Transcriptomic analysis of Chaetoceros muelleri in response to different nitrogen concentrations reveals the activation of pathways to enable efficient nitrogen uptake.

Nitrogen is the principal nutrient deficiency that increases lipids and carbohydrate content in diatoms but negatively affects biomass production. Marine diatom Chaetoceros muelleri is characterized by lipid and carbohydrate accumulation under low nitrogen concentration without affecting biomass. To elucidate the molecular effects of nitrogen concentrations, we performed an RNA-seq analysis of C. muelleri grown under four nitrogen concentrations (3.53 mM, 1.76 mM, 0.44 mM, and 0.18 mM of NaNO3). This research revealed that changes in global transcription in C. muelleri are differentially expressed by nitrogen concentration. "Energetic metabolism", "Carbohydrate metabolism" and "Lipid metabolism" pathways were identified as the most upregulated by N deficiency. Due to N limitation, alternative pathways to self-supply nitrogen employed by microalgal cells were identified. Additionally, nitrogen limitation decreased chlorophyll content and caused a greater response at the transcriptional level with a higher number of unigenes differentially expressed. By contrast, the highest N concentration (3.53 mM) recorded the lowest number of differentially expressed genes. Amt1, Nrt2, Fad2, Skn7, Wrky19, and Dgat2 genes were evaluated by RT-qPCR. In conclusion, C. muelleri modify their metabolic pathways to optimize nitrogen utilization and minimize nitrogen losses. On the other hand, the assembled transcriptome serves as the basis for metabolic engineering focused on improving the quantity and quality of the diatom for biotechnological applications. However, proteomic and metabolomic analysis is also required to compare gene expression, protein, and metabolite accumulation.

Large scale sampling of Mexican maize landraces for the presence of transgenes.

The presence and levels of transgenic maize in Mexico and the effect this could have on local landraces or closely related species such as teosinte has been the subject of several previous reports, some showing contrasting results. Cultural, social and political factors all affect maize cultivation in Mexico and although since 1998 there has been a moratorium on the commercial cultivation of transgenic maize, Mexico imports maize, mainly from the USA where transgenic cultivars are widely grown. Additionally extensive migration between rural areas in Mexico and the USA and customs of seed exchange between farmers may also play an unintentional role in the establishment of transgenic seed. A comprehensive study of all Mexican maize landraces throughout the country is not feasible, however this report presents data based on analysis of 3204 maize accessions obtained from the central region of Mexico (where permits have never been authorized for cultivation of transgenic maize) and the northern region (where for a short period authorization for experimental plots was granted). The results of the study confirm that transgenes are present in all the geographical areas sampled and were more common in germplasm obtained in the northern region. However, there was no evidence that regions where field trials had been authorized showed higher levels of transgene presence or that the morphology of seed lots harboring transgenic material was significantly modified in favor of expected transgenic phenotypes.

Gene Functional Networks from Time Expression Profiles: A Constructive Approach Demonstrated in Chili Pepper (Capsicum annuum L.).

Gene co-expression networks are powerful tools to understand functional interactions between genes. However, large co-expression networks are difficult to interpret and do not guarantee that the relations found will be true for different genotypes. Statistically verified time expression profiles give information about significant changes in expressions through time, and genes with highly correlated time expression profiles, which are annotated in the same biological process, are likely to be functionally connected. A method to obtain robust networks of functionally related genes will be useful to understand the complexity of the transcriptome, leading to biologically relevant insights. We present an algorithm to construct gene functional networks for genes annotated in a given biological process or other aspects of interest. We assume that there are genome-wide time expression profiles for a set of representative genotypes of the species of interest. The method is based on the correlation of time expression profiles, bound by a set of thresholds that assure both, a given false discovery rate, and the discard of correlation outliers. The novelty of the method consists in that a gene expression relation must be repeatedly found in a given set of independent genotypes to be considered valid. This automatically discards relations particular to specific genotypes, assuring a network robustness, which can be set a priori. Additionally, we present an algorithm to find transcription factors candidates for regulating hub genes within a network. The algorithms are demonstrated with data from a large experiment studying gene expression during the development of the fruit in a diverse set of chili pepper genotypes. The algorithm is implemented and demonstrated in a new version of the publicly available R package "Salsa" (version 1.0).

Transcriptomic analysis of a wild and a cultivated varieties of Capsicum annuum over fruit development and ripening.

Chili pepper (Capsicum annuum) is one of the most important crops worldwide. Its fruits contain metabolites produced over the maturation process like capsaicinoids and carotenoids. This metabolic process produces internal changes in flavor, color, texture, and aroma in fruits to make them more attractive for seed dispersal organisms. The chiltepin (C. annuum L. var. glabriusculum) is a wild variety of the C. annuum L. species that is considered a source of genetic resources that could be used to improve the current chili crops. In this study, we performed a transcriptomic analysis on two fruit maturation stages: immature stage (green fruit) and mature stage (red fruit) of a wild and a cultivated pepper variety. We found 19,811 genes expressed, and 1,008 genes differentially expressed (DEGs) in at least one of the five contrast used; 730 DEGs were found only in one contrast, and most DEGs in all contrasts were downregulated. GO enrichment analysis showed that the majority of DEGs are related to stress responses. KEGG enrichment analysis detected differences in expression patterns in metabolic pathways related to phenylpropanoid biosynthesis, secondary metabolites, plant hormone signal transduction, carotenoid biosynthesis and sesquiterpenoid and triterpenoid biosynthesis. We selected 105 tomato fruit ripening-related genes, and found 53 pepper homologs differentially expressed related to shape, size, and secondary metabolite biosynthesis. According to the transcriptome analysis, the two peppers showed very similar gene expression patterns; differences in expression patterns of genes related to shape, size, ethylene and secondary metabolites biosynthesis suggest that changes produced by domestication of chilli pepper could be very specific to the expression of genes related to traits desired in commercial fruits.

Biochemical and Molecular Aspects of Phosphorus Limitation in Diatoms and Their Relationship with Biomolecule Accumulation.

Diatoms are the most abundant group of phytoplankton, and their success lies in their significant adaptation ability to stress conditions, such as nutrient limitation. Phosphorus (P) is a key nutrient involved in the transfer of energy and the synthesis of several cellular components. Molecular and biochemical mechanisms related to how diatoms cope with P deficiency are not clear, and research into this has been limited to a few species. Among the molecular responses that have been reported in diatoms cultured under P deficient conditions is the upregulation of genes encoding enzymes related to the transport, assimilation, remobilization and recycling of this nutrient. Regarding biochemical responses, due to the reduction of the requirements for carbon structures for the synthesis of proteins and phospholipids, more CO2 is fixed than is consumed by the Calvin cycle. To deal with this excess, diatoms redirect the carbon flow toward the synthesis of storage compounds such as triacylglycerides and carbohydrates, which are excreted as extracellular polymeric substances. This review aimed to gather all current knowledge regarding the biochemical and molecular mechanisms of diatoms related to managing P deficiency in order to provide a wider insight into and understanding of their responses, as well as the metabolic pathways affected by the limitation of this nutrient.

Transcriptome Analyses Throughout Chili Pepper Fruit Development Reveal Novel Insights into the Domestication Process.

Chili pepper (Capsicum spp.) is an important crop, as well as a model for fruit development studies and domestication. Here, we performed a time-course experiment to estimate standardized gene expression profiles with respect to fruit development for six domesticated and four wild chili pepper ancestors. We sampled the transcriptomes every 10 days from flowering to fruit maturity, and found that the mean standardized expression profiles for domesticated and wild accessions significantly differed. The mean standardized expression was higher and peaked earlier for domesticated vs. wild genotypes, particularly for genes involved in the cell cycle that ultimately control fruit size. We postulate that these gene expression changes are driven by selection pressures during domestication and show a robust network of cell cycle genes with a time shift in expression, which explains some of the differences between domesticated and wild phenotypes.

Metagenomics of mine tailing rhizospheric communities and its selection for plant establishment towards bioremediation.

Mining operations often generate tailing dams that contain toxic residues and are a source of contamination when left unconfined. The establishment of a plant community over the tailings has been proposed as a containment strategy known as phytostabilization. Previously, we described naturally occurring mine tailing colonizing plants such as Acacia farnesiana, Brickellia coulteri, Baccharis sarothroides, and Gnaphalium leucocephalum without finding local adaptation. We explored the rhizosphere microbes as contributors in plant establishment and described both the culturable and in situ diversity of rhizospheric bacteria using the 16S rRNA gene and metagenomic shotgun sequencing. We built a synthetic community (SC) of culturable rhizosphere bacteria from the mine tailings. The SC was then the foundation for a serial passes experiment grown in plant-derived nutrient sources, selecting for heavy metals tolerance, community cooperation, and competition. The outcome of the serial passes was named the 'final synthetic community' (FSC). Overall, diversity decreased from in situ uncultivable microbes from roots (399 bacteria genera) to the cultivated communities (291 genera), the SC (94 genera), and the lowest diversity was in the FSC (43 genera). Metagenomic diversity clustered into 94,245 protein families, where we found plant growth promotion-related genes such as the csgBAC and entCEBAH, coded in a metagenome-assembled genome named Kosakonia sp. Nacozari. Finally, we used the FSC to inoculate mine tailing colonizing plants in a greenhouse experiment. The plants with the FSC inocula observed higher relative plant growth rates in sterile substrates. The FSC presents promising features that might make it useful for phytostabilization tailored strategies.

Determination of Chemical Constituents and Antioxidant Activities of Leaves and Stems from Jatropha cinerea (Ortega) Müll. Arg and Jatropha cordata (Ortega) Müll. Arg.

Jatropha species have been shown to be an important source of secondary metabolites with different biological effects. Jatropha cinerea (Ortega) Müll. Arg and Jatropha cordata (Ortega) Müll. Arg are distributed in the Northwestern region of Mexico, are adapted to extreme weather conditions and are widely used (stems, leaves, and sap) in traditional medicine. The aim of the present study was to carry out the phytochemical characterization and the evaluation of the antioxidant activity in methanolic extracts of stems and leaves from J. cinerea and J. cordata. The compounds present in the extracts of both species were characterized by ESI-IT-MS/MS and quantified by HPLC-DAD. The results showed that the stem extracts of both species are rich in phenolic acids, while the leaf extracts are rich in flavonoids. Some of the main compounds found were gallic acid, gentisic acid, 3,4-Dihydroxybenzoic acid, vitexin, isovitexin, and catechol. Both species showed high concentrations of phenols and total flavonoids and antioxidant activity. J.cordata showed the highest antioxidant capacity and the highest concentration of phenolic compounds. Overall, both Jatropha species are a natural source of antioxidant compounds with potential biotechnological uses.

Compacta: a fast contig clustering tool for de novo assembled transcriptomes.

BACKGROUND: RNA-Seq is the preferred method to explore transcriptomes and to estimate differential gene expression. When an organism has a well-characterized and annotated genome, reads obtained from RNA-Seq experiments can be directly mapped to that genome to estimate the number of transcripts present and relative expression levels of these transcripts. However, for unknown genomes, de novo assembly of RNA-Seq reads must be performed to generate a set of contigs that represents the transcriptome. These contig sets contain multiple transcripts, including immature mRNAs, spliced transcripts and allele variants, as well as products of close paralogs or gene families that can be difficult to distinguish. Thus, tools are needed to select a set of less redundant contigs to represent the transcriptome for downstream analyses. Here we describe the development of Compacta to produce contig sets from de novo assemblies. RESULTS: Compacta is a fast and flexible computational tool that allows selection of a representative set of contigs from de novo assemblies. Using a graph-based algorithm, Compacta groups contigs into clusters based on the proportion of shared reads. The user can determine the minimum coverage of the contigs to be clustered, as well as a threshold for the proportion of shared reads in the clustered contigs, thus providing a dynamic range of transcriptome compression that can be adapted according to experimental aims. We compared the performance of Compacta against state of the art clustering algorithms on assemblies from Arabidopsis, mouse and mango, and found that Compacta yielded more rapid results and had competitive precision and recall ratios. We describe and demonstrate a pipeline to tailor Compacta parameters to specific experimental aims. CONCLUSIONS: Compacta is a fast and flexible algorithm for the determination of optimum contig sets that represent the transcriptome for downstream analyses.

Simulated Gastrointestinal Digestion, Bioaccessibility and Antioxidant Capacity of Polyphenols from Red Chiltepin (Capsicum annuum L. Var. glabriusculum) Grown in Northwest Mexico.

Chiltepin, a wild chili mostly used in different traditional foods and traditional medicine in Northwest Mexico, represents a source of polyphenols. However, studies about the bioaccessibility of polyphenols as a parameter to measure the nutritional quality and bioefficacy of them in the fruit after consumption are scarce. Chiltepin showed phenolic acids and flavonoids contents between 387 and 65 µg/g, respectively. Nevertheless, these values decreased after the digestion process. Before digestion, gallic acid, 4-hydroxibenzoinc acid, chlorogenic acid, caffeic acid, p-coumaric acid, quercetin and luteolin were the main polyphenols found in chiltepin by HPLC-DAD and confirmed by FIA-ESI-IT-MS/MS. Gallic and chlorogenic acids were non-detected in the gastric phase, while only p-coumaric acid (5.35 ± 3.89 µg/g), quercetin (5.91 ± 0.92 µg/g) and luteolin (2.86 ± 0.62 µg/g) were found in the intestinal phase. The bioaccessibility of phenolic acids, flavonoids, and total polyphenols after the intestinal phase was around 24, 17 and 23%, respectively. Overall, results indicated that release of polyphenols from chiltepin fruit might be affected by the food matrix and gastrointestinal conditions due to the low bioaccessibility values observed.

Expression analysis of genes involved in the synthesis of oleic and linoleic acids in Jatropha cinerea seeds from Northwestern Mexico / Análise de expressão de genes envolvidos na síntese de ácidos oleico e linoleico em sementes de Jatropha cinerea do noroeste do México

ABSTRACT: Plants belonging to genus Jatropha has arisen interest because of its high oil content that could be used to produce biodiesel. It is also widely reported that the main fatty acids in Jatropha oilseed are oleic and linoleic acids. However, there are scarce studies related to native species of Jatropha from Northwestern Mexico which are adapted to arid conditions, and the expression of genes involved in fatty acid synthesis for these species is still unknown. Therefore, the aim of this study was to analyze the expression of five genes, ACP1, KASII, D9SD, FAD2-1 and FAD2-2, which are involved in the oleic and linoleic acids synthesis in mature wild-seeds of Jatropha cinerea, a native species from Sonoran Desert, using semi-quantitative RT-PCR. The J. cinerea seeds were randomly collected in Bahía de Kino, Sonora (México) which is a region characterized by its harsh environments such as saline soils and extreme temperature changes and J. curcas mature seeds from a non-toxic variety from Veracruz, Mexico were used as a reference. The RT-PCR analysis of three biological replicates were considered to ensure data consistent. Our analysis showed a higher expression of KASII and FAD2-1 genes in J. cinerea seeds compared to J. curcas, meanwhile the expression of ACP1, D9SD and FAD2-2 were higher in J. curcas. Furthermore, Actin and FAD2-1 genes sequences here obtained are the first reported for J. cinerea, thus providing information to develop further studies.

RESUMO: Plantas pertencentes ao gênero Jatropha têm despertado interesse devido ao seu alto teor de óleo que poderia ser usado para produzir biodiesel. Também é amplamente relatado que os principais ácidos graxos das oleaginosas da Jatropha são os ácidos oléico e linoleico. No entanto, existem poucos estudos relacionados a espécies nativas de pinhão-manso do noroeste do México que estão adaptados a condições áridas, e a expressão de genes envolvidos na síntese de ácidos graxos para essas espécies ainda é desconhecida. Portanto, o objetivo deste estudo foi analisar a expressão de cinco genes, ACP1, KASII, D9SD, FAD2-1 e FAD2-2, que estão envolvidos na síntese de ácidos oleico e linoleico em sementes selvagens maduras de Jatropha cinerea, espécies nativas do Deserto de Sonora, usando RT-PCR semi-quantitativa. Sementes de J. cinerea foram coletadas aleatoriamente em Bahía de Kino, Sonora (México), região caracterizada por ambientes agressivos como solos salinos e mudanças extremas de temperatura, e sementes de J. curcas maduras de uma variedade não-tóxica de Veracruz, México, usado como referência. Análise RT-PCR de três repetições biológicas foram consideradas para garantir dados consistentes. Nossa análise mostrou uma maior expressão dos genes KASII e FAD2-1 em sementes de J. cinerea comparado a J. curcas, enquanto a expressão de ACP1, D9SD e FAD2-2 foi maior em J. curcas. Além disso, as sequências dos genes Actin e FAD2-1 obtidas são as primeiras relatadas para J. cinerea, fornecendo, assim, informações para o desenvolvimento de novos estudos.

An SSR-based approach incorporating a novel algorithm for identification of rare maize genotypes facilitates criteria for landrace conservation in Mexico.

As maize was domesticated in Mexico around 9,000 years ago, local farmers have selected and maintained seed stocks with particular traits and adapted to local conditions. In the present day, many of these landraces are still cultivated; however, increased urbanization and migration from rural areas implies a risk that this invaluable maize germplasm may be lost. In order to implement an efficient mechanism of conservation in situ, the diversity of these landrace populations must be estimated. Development of a method to select the minimum number of samples that would include the maximum number of alleles and identify germplasm harboring rare combinations of particular alleles will also safeguard the efficient ex-situ conservation of this germplasm. To reach this goal, a strategy based on SSR analysis and a novel algorithm to define a minimum collection and rare genotypes using landrace populations from Puebla State, Mexico, was developed as a "proof of concept" for methodology that could be extended to all maize landrace populations in Mexico and eventually to other native crops. The SSR-based strategy using bulked DNA samples allows rapid processing of large numbers of samples and can be set up in most laboratories equipped for basic molecular biology. Therefore, continuous monitoring of landrace populations locally could easily be carried out. This methodology can now be applied to support incentives for small farmers for the in situ conservation of these traditional cultivars.

Regulatory network analysis reveals novel regulators of seed desiccation tolerance in Arabidopsis thaliana.

Desiccation tolerance (DT) is a remarkable process that allows seeds in the dry state to remain viable for long periods of time that in some instances exceed 1,000 y. It has been postulated that seed DT evolved by rewiring the regulatory and signaling networks that controlled vegetative DT, which itself emerged as a crucial adaptive trait of early land plants. Understanding the networks that regulate seed desiccation tolerance in model plant systems would provide the tools to understand an evolutionary process that played a crucial role in the diversification of flowering plants. In this work, we used an integrated approach that included genomics, bioinformatics, metabolomics, and molecular genetics to identify and validate molecular networks that control the acquisition of DT in Arabidopsis seeds. Two DT-specific transcriptional subnetworks were identified related to storage of reserve compounds and cellular protection mechanisms that act downstream of the embryo development master regulators LEAFY COTYLEDON 1 and 2, FUSCA 3, and ABSCICIC ACID INSENSITIVE 3. Among the transcription factors identified as major nodes in the DT regulatory subnetworks, PLATZ1, PLATZ2, and AGL67 were confirmed by knockout mutants and overexpression in a desiccation-intolerant mutant background to play an important role in seed DT. Additionally, we found that constitutive expression of PLATZ1 in WT plants confers partial DT in vegetative tissues.

Methylome analysis reveals an important role for epigenetic changes in the regulation of the Arabidopsis response to phosphate starvation.

Phosphate (Pi) availability is a significant limiting factor for plant growth and productivity in both natural and agricultural systems. To cope with such limiting conditions, plants have evolved a myriad of developmental and biochemical strategies to enhance the efficiency of Pi acquisition and assimilation to avoid nutrient starvation. In the past decade, these responses have been studied in detail at the level of gene expression; however, the possible epigenetic components modulating plant Pi starvation responses have not been thoroughly investigated. Here, we report that an extensive remodeling of global DNA methylation occurs in Arabidopsis plants exposed to low Pi availability, and in many instances, this effect is related to changes in gene expression. Modifications in methylation patterns within genic regions were often associated with transcriptional activation or repression, revealing the important role of dynamic methylation changes in modulating the expression of genes in response to Pi starvation. Moreover, Arabidopsis mutants affected in DNA methylation showed that changes in DNA methylation patterns are required for the accurate regulation of a number of Pi-starvation-responsive genes and that DNA methylation is necessary to establish proper morphological and physiological phosphate starvation responses.

Analysis and optimization of bulk DNA sampling with binary scoring for germplasm characterization.

The strategy of bulk DNA sampling has been a valuable method for studying large numbers of individuals through genetic markers. The application of this strategy for discrimination among germplasm sources was analyzed through information theory, considering the case of polymorphic alleles scored binarily for their presence or absence in DNA pools. We defined the informativeness of a set of marker loci in bulks as the mutual information between genotype and population identity, composed by two terms: diversity and noise. The first term is the entropy of bulk genotypes, whereas the noise term is measured through the conditional entropy of bulk genotypes given germplasm sources. Thus, optimizing marker information implies increasing diversity and reducing noise. Simple formulas were devised to estimate marker information per allele from a set of estimated allele frequencies across populations. As an example, they allowed optimization of bulk size for SSR genotyping in maize, from allele frequencies estimated in a sample of 56 maize populations. It was found that a sample of 30 plants from a random mating population is adequate for maize germplasm SSR characterization. We analyzed the use of divided bulks to overcome the allele dilution problem in DNA pools, and concluded that samples of 30 plants divided into three bulks of 10 plants are efficient to characterize maize germplasm sources through SSR with a good control of the dilution problem. We estimated the informativeness of 30 SSR loci from the estimated allele frequencies in maize populations, and found a wide variation of marker informativeness, which positively correlated with the number of alleles per locus.

First draft genome sequence of a strain from the genus Citricoccus.

Bacteria of the genus Citricoccus have been isolated from ecological niches characterized by diverse abiotic stress conditions. Here we report the first genome draft of a strain of the genus Citricoccus isolated from the extremely oligotrophic Churince system in the Cuatro Ciénegas Basin (CCB) in Coahuila, Mexico.

Analysis of gene expression and physiological responses in three Mexican maize landraces under drought stress and recovery irrigation.

BACKGROUND: Drought is one of the major constraints for plant productivity worldwide. Different mechanisms of drought-tolerance have been reported for several plant species including maize. However, the differences in global gene expression between drought-tolerant and susceptible genotypes and their relationship to physiological adaptations to drought are largely unknown. The study of the differences in global gene expression between tolerant and susceptible genotypes could provide important information to design more efficient breeding programs to produce maize varieties better adapted to water limiting conditions. METHODOLOGY/PRINCIPAL FINDINGS: Changes in physiological responses and gene expression patterns were studied under drought stress and recovery in three Mexican maize landraces which included two drought tolerant (Cajete criollo and Michoacán 21) and one susceptible (85-2) genotypes. Photosynthesis, stomatal conductance, soil and leaf water potentials were monitored throughout the experiment and microarray analysis was carried out on transcripts obtained at 10 and 17 days following application of stress and after recovery irrigation. The two tolerant genotypes show more drastic changes in global gene expression which correlate with different physiological mechanisms of adaptation to drought. Differences in the kinetics and number of up- and down-regulated genes were observed between the tolerant and susceptible maize genotypes, as well as differences between the two tolerant genotypes. Interestingly, the most dramatic differences between the tolerant and susceptible genotypes were observed during recovery irrigation, suggesting that the tolerant genotypes activate mechanisms that allow more efficient recovery after a severe drought. CONCLUSIONS/SIGNIFICANCE: A correlation between levels of photosynthesis and transcription under stress was observed and differences in the number, type and expression levels of transcription factor families were also identified under drought and recovery between the three maize landraces. Gene expression analysis suggests that the drought tolerant landraces have a greater capacity to rapidly modulate more genes under drought and recovery in comparison to the susceptible landrace. Modulation of a greater number of differentially expressed genes of different TF gene families is an important characteristic of the tolerant genotypes. Finally, important differences were also noted between the tolerant landraces that underlie different mechanisms of achieving tolerance.