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Made available in DSpace on 2021-05-04T12:31:44Z (GMT). No. of bitstreams: 1 3319-Texto-do-artigo-15721-1-10-20210118.pdf: 8641979 bytes, checksum: fac72fa024018329750b3412c0d1dc72 (MD5) Previous issue date: 2021 Título em português: Análise de transcriptoma de experimentos de RNA-Seq com e sem repetições biológicas: revisão.

Made available in DSpace on 2020-01-20T18:07:21Z (GMT). No. of bitstreams: 1 abertop3243UAVbasedthermalimagingintheassessmentofwaterstatusofsoybeanplants.pdf: 4532732 bytes, checksum: 97d48e35655a6e440885c0909f489306 (MD5) Previous issue date: 2020

Summary MicroRNAs (miRNAs) modulate the abundance and spatial–temporal accumulation of target mRNAs and indirectly regulate several plant processes. Transcriptional regulation of the genes encoding miRNAs (MIR genes) can be activated by numerous transcription factors, which themselves are regulated by other miRNAs. Fine‐tuning of MIR genes or miRNAs is a powerful biotechnological strategy to improve tolerance to abiotic or biotic stresses in crops of economic importance. Current approaches for miRNA fine‐tuning are based on the down‐ or up‐regulation of MIR gene transcription and the use of genetic engineering tools to manipulate the final concentration of these miRNAs in the cytoplasm. Transgenesis, cisgenesis, intragenesis, artificial MIR genes, endogenous and artificial target mimicry, MIR genes editing using Meganucleases, ZNF proteins, TALENs and CRISPR/Cas9 or CRISPR/Cpf1, CRISPR/dCas9 or dCpf1, CRISPR13a, topical delivery of miRNAs and epigenetic memory have been successfully explored to MIR gene or miRNA modulation and improve agronomic traits in several model or crop plants. However, advantages and drawbacks of each of these new biotechnological tools (NBTs) are still not well understood. In this review, we provide a brief overview of the biogenesis and role of miRNAs in response to abiotic or biotic stresses, we present critically the main NBTs used for the manipulation of MIR genes and miRNAs, we show current efforts and findings with the MIR genes and miRNAs modulation in plants, and we summarize the advantages and drawbacks of these NBTs and provide some alternatives to overcome. Finally, challenges and future perspectives to miRNA modulating in important crops are also discussed.

Abiotic stresses, especially drought, may seriously affect soybean yield. Due to the complexity of drought tolerance, one of the difficulties in selecting genotypes is the identification of traits contributing to improve stress tolerance. This study carried out soybean phenotyping under water deficit in vegetative and reproductive stages in field conditions at two consecutive crop seasons (2012/13 and 2013/14). The experiment was performed in Londrina, PR, Brazil, with two soybean cultivars with distinct levels of drought tolerance, BR 16 (sensitive) and Embrapa 48 (less sensitive). Water deficit was applied through rainout shelters, moving on rails to cover plots when the rainfall begins and uncover them when it ends. Then, some agronomic and morphological traits were measured. Multivariate statistics through the principal component analysis (PCA) associated with the biplot graph identified traits contributing to greater yield stability under drought. Result showed that water deficit affected soybean yield, mainly in the reproductive stage, in which Embrapa 48 had greater yield stability when compared to BR 16. However, opposite results were obtained for stress induced in the vegetative stage, when Embrapa 48 was more negatively affected than BR16 cultivar. Seed weight influenced yield differences between cultivars. Lighter seeds, but in larger number, constituted an advantage under water deficit. Due to the difficulty in introducing all drought tolerance mechanisms in one genotype, breeding programs need to define selection parameters according to regional drought conditions.

Abiotic stress, such as drought and flooding, are responsible for considerable losses in grain production worldwide. Soybean, the main cultivated oilseed in the world, is sensitive to both stresses. Plant molecular mechanisms answer via crosstalk of several signaling pathways, in which particular genes can respond to different stresses. Previous studies confirmed that overexpression of transcription factor AtAREB1 confers drought tolerance in soybean. However, plants containing this gene have not yet been tested under flooding. Thus, the objective of this study was to characterize genetically modified (GM) soybean plants overexpressing AtAREB1 under drought and flooding conditions in comparison to its genetic background. Physiological and biochemical measurements were performed. In addition, the expression level of genes commonly activated under both stresses was evaluated. The results supported the role of the AtAREB1 gene in conferring tolerance to water deficit in soybeans. Furthermore, under flooding, the GM line was efficient in maintaining a higher photosynthetic rate, intrinsic efficiency in water use, and instantaneous carboxylation efficiency, resulting in higher grain yield under stress. The GM line also presented higher protein content, lower concentration of hydrogen peroxide, and lower expression levels of genes related to fermentative metabolism and alanine biosynthesis. These results indicate that in addition to drought stress, plants overexpressing AtAREB1 exhibited better performance under flooding when compared to the non-GM line, suggesting a cross-signaling response to both abiotic factors.

Monitoring of soybean genotypes is important because of intellectual property over seed technology, better management over seed genetics, and more efficient strategies for its agricultural production process. This paper aims at spectrally classifying soybean genotypes submitted to diverse water availability levels at different phenological stages using leaf-based hyperspectral reflectance. Leaf reflectance spectra were collected using a hyperspectral proximal sensor. Two experiments were conducted as field trials: one experiment was at Embrapa Soja in the 2016/2017, 2017/2018, and 2018/2019 cropping seasons, where ten soybean genotypes were grown under four water conditions; and another experiment was in the experimental farm of Unoeste University in the 2018/2019 cropping season, where nine soybean genotypes were evaluated. The spectral data collected was divided into nine spectral datasets, comprising single and multiple cropping seasons (from 2016 to 2019), and two contrasting crop-growing environments. Principal component analysis, applied as an indicator of the explained variance of the reflectance spectra among genotypes within each spectral dataset, explained over 94% of the spectral variance in the first three principal components. Linear discriminant analysis, used to obtain a model of classification of each reflectance spectra of soybean leaves into each soybean genotype, achieved accuracy between 61% and 100% in the calibration procedure and between 50% and 100% in the validation procedure. Misclassification was observed only between genotypes from the same genetic background. The results demonstrated the great potential of the spectral classification of soybean genotypes at leaf-scale, regardless of the phenological stages or water status to which plants were submitted.

Although the information on the Normalized Difference Vegetation Index (NDVI) in plants under water deficit is often obtained from sensors attached to satellites, the increasing data acquisition with portable sensors has wide applicability in agricultural production because it is a fast, nondestructive method, and is less prone to interference problems. Thus, we carried out a set of experiments to investigate the influence of time, spatial plant arrangements, sampling size, height of the sensor and water regimes on NDVI readings in different soybean cultivars in greenhouse and field trials during the crop seasons 2011/12, 2012/13 and 2013/14. In experiments where plants were always evaluated under well-watered conditions, we observed that 9 a.m. was the most suitable time for NDVI readings regardless of the soybean cultivar, spatial arrangement or environment. Furthermore, there was no difference among NDVI readings in relation to the sampling size, regardless of the date or cultivar. We also observed that NDVI tended to decrease according to the higher height of the sensor in relation to the canopy top, with higher values tending to be at 0.8 m, but with no significant difference relative to 1.0 m—the height we adopted in our experiments. When different water regimes were induced under field conditions, NDVI readings measured at 9 a.m. by using a portable sensor were successful to differentiate soybean cultivars with contrasting responses to drought.

NMR and chromatography methods combined with mass spectrometry are the most important analytical techniques employed for plant metabolomics screening. Metabolomic analysis integrated to transcriptome screening add an important extra dimension to the information flow from DNA to RNA to protein. The most useful NMR experiment in metabolomics analysis is the proton spectra due the high receptivity of 1H and important structural information, through proton–proton scalar coupling. Routinely, databases have been used in identification of primary metabolites, however, there is currently no comparable data for identification of secondary metabolites, mainly, due to signal overlap in normal 1H NMR spectra and natural variation of plant. Related to spectra overlap, alternatively, better resolution can be find using 1H pure shift and 2D NMR pulse sequence in complex samples due to spreading the resonances in a second dimension. Thus, in data brief we provide a catalogue of metabolites and expression levels of genes identified in soy leaves and roots under flooding stress. Keywords: Soybean, Nuclear magnetic resonance, Primary metabolites, Secondary metabolites

Acidic soils occupy a vast area in the world, and the aluminum (Al) in these soils can directly interact with plant cells and tissues to inhibit their growth and reduce yields. The signalgrass Urochloa decumbens cv. Basilisk (syn. Brachiaria decumbens cv. Basilisk), a widely sown tropical forage grass, is recognized for its high productivity under intensive use, vigorous growth, ease of establishment, and good forage value throughout the year, as well as its exceptional adaptation to infertile acid soils. We sequenced the transcriptome from roots of U. decumbens cv. Basilisk under two conditions, with and without Al, using Illumina paired-end sequencing technology and performed de novo assembly of those reads, which yielded 164,920 transcripts. Of these transcripts, 113,918 were assigned a putative protein function through comparisons with different gene set databases. Additionally, 13,375 simple sequence repeat (SSR) markers were identified. Digital gene expression analyses were conducted to identify 6698 differentially expressed genes between treatments, revealing a great differences in the root transcriptional landscape when exposed to aluminum. An extensive annotation of the differentially expressed genes (DEGs), made possible to identify several transcripts with putative functions correlated to aluminum exposure, most belonging to vesicle transportation, cell wall modifications and metal handling ontologies. In this work, abundant, high-quality transcripts were obtained, providing a reference platform for future biotechnological studies and breeding programs for this species and its close relatives. Made available in DSpace on 2017-11-10T23:26:44Z (GMT). No. of bitstreams: 1 DenovoRNAsequencingandanalysis.pdf: 3315872 bytes, checksum: 114c260ce9b66a898f78ef1f1e06c51d (MD5) Previous issue date: 2017-11-10

Made available in DSpace on 2015-03-18T15:56:26Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-01-01Bitstream added on 2015-03-18T16:28:21Z : No. of bitstreams: 1 WOS000340383600167.pdf: 2463395 bytes, checksum: 0cfcb8fa6493ded313d5a66c62c6782c (MD5) Science and Technology Research Partnership for Sustainable Development (SATREPS) of the Japan Science and Technology Agency/Japan International Cooperation Agency (JIRCAS) Empresa Brasileira de Pesquisa Agropecuaria Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Faculdade de Ciencias Agrarias e Veterinaria Universidade Estadual Paulista (FCAV-UNESP), Jaboticabal, Brazil Abscisic acid-responsive element binding protein (AREB1) is a basic domain/leucine zipper transcription factor that binds to the abscisic acid (ABA)-responsive element motif in the promoter region of ABA-inducible genes. Because AREB1 is not sufficient to direct the expression of downstream genes under non-stress conditions, an activated form of AREB1 (AREB1 Delta QT) was created. Several reports claim that plants overexpressing AREB1 or AREB1 Delta QT show improved drought tolerance. In our studies, soybean plants overexpressing AREB1 Delta QT were characterized molecularly, and the phenotype and drought response of three lines were accessed under greenhouse conditions. Under conditions of water deficit, the transformed plants presented a higher survival rate (100%) than those of their isoline, cultivar BR 16 (40%). Moreover, the transformed plants displayed better water use efficiency and had a higher number of leaves than their isoline. Because the transgenic plants had higher stomatal conductance than its isoline under well-watered conditions, it was suggested that the enhanced drought response of AREB1 Delta QT soybean plants might not be associated with altered transpiration rates mediated by ABA-dependent stomatal closure. However, it is possible that the smaller leaf area of the transgenic plants reduced their transpiration and water use, causing delayed stress onset. The difference in the degree of wilting and percentage of survival between the 35S-AREB1 Delta QT and wildtype plants may also be related to the regulation of genes that protect against dehydration because metabolic impairment of photosynthesis, deduced by an increasing internal CO2 concentration, was not observed in the transgenic plants. Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada Agr, Jaboticabal, SP, Brazil Embrapa Soja, Lab Biotecnol Vegetal, Londrina, PR, Brazil Univ Estadual Londrina, Dept Biol Geral, Londrina, PR, Brazil Embrapa Soja, Dept Biometr, Londrina, PR, Brazil Embrapa Soja, Lab Ecofisiol Vegetal, Londrina, PR, Brazil Japan Int Res Ctr Agr Sci, Biol Resources & Postharvest Div, Tsukuba, Ibaraki, Japan Univ Tokyo, Bunkyo Ku, Lab Plant Mol Physiol, Tokyo 113, Japan Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Biol Aplicada Agr, Jaboticabal, SP, Brazil

Rhythms produced by the endogenous circadian clock play a critical role in allowing plants to respond and adapt to the environment. While there is a well-established regulatory link between the circadian clock and responses to abiotic stress in model plants, little is known of the circadian system in crop species like soybean. This study examines how drought impacts diurnal oscillation of both drought responsive and circadian clock genes in soybean. Drought stress induced marked changes in gene expression of several circadian clock-like components, such as LCL1-, GmELF4- and PRR-like genes, which had reduced expression in stressed plants. The same conditions produced a phase advance of expression for the GmTOC1- like, GmLUX-like and GmPRR7-like genes. Similarly, the rhythmic expression pattern of the soybean drought-responsive genes DREB-, bZIP-, GOLS-, RAB18- and Remorin-like changed significantly after plant exposure to drought. In silico analysis of promoter regions of these genes revealed the presence of cis-elements associated both with stress and circadian clock regulation. Furthermore, some soybean genes with upstream ABRE elements were responsive to abscisic acid treatment. Our results indicate that some connection between the drought response and the circadian clock may exist in soybean since (i) drought stress affects gene expression of circadian clock components and (ii) several stress responsive genes display diurnal oscillation in soybeans.

The objective of this work was to validate, by quantitative PCR in real time (RT-qPCR), genes to be used as reference in studies of gene expression in soybean in drought-stressed trials. Four genes commonly used in soybean were evaluated: GmB-actin, GmGAPDH, GmLectin and GmRNAr18S. Total RNA was extracted from six samples: three from roots in a hydroponic system with different drought intensities (0, 25, 50, 75 and 100 minutes of water stress), and three from leaves of plants grown in sand with different soil moistures (15, 5 and 2.5% gravimetric humidity). The raw cycle threshold (Ct) data were analyzed, and the efficiency of each primer was calculated for an overall analysis of the Ct range among the different samples. The GeNorm application was used to evaluate the best reference gene, according to its stability. The GmGAPDH was the least stable gene, with the highest mean values of expression stability (M), and the most stable genes, with the lowest M values, were the GmB-actin and GmRNAr18S, when both root and leaves samples were tested. These genes can be used in RT-qPCR as reference gene for expression analysis.

The objectives of this work were to determine the heliotropic movements of the upper trifoliates for two soybean cultivars, BR 16 and Embrapa 48, during a daily cycle, in three phenological stages and two water regimes, and to estimate the impact of irrigation and daily leaflet movements on agronomic characteristics and grain yield. Heliotropic movements were studied in three phenological stages: V4-V6, V7-V10, and R5 in irrigated and non-irrigated plots. For each stage, the leaflet elevation and azimuth were measured hourly. Under a low (V4-V6 stage) and mid (V7-V10 stage) leaf area index (LAI) the diaheliotropism was slightly more frequent and intensive in non-irrigated than in irrigated plants, only at early morning and late afternoon hours. At R5 stage (high LAI) the paraheliotropism of superior trifoliates was predominant and more intensive in non-irrigated plants. The heliotropic movements are correlated to carbon gain, but not to environment (light intensity or temperature), for measurements at 11h. 'Embrapa 48' expresses greater paraheliotropism than 'BR 16' at high LAI, while 'BR 16' displays lower heliotropic plasticity under irrigation. In spite of significant heliotropic differences, genotype and water availability treatments did not influence the final grain yield. Os objetivos deste trabalho foram determinar o movimento heliotrópico dos trifólios superiores de duas cultivares de soja, BR 16 e Embrapa 48, durante o dia, em três estádios fenológicos e em dois regimes hídricos e determinar o impacto da irrigação e do movimento foliar diário nas características agronômicas e na produção de grãos. Movimentos heliotrópicos foram determinados a cada hora, em três estádios fenológicos: V4-V6, V7-V10 e R5, em parcelas irrigadas e não irrigadas. Para cada estágio, a elevação do folíolo e o azimute foram medidos por hora. Em índices de área foliar (IAF) baixo e médio (V4-V6 e V7-V10), o diaheliotropismo foi levemente mais intenso e frequente nas plantas sem irrigação do que nas irrigadas, e somente nas primeiras horas matinais e últimas horas da tarde. Em R5 (alto IAF), o paraheliotropismo dos trifólios superiores foi predominante e mais intenso em plantas não irrigadas. Os movimentos heliotrópicos se correlacionaram com o ganho de carbono mas não com o ambiente (intensidade de luz ou temperatura), para medidas às 11 horas. 'Embrapa 48' apresenta maior paraheliotropismo do que 'BR 16', em plantas com alto IAF enquanto 'BR 16' apresenta menor plasticidade nas respostas heliotrópicas nas parcelas irrigadas. Apesar de diferenças no heliotropismo, o genótipo e disponibilidade de água não influenciaram a produção de grãos.