Biomass Accumulation and Cell Wall Structure of Rice Plants Overexpressing a Dirigent-Jacalin of Sugarcane (ShDJ) Under Varying Conditions of Water Availability

A sugarcane gene encoding a dirigent-jacalin, ShDJ, was induced under drought stress. To elucidate its biological function, we integrated a ShDJ-overexpression construction into the rice Nipponbare genome via Agrobacterium-mediated transformation. Two transgenic lines with a single copy gene in T0 were selected and evaluated in both the T1 and T4 generations. Transgenic lines had drastically improved survival rate under water deficit conditions, at rates close to 100%, while WT did not survive. Besides, transgenic lines had improved biomass production and higher tillering under water deficit conditions compared with WT plants. Reduced pectin and hemicellulose contents were observed in transgenic lines compared with wild-type plants under both well-watered and water deficit conditions, whereas cellulose content was unchanged in line #17 and reduced in line #29 under conditions of low water availability. Changes in lignin content under water deficit were only observed in line #17. However, improvements in saccharification were found in both transgenic lines along with changes in the expression of OsNTS1/2 and OsMYB58/63 secondary cell wall biosynthesis genes. ShDJ-overexpression up-regulated the expression of the OsbZIP23, OsGRAS23, OsP5CS, and OsLea3 genes in rice stems under well-watered conditions. Taken together, our data suggest that ShDJ has the potential for improving drought tolerance, plant biomass accumulation, and saccharification efficiency

Sugarcane (Saccharum X officinarum): A Reference Study for the Regulation of Genetically Modified Cultivars in Brazil

Global interest in sugarcane has increased significantly in recent years due to its economic impact on sustainable energy production. Sugarcane breeding and better agronomic practices have contributed to a huge increase in sugarcane yield in the last 30 years. Additional increases in sugarcane yield are expected to result from the use of biotechnology tools in the near future. Genetically modified (GM) sugarcane that incorporates genes to increase resistance to biotic and abiotic stresses could play a major role in achieving this goal. However, to bring GM sugarcane to the market, it is necessary to follow a regulatory process that will evaluate the environmental and health impacts of this crop. The regulatory review process is usually accomplished through a comparison of the biology and composition of the GM cultivar and a non-GM counterpart. This review intends to provide information on non-GM sugarcane biology, genetics, breeding, agronomic management, processing, products and byproducts, as well as the current technologies used to develop GM sugarcane, with the aim of assisting regulators in the decision-making process regarding the commercial release of GM sugarcane cultivars

Evaluation of genetic variability in musa spp. using microsattelite markers.

Em todo o mundo, a cultura da banana tem enfrentado uma série de problemas relacionados a ocorrência de doenças e pragas, para as quais, a obtenção de cultivares resistentes é a forma mais viável de controle. Híbridos tetraplóides promissores, obtidos a partir do cruzamento de cultivares triplóides com diplóides cultivados, selvagens ou melhorados, têm sido produzidos pelos diferentes programas de melhoramento. No entanto, o sucesso num programa de melhoramento depende inicialmente do completo conhecimento da diversidade genética do germoplasma disponível. Os marcadores microssatélites constituem-se em ferramentas valiosas para este fim. Este trabalho objetivou caracterizar, por marcadores microssatélites, 84 genótipos de Musa, a partir do estabelecimento de uma metodologia de detecção do polimorfismo em géis de seqüenciamento, por meio da coloração com prata. Os genótipos foram analisados em dois experimentos distintos. No primeiro experimento, as relações genéticas existentes entre 35 genótipos, compreendendo cultivares triplóides, raças locais (landraces) e híbridos tetraplóides foram investigadas empregando-se 11 primers SSRs. A análise fenética obtida mostrou concordância com a caracterização baseada em descritores morfológicos. Os genótipos agruparam-se com base na composição genômica, grupo genômico e subgrupo. Alguns híbridos tetraplóides apresentaram distorções na proporção de alelos doados pelo genitor feminino triplóide, o que suporta constatações recentes sobre a ocorrência de recombinação durante a formação dos gametas femininos. No segundo experimento, nove primers SSRs foram empregados na caracterização de 49 genótipos diplóides, divididos em três 'subgrupos' (cultivados, selvagens e melhorados) e na determinação das relações genéticas existentes entre estes materiais e nove cultivares comerciais triplóides. A análise fenética conduzida sobre todos os genótipos, não evidenciou uma perfeita separação dos genótipos diplóides em seus respectivos subgrupos, possivelmente devido a existência de muitos alelos comuns a eles. A estatística Rst confirmou este resultado. Alguns genótipos diplóides exibiram uma forte relação com as cultivares triplóides AAA tipo exportação. Os marcadores microssatélites mostraram-se altamente eficientes na caracterização e discriminação do germoplasma de Musa, gerando fingerprinting únicos para um grande número de genótipos. Um grande número de alelos pôde ser detectado, o que comprovou a natureza multialélica deste tipo de marcador. Os genótipos diplóides foram os que apresentaram a maior diversidade, refletida pelo maior número de alelos detectados e pela baixa similaridade entre os clones. Nos dois experimentos, observou-se uma alta proporção de alelos "multiplex", bem como locos duplicados, o que resultou na perda do caráter codominante dos marcadores SSRs. As implicações decorrentes destas observações foram consideradas. A metodologia de coloração com prata apresentada mostrou ser um procedimento eficiente, rápido, simples e de baixo custo na detecção do polimorfismo SSRs.The banana (Musa) industry has faced various disease and pest problems all over the world, and the development of resistant cultivars is the most attractive way of control. Promising tetraploid hybrids, derived from crosses between triploid cultivars and wild, cultivated or selected diploids, have been developed from different breeding programs. However, the success in a breeding program depends on the knowledge about the genetic diversity of the available germplasm. Microsattelite is an important tool for estimating the genetic diversity. This work aimed to characterize 84 Musa genotypes using microsattelite markers, based on a method for polymorphism detection with electrophoresis on sequencing gel stained with silver nitrate. The genotypes were analyzed in two experiments. In the first one, the genetic relationships between 35 genotypes, including triploid cultivars, landraces and tetraploid hybrids, were investigated using 11 microsattelite primers. The phenetic analysis disclosed a grouping in agreement with the characterization based on morphological descriptors. The genotypes clustered according to genomic composition, genomic group and subgroup. Some tetraploid hybrids presented distortion of the proportion of alleles donated by the female triploid genitor, in support to recent description about the occurrence of recombination during female gamete formation. In the second experiment, 9 microsattelite primers were used to characterize 49 diploid genotypes, divided into three subgroups (cultivated, wild and selected) and to determine the genetic relationships between these genotypes and 9 commercial triploid cultivars. The phenetic analysis of all the genotypes did not demonstrate a separation of the diploid genotypes into the respective subgroups, possibly because of the occurrence of various alleles in common The statistic Rst confirmed this result. Some diploid genotypes showed a strong relationship with export-type triploid AAA cultivars. Microsattelite markers were shown to be highly efficient for Musa germplasm characterization and discrimination, generating unique fingerprinting for a large number of genotypes. A large number of alleles was detected, demonstrating the multi-allelic nature of this marker. The diploid genotypes presented the largest diversity, reflected by the largest number of detected alleles and by the low similarity between clones. In both experiments, a large proportion of multiplex alleles was, as well as duplicated loci, resulting in the loss of the codominant character of the marker. The resulting implications were considered. The methods of silver staining showed to be a quick, simple, efficient, and low-cost procedure to detect SSR polymorphism

Drought-triggered leaf transcriptional responses disclose key molecular pathways underlying leaf water use efficiency in sugarcane (Saccharum spp.)

Drought is a major constraint to sugarcane (Saccharum spp.) production and improving the water use efficiency (WUE) is a critical trait for the sustainability of this bioenergy crop. The molecular mechanism underlying WUE remains underexplored in sugarcane. Here, we investigated the drought-triggered physiological and transcriptional responses of two sugarcane cultivars contrasting for drought tolerance, ‘IACSP97-7065’ (sensitive) and ‘IACSP94-2094’ (tolerant). After 21 days without irrigation (DWI), only ‘IACSP94-2094’ exhibited superior WUE and instantaneous carboxylation efficiency, with the net CO2 assimilation being less impacted when compared with ‘IACSP97-7065’. RNA-seq of sugarcane leaves at 21 DWI revealed a total of 1,585 differentially expressed genes (DEGs) for both genotypes, among which ‘IACSP94-2094’ showed 617 (38.9%) exclusive transcripts (212 up- and 405 down-regulated). Functional enrichment analyses of these unique DEGs revealed several relevant biological processes, such as photosynthesis, transcription factors, signal transduction, solute transport, and redox homeostasis. The better drought-responsiveness of ‘IACSP94-2094’ suggested signaling cascades that foster transcriptional regulation of genes implicated in the Calvin cycle and transport of water and carbon dioxide, which are expected to support the high WUE and carboxylation efficiency observed for this genotype under water deficit. Moreover, the robust antioxidant system of the drought-tolerant genotype might serve as a molecular shield against the drought-associated overproduction of reactive oxygen species. This study provides relevant data that may be used to develop novel strategies for sugarcane breeding programs and to understand the genetic basis of drought tolerance and WUE improvement of sugarcane

Metabolite Profiling of Sugarcane Genotypes and Identification of Flavonoid Glycosides and Phenolic Acids

Sugarcane is an important agricultural crop in the economy of tropical regions, and Brazil has the largest cultivated acreage in the world. Sugarcane accumulates high levels of sucrose in its stalks. Other compounds produced by sugarcane are currently not of economic importance. To explore potential coproducts, we have studied the chemical diversity of sugarcane genotypes, via metabolite profiling of leaves by NMR and LC-DAD-MS. Metabolites were identified via in-house and public databases. From the analysis of 60 HPLC-fractionated extracts, LC-DAD-MS detected 144 metabolites, of which 56 were identified (MS-MS and 1H NMR), including 19 phenolics and 25 flavones, with a predominance of isomeric flavone C-glycosides. Multivariate analysis of the profiles from genotypes utilized in Brazilian breeding programs revealed clustering according to sugar, phenolic acid, and flavone contents

Genetic variability among sugarcane genotypes based on polymorphisms in sucrose metabolism and drought tolerance genes

Target region amplification polymorphism (TRAP) markers were used to estimate the genetic similarity (GS) among 53 sugarcane varieties and five species of the Saccharum complex. Seven fixed primers designed from candidate genes involved in sucrose metabolism and three from those involved in drought response metabolism were used in combination with three arbitrary primers. The clustering of the genotypes for sucrose metabolism and drought response were similar, but the GS based on Jaccard`s coefficient changed. The GS based on polymorphism in sucrose genes estimated in a set of 46 Brazilian varieties, all of which belong to the three Brazilian breeding programs, ranged from 0.52 to 0.9, and that based on drought data ranged from 0.44 to 0.95. The results suggest that genetic variability in the evaluated genes was lower in the sucrose metabolism genes than in the drought response metabolism ones

Molecular variability and genetic relationship among Brazilian strains of the sugarcane smut fungus

Sporisorium scitamineum is the fungus that causes sugarcane smut disease. Despite of the importance of sugarcane for Brazilian agribusiness and the persistence of the pathogen in most cropping areas, genetic variation studies are still missing for Brazilian isolates. In this study, sets of isolates were analyzed using two molecular markers (AFLP and telRFLP) and ITS sequencing. Twenty-two whips were collected from symptomatic plants in cultivated sugarcane fields of Brazil. A total of 41 haploid strains of compatible mating types were selected from individual teliospores and used for molecular genetic analyses. telRFLP and ITS analyses were expanded to six Argentine isolates, where the sugarcane smut was first recorded in America. Genetic relationship among strains suggests the human-mediated dispersal of S. scitamineum within the Brazilian territory and between the two neighboring countries. Two genetically distinct groups were defined by the combined analysis of AFLP and telRFLP. The opposite mating-type strains derived from single teliospores were clustered together into these main groups, but had not always identical haplotypes. telRFLP markers analyzed over two generations of selfing and controlled outcrossing confirmed the potential for emergence of new variants and occurrence of recombination, which are relevant events for evolution of virulence and environmental adaptation.Fil: Benevenuto, Juliana. Universidade de Sao Paulo; BrasilFil: Longatto, Daniel P.. Universidade de Sao Paulo; BrasilFil: Reis, Gislaine V.. Universidade de Sao Paulo; BrasilFil: Mielnichuk, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Palhares, Alessandra C.. Universidade de Sao Paulo; BrasilFil: Carvalho, Giselle. Universidade de Sao Paulo; BrasilFil: Saito, Suzane. Universidade de Sao Paulo; BrasilFil: Quecine, Maria C.. Universidade de Sao Paulo; BrasilFil: Sanguino, Alvaro. Agencia Paulista de Tecnologia Dos Agronegocios; BrasilFil: Vieira, Maria Lucia C.. Universidade de Sao Paulo; BrasilFil: Camargo, Luis Eduardo A.. Universidade de Sao Paulo; BrasilFil: Creste, Silvana. Agencia Paulista de Tecnologia Dos Agronegocios; BrasilFil: Monteiro-Vitorello, Claudia B.. Universidade de Sao Paulo; Brasi

Lignin biosynthesis in sugarcane is affected by low temperature

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Sugarcane is an abundant and promising source of plant biomass for biofuel production. The use of biomass for conversion to ethanol is limited by the recalcitrance of lignocellulosic material mainly due to the presence of lignin. This study characterised some aspects of lignification in the stem of two sugarcane genotypes grown under low (Cl') and warmer (HT - reference) temperatures. Stems were separated in young and mature culms and the culms were separated in cortex (rind) and medulla (pith). Plants of the genotype IACSP04-627, which has more lignin in the stem, grew better (fresh weight of the stem) than IACSP04-065 under HT. Cold negatively affected plant growth but apparently IACSPO4-065 was more sensitive than the other genotype. Lignin content was significantly increased in the young rind of IACSP04-627 plants at CT, what could not be directly correlated with the expression profile of genes of the monolignols biosynthesis. Lignin content in mature rind was reduced in IACSP04-065 plants exposed to CT, what could be correlated with the low expression level of the genes ShCAD2, ShCOMT1 and ShCCoAOMT1. The high expression of ShF5H occurred preferentially in mature pith of both sugarcane genotypes, what is possibly related with the early formation of the secondary cell wall induced by low temperature. In conclusion, lignin deposition in sugarcane under low temperature seems to be differentially regulated in rind and pith tissues and it is genotype-dependent. (C) 2015 Elsevier B.V. All rights reserved.Sugarcane is an abundant and promising source of plant biomass for biofuel production. The use of biomass for conversion to ethanol is limited by the recalcitrance of lignocellulosic material mainly due to the presence of lignin. This study characterised1203142FAPESP - FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULOCAPES - COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIORCNQP - CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2008/58035-6]2008/58035-6SEM INFORMAÇÃOSEM INFORMAÇÃOThis study was funded by Fundac ̧ão de Amparo à Pesquisa doEstado de São Paulo (FAPESP – grant 2008/58035-6). The authors thank Coordenac ̧ão de Aperfeic ̧oamento de Pessoal de Nível Supe-rior (Capes – Brazil) and Conselho Nacional de DesenvolvimentoCien