol showed the best detannization results, with increased acetaldehyde content, reduction of soluble tannins and astringency. However ethylene and ethanol treatments induced excessive softening of the fruit. Storage under PMAR kept the purple-blue color of fruits and, besides preventing fungal incidence, it aided the maintenance of the total phenolic content, ascorbic acid and antioxidant activity of fruits.