Plant Genomics
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Organ-specific alterations in tobacco transcriptome caused by the PVX-derived P25 silencing suppressor transgene

RNA silencing affects a broad range of regulatory processes in all eukaryotes ranging from chromatin structure maintenance to transcriptional and translational regulation and longevity of the mRNAs.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

RNA silencing affects a broad range of regulatory processes in all eukaryotes ranging from chromatin structure maintenance to transcriptional and translational regulation and longevity of the mRNAs. Particularly in plants, it functions as the major defense mechanism against viruses. To counter-act this defense, plant viruses produce suppressors of RNA silencing (Viral suppressors of RNA silencing, VSRSs), which are essential for viruses to invade their specific host plants. Interactions of these VSRSs with the hosts' silencing pathways, and their direct and indirect interference with different cellular regulatory networks constitute one of the main lines of the molecular virus-host interactions. Here we have used a microarray approach to study the effects of the Potato virus X Potexvirus (PVX)-specific P25 VSRS protein on the transcript profile of tobacco plants, when expressed as a transgene in these plants.

Results

The expression of the PVX-specific P25 silencing suppressor in transgenic tobacco plants caused significant up-regulation of 1350 transcripts, but down-regulation of only five transcripts in the leaves, and up- and down-regulation of 51 and 13 transcripts, respectively, in the flowers of these plants, as compared to the wild type control plants. Most of the changes occurred in the transcripts related to biotic and abiotic stresses, transcription regulation, signaling, metabolic pathways and cell wall modifications, and many of them appeared to be induced through up-regulation of the signaling pathways regulated by ethylene, jasmonic acid and salicylic acid. Correlations of these alterations with the protein profile and related biological functions were analyzed. Surprisingly, they did not cause significant alterations in the protein profile, and caused only very mild alteration in the phenotype of the plants.

Conclusion

Expression of the PVX-specific P25 VSRS protein causes major alterations in the transcriptome of the leaves of transgenic tobacco plants, but very little of any effects in the young flowers of the same plants. The fairly stable protein profile in the leaves and lack of any major changes in the plant phenotype indicate that the complicated interplay and interactions between different regulatory levels are able to maintain homeostasis in the plants.

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Transcription Activator-Like Effector Nucleases Enabl Efficient Plant Genome Engineering

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The ability to precisely engineer plant genomes offers much potential for advancing basic and applied plant biology. Here, we
describe methods for the targeted modification of plant genomes using transcription activator-like effector nucleases (TALENs).
Methods were optimized using tobacco (Nicotiana tabacum) protoplasts and TALENs targeting the acetolactate synthase (ALS)
gene. Optimal TALEN scaffolds were identified using a protoplast-based single-strand annealing assay in which TALEN
cleavage creates a functional yellow fluorescent protein gene, enabling quantification of TALEN activity by flow cytometry.
Single-strand annealing activity data for TALENs with different scaffolds correlated highly with their activity at endogenous
targets, as measured by high-throughput DNA sequencing of polymerase chain reaction products encompassing the TALEN
recognition sites. TALENs introduced targeted mutations in ALS in 30% of transformed cells, and the frequencies of targeted
gene insertion approximated 14%. These efficiencies made it possible to recover genome modifications without selection or
enrichment regimes: 32% of tobacco calli generated from protoplasts transformed with TALEN-encoding constructs had
TALEN-induced mutations in ALS, and of 16 calli characterized in detail, all had mutations in one allele each of the
duplicate ALS genes (SurA and SurB). In calli derived from cells treated with a TALEN and a 322-bp donor molecule
differing by 6 bp from the ALS coding sequence, 4% showed evidence of targeted gene replacement. The optimized reagents
implemented in plant protoplasts should be useful for targeted modification of cells from diverse plant species and using a
variety of means for reagent delivery.

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BMC Plant Biology | Abstract | Selaginella moellendorffii has a reduced and highly conserved expansin superfamily with genes more closely related to angiosperms than to bryophytes

Expansins are plant cell wall loosening proteins encoded by a large superfamily of genes, consisting of four families named EXPA, EXPB, EXLA, and EXLB.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Expansins are plant cell wall loosening proteins encoded by a large superfamily of genes, consisting of four families named EXPA, EXPB, EXLA, and EXLB. The evolution of the expansin superfamily is well understood in angiosperms, thanks to synteny-based evolutionary studies of the gene superfamily in Arabidopsis, rice, and Populus. Analysis of the expansin superfamily in the moss Physcomitrella patens revealed a superfamily without EXLA or EXLB genes that has evolved considerably and independently of angiosperm expansins. The sequencing of the Selaginella moellendorffii genome has allowed us to extend these analyses into an early diverging vascular plant.

Results

The expansin superfamily in Selaginella moellendorffii has now been assembled from genomic scaffolds. A smaller (and less diverse) superfamily is revealed, consistent with studies of other gene families in Selaginella. Selaginella has an expansin superfamily, which, like Physcomitrella, lacks EXLA or EXLB genes, but does contain two EXPA genes that are related to a particular Arabidopsis-rice clade involved in root hair development.

Conclusions

From sequence-based phylogenetic analysis, most Selaginella expansins lie outside the Arabidopsis-rice clades, leading us to estimate the minimum number of expansins present in the last common ancestor of Selaginella and angiosperms at 2 EXPA genes and 1 EXPB gene. These results confirm Selaginella as an important intermediary between bryophytes and angiosperms.

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Population genomic and genome-wide association studies of agroclimatic traits in sorghum

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Abstract

Accelerating crop improvement in sorghum, a staple food for people in semiarid regions across the developing world, is key to ensuring global food security in the context of climate change. To facilitate gene discovery and molecular breeding in sorghum, we have characterized ∼265,000 single nucleotide polymorphisms (SNPs) in 971 worldwide accessions that have adapted to diverse agroclimatic conditions. Using this genome-wide SNP map, we have characterized population structure with respect to geographic origin and morphological type and identified patterns of ancient crop diffusion to diverse agroclimatic regions across Africa and Asia. To better understand the genomic patterns of diversification in sorghum, we quantified variation in nucleotide diversity, linkage disequilibrium, and recombination rates across the genome. Analyzing nucleotide diversity in landraces, we find evidence of selective sweeps around starch metabolism genes, whereas in landrace-derived introgression lines, we find introgressions around known height and maturity loci. To identify additional loci underlying variation in major agroclimatic traits, we performed genome-wide association studies (GWAS) on plant height components and inflorescence architecture. GWAS maps several classical loci for plant height, candidate genes for inflorescence architecture. Finally, we trace the independent spread of multiple haplotypes carrying alleles for short stature or long inflorescence branches. This genome-wide map of SNP variation in sorghum provides a basis for crop improvement through marker-assisted breeding and genomic selection.

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Genome Biology | Abstract | Q & A: the Snyderome

Michael Snyder answers Genome Biology's questions on the human and professional stories underlying his Snyderome integrative omics project.
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Michael Snyder answers Genome Biology's questions on the human and professional stories underlying his Snyderome integrative omics project.

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BMC Genomics | Full text | A comparative transcriptomic, fluxomic and metabolomic analysis of the response of Saccharomyces cerevisiae to increases in NADPH oxidation

Redox homeostasis is essential to sustain metabolism and growth.
Biswapriya Biswavas Misra's insight:
AbstractBackground

Redox homeostasis is essential to sustain metabolism and growth. We recently reported that yeast cells meet a gradual increase in imposed NADPH demand by progressively increasing flux through the pentose phosphate (PP) and acetate pathways and by exchanging NADH for NADPH in the cytosol, via a transhydrogenase-like cycle. Here, we studied the mechanisms underlying this metabolic response, through a combination of gene expression profiling and analyses of extracellular and intracellular metabolites and 13 C-flux analysis.

Results

NADPH oxidation was increased by reducing acetoin to 2,3-butanediol in a strain overexpressing an engineered NADPH-dependent butanediol dehydrogenase cultured in the presence of acetoin. An increase in NADPH demand to 22 times the anabolic requirement for NADPH was accompanied by the intracellular accumulation of PP pathway metabolites consistent with an increase in flux through this pathway. Increases in NADPH demand were accompanied by the successive induction of several genes of the PP pathway. NADPH-consuming pathways, such as amino-acid biosynthesis, were upregulated as an indirect effect of the decrease in NADPH availability. Metabolomic analysis showed that the most extreme modification of NADPH demand resulted in an energetic problem. Our results also highlight the influence of redox status on aroma production.

Conclusions

Combined 13 C-flux, intracellular metabolite levels and microarrays analyses revealed that NADPH homeostasis, in response to a progressive increase in NADPH demand, was achieved by the regulation, at several levels, of the PP pathway. This pathway is principally under metabolic control, but regulation of the transcription of PP pathway genes can exert a stronger effect, by redirecting larger amounts of carbon to this pathway to satisfy the demand for NADPH. No coordinated response of genes involved in NADPH metabolism was observed, suggesting that yeast has no system for sensing NADPH/NADP+ ratio. Instead, the induction of NADPH-consuming amino-acid pathways in conditions of NADPH limitation may indirectly trigger the transcription of a set of PP pathway genes.

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The genome of Prunus mume : Nature Communications : Nature Publishing Group

The genome of Prunus mume : Nature Communications : Nature Publishing Group | Plant Genomics | Scoop.it
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Prunus mume (mei), which was domesticated in China more than 3,000 years ago as ornamental plant and fruit, is one of the first genomes among Prunus subfamilies of Rosaceae been sequenced. Here, we assemble a 280M genome by combining 101-fold next-generation sequencing and optical mapping data. We further anchor 83.9% of scaffolds to eight chromosomes with genetic map constructed by restriction-site-associated DNA sequencing. Combining P. mume genome with available data, we succeed in reconstructing nine ancestral chromosomes of Rosaceae family, as well as depicting chromosome fusion, fission and duplication history in three major subfamilies. We sequence the transcriptome of various tissues and perform genome-wide analysis to reveal the characteristics of P. mume, including its regulation of early blooming in endodormancy, immune response against bacterial infection and biosynthesis of flower scent. The P. mume genome sequence adds to our understanding of Rosaceae evolution and provides important data for improvement of fruit trees.

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BMC Plant Biology | Abstract | Analysis of ripening-related gene expression in papaya using an Arabidopsis-based microarray

BMC Plant Biology | Abstract | Analysis of ripening-related gene expression in papaya using an Arabidopsis-based microarray | Plant Genomics | Scoop.it
Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals. Despite its importance, little is known about transcriptional modifications during papaya fruit ripening and their control. In this study we report the analysis of ripe papaya transcriptome by using a cross-species (XSpecies) microarray technique based on the phylogenetic proximity between papaya and Arabidopsis thaliana.

Results

Papaya transcriptome analyses resulted in the identification of 414 ripening-related genes with some having their expression validated by qPCR. The transcription profile was compared with that from ripening tomato and grape. There were many similarities between papaya and tomato especially with respect to the expression of genes encoding proteins involved in primary metabolism, regulation of transcription, biotic and abiotic stress and cell wall metabolism. XSpecies microarray data indicated that transcription factors (TFs) of the MADS-box, NAC and AP2/ERF gene families were involved in the control of papaya ripening and revealed that cell wall-related gene expression in papaya had similarities to the expression profiles seen in Arabidopsis during hypocotyl development.

Conclusion

The cross-species array experiment identified a ripening-related set of genes in papaya allowing the comparison of transcription control between papaya and other fruit bearing taxa during the ripening process.

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BMC Genomics | Abstract | Development of high-throughput SNP-based genotyping in Acacia auriculiformis x A. mangium hybrids using short-read transcriptome data

Next Generation Sequencing has provided comprehensive, affordable and high-throughput DNA sequences for Single Nucleotide Polymorphism (SNP) discovery in Acacia auriculiformis and Acacia mangium.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Next Generation Sequencing has provided comprehensive, affordable and high-throughput DNA sequences for Single Nucleotide Polymorphism (SNP) discovery in Acacia auriculiformis and Acacia mangium. Like other non-model species, SNP detection and genotyping in Acacia are challenging due to lack of genome sequences. The main objective of this study is to develop the first high-throughput SNP genotyping assay for linkage map construction of A. auriculiformis x A. mangium hybrids.

Results

We identified a total of 37,786 putative SNPs by aligning short read transcriptome data from four parents of two Acacia hybrid mapping populations using Bowtie against 7,839 de novo transcriptome contigs. Given a set of 10 validated SNPs from two lignin genes, our in silico SNP detection approach is highly accurate (100%) compared to the traditional in vitro approach (44%). Further validation of 96 SNPs using Illumina GoldenGate Assay gave an overall assay success rate of 89.6% and conversion rate of 37.5%. We explored possible factors lowering assay success rate by predicting exon-intron boundaries and paralogous genes of Acacia contigs using Medicago truncatula genome as reference. This assessment revealed that presence of exon-intron boundary is the main cause (50%) of assay failure. Subsequent SNPs filtering and improved assay design resulted in assay success and conversion rate of 92.4% and 57.4%, respectively based on 768 SNPs genotyping. Analysis of clustering patterns revealed that 27.6% of the assays were not reproducible and flanking sequence might play a role in determining cluster compression. In addition, we identified a total of 258 and 319 polymorphic SNPs in A. auriculiformis and A. mangium natural germplasms, respectively.

Conclusion

We have successfully discovered a large number of SNP markers in A. auriculiformis x A. mangium hybrids using next generation transcriptome sequencing. By using a reference genome from the most closely related species, we converted most SNPs to successful assays. We also demonstrated that Illumina GoldenGate genotyping together with manual clustering can provide high quality genotypes for a non-model species like Acacia. These SNPs markers are not only important for linkage map construction, but will be very useful for hybrid discrimination and genetic diversity assessment of natural germplasms in the future.

 
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Rescooped by Biswapriya Biswavas Misra from Plant Biology Teaching Resources (Higher Education)
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iRootHair - Genomics database of ... root hair information!

iRootHair - Genomics database of ... root hair information! | Plant Genomics | Scoop.it

Root Hair Genomics Database.. genes, processes, mutants, references etc, with the option of searching just monocots. Nice! 

 

The accompanying paper is just out in Plant Physiology:

http://www.plantphysiol.org/content/early/2012/11/05/pp.112.206441.abstract


Via Mary Williams
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PLOS ONE: Comparative Transcriptome Profiling of the Early Response to Magnaporthe oryzae in Durable Resistant vs Susceptible Rice (Oryza sativa L.) Genotypes

PLOS ONE: Comparative Transcriptome Profiling of the Early Response to Magnaporthe oryzae in Durable Resistant vs Susceptible Rice (Oryza sativa L.) Genotypes | Plant Genomics | Scoop.it
PLOS ONE: an inclusive, peer-reviewed, open-access resource from the PUBLIC LIBRARY OF SCIENCE. Reports of well-performed scientific studies from all disciplines freely available to the whole world.
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Abstract

 

Durable resistance to blast, the most significant fungal disease of rice, represents an agronomically relevant character. Gigante Vercelli (GV) and Vialone Nano (VN) are two old temperate japonica Italian rice cultivars with contrasting response to blast infection: GV displays durable and broad resistance while VN is highly susceptible. RNA-seq was used to dissect the early molecular processes deployed during the resistance response of GV at 24 h after blast inoculation. Differential gene expression analysis identified 1,070 and 1,484 modulated genes, of which 726 and 699 were up regulated in response to infection in GV and VN, respectively. Gene ontology (GO) enrichment analyses revealed a set of GO terms enriched in both varieties but, despite this commonality, the gene sets contributing to common GO enriched terms were dissimilar. The expression patterns of genes grouped in GV-specific enriched GO terms were examined in detail including at the transcript isoform level. GV exhibited a dramatic up-regulation of genes encoding diterpene phytoalexin biosynthetic enzymes, flavin-containing monooxygenase, class I chitinase and glycosyl hydrolase 17. The sensitivity and high dynamic range of RNA-seq allowed the identification of genes critically involved in conferring GV resistance during the early steps of defence perception-signalling. These included chitin oligosaccharides sensing factors, wall associated kinases, MAPK cascades and WRKY transcription factors. Candidate genes with expression patterns consistent with a potential role as GV-specific functional resistance (R) gene(s) were also identified. This first application of RNA-seq to dissect durable blast resistance supports a crucial role of the prompt induction of a battery of responses including defence-related genes as well as members of gene families involved in signalling and pathogen-related gene expression regulation.

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BMC Biology | Full text | A spruce gene map infers ancient plant genome reshuffling and subsequent slow evolution in the gymnosperm lineage leading to extant conifers

Seed plants are composed of angiosperms and gymnosperms, which diverged from each other around 300 million years ago.
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AbstractBackground

Seed plants are composed of angiosperms and gymnosperms, which diverged from each other around 300 million years ago. While much light has been shed on the mechanisms and rate of genome evolution in flowering plants, such knowledge remains conspicuously meagre for the gymnosperms. Conifers are key representatives of gymnosperms and the sheer size of their genomes represents a significant challenge for characterization, sequencing and assembling.

Results

To gain insight into the macro-organisation and long-term evolution of the conifer genome, we developed a genetic map involving 1,801 spruce genes. We designed a statistical approach based on kernel density estimation to analyse gene density and identified seven gene-rich isochors. Groups of co-localizing genes were also found that were transcriptionally co-regulated, indicative of functional clusters. Phylogenetic analyses of 157 gene families for which at least two duplicates were mapped on the spruce genome indicated that ancient gene duplicates shared by angiosperms and gymnosperms outnumbered conifer-specific duplicates by a ratio of eight to one. Ancient duplicates were much more translocated within and among spruce chromosomes than conifer-specific duplicates, which were mostly organised in tandem arrays. Both high synteny and collinearity were also observed between the genomes of spruce and pine, two conifers that diverged more than 100 million years ago.

Conclusions

Taken together, these results indicate that much genomic evolution has occurred in the seed plant lineage before the split between gymnosperms and angiosperms, and that the pace of evolution of the genome macro-structure has been much slower in the gymnosperm lineage leading to extent conifers than that seen for the same period of time in flowering plants. This trend is largely congruent with the contrasted rates of diversification and morphological evolution observed between these two groups of seed plants.

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BMC Plant Biology | Abstract | Analysis of ripening-related gene expression in papaya using an Arabidopsis-based microarray

Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals. Despite its importance, little is known about transcriptional modifications during papaya fruit ripening and their control. In this study we report the analysis of ripe papaya transcriptome by using a cross-species (XSpecies) microarray technique based on the phylogenetic proximity between papaya and Arabidopsis thaliana.

Results

Papaya transcriptome analyses resulted in the identification of 414 ripening-related genes with some having their expression validated by qPCR. The transcription profile was compared with that from ripening tomato and grape. There were many similarities between papaya and tomato especially with respect to the expression of genes encoding proteins involved in primary metabolism, regulation of transcription, biotic and abiotic stress and cell wall metabolism. XSpecies microarray data indicated that transcription factors (TFs) of the MADS-box, NAC and AP2/ERF gene families were involved in the control of papaya ripening and revealed that cell wall-related gene expression in papaya had similarities to the expression profiles seen in Arabidopsis during hypocotyl development.

Conclusion

The cross-species array experiment identified a ripening-related set of genes in papaya allowing the comparison of transcription control between papaya and other fruit bearing taxa during the ripening process.

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Genetic Adaptation Associated with Genome-Doubling in Autotetraploid Arabidopsis arenosa

Genetic Adaptation Associated with Genome-Doubling in Autotetraploid Arabidopsis arenosa | Plant Genomics | Scoop.it
PLOS Genetics is an open-access
Biswapriya Biswavas Misra's insight:

Genome duplication, which results in polyploidy, is disruptive to fundamental biological processes. Genome duplications occur spontaneously in a range of taxa and problems such as sterility, aneuploidy, and gene expression aberrations are common in newly formed polyploids. In mammals, genome duplication is associated with cancer and spontaneous abortion of embryos. Nevertheless, stable polyploid species occur in both plants and animals. Understanding how natural selection enabled these species to overcome early challenges can provide important insights into the mechanisms by which core cellular functions can adapt to perturbations of the genomic environment. Arabidopsis arenosa includes stable tetraploid populations and is related to well-characterized diploids A. lyrata and A. thaliana. It thus provides a rare opportunity to leverage genomic tools to investigate the genetic basis of polyploid stabilization. We sequenced the genomes of twelve A. arenosa individuals and found signatures suggestive of recent and ongoing selective sweeps throughout the genome. Many of these are at genes implicated in genome maintenance functions, including chromosome cohesion and segregation, DNA repair, homologous recombination, transcriptional regulation, and chromatin structure. Numerous encoded proteins are predicted to interact with one another. For a critical meiosis gene, ASYNAPSIS1, we identified a non-synonymous mutation that is highly differentiated by cytotype, but present as a rare variant in diploid A. arenosa, indicating selection may have acted on standing variation already present in the diploid. Several genes we identified that are implicated in sister chromatid cohesion and segregation are homologous to genes identified in a yeast mutant screen as necessary for survival of polyploid cells, and also implicated in genome instability in human diseases including cancer. This points to commonalities across kingdoms and supports the hypothesis that selection has acted on genes controlling genome integrity in A. arenosa as an adaptive response to genome doubling.

 
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Regulatory Impact of RNA Secondary Structure across the Arabidopsis Transcriptome

Regulatory Impact of RNA Secondary Structure across the Arabidopsis Transcriptome | Plant Genomics | Scoop.it
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Abstract

The secondary structure of an RNA molecule plays an integral role in its maturation, regulation, and function. However, the global influence of this feature on plant gene expression is still largely unclear. Here, we use a high-throughput, sequencing-based, structure-mapping approach in conjunction with transcriptome-wide sequencing of rRNA-depleted (RNA sequencing), small RNA, and ribosome-bound RNA populations to investigate the impact of RNA secondary structure on gene expression regulation in Arabidopsis thaliana. From this analysis, we find that highly unpaired and paired RNAs are strongly correlated with euchromatic and heterochromatic epigenetic histone modifications, respectively, providing evidence that secondary structure is necessary for these RNA-mediated posttranscriptional regulatory pathways. Additionally, we uncover key structural patterns across protein-coding transcripts that indicate RNA folding demarcates regions of protein translation and likely affects microRNA-mediated regulation of mRNAs in this model plant. We further reveal that RNA folding is significantly anticorrelated with overall transcript abundance, which is often due to the increased propensity of highly structured mRNAs to be degraded and/or processed into small RNAs. Finally, we find that secondary structure affects mRNA translation, suggesting that this feature regulates plant gene expression at multiple levels. These findings provide a global assessment of RNA folding and its significant regulatory effects in a plant transcriptome.

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BMC Plant Biology | Abstract | Partial functional conservation of IRX10 homologs in physcomitrella patens and Arabidopsis thaliana indicates an evolutionary step contributing to vascular formation ...

Plant cell walls are complex multicomponent structures that have evolved to fulfil an essential function in providing strength and protection to cells.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Plant cell walls are complex multicomponent structures that have evolved to fulfil an essential function in providing strength and protection to cells. Hemicelluloses constitute a key component of the cell wall and recently a number of the genes thought to encode the enzymes required for its synthesis have been identified in Arabidopsis. The acquisition of hemicellulose synthesis capability is hypothesised to have been an important step in the evolution of higher plants.

Results

Analysis of the Physcomitrella patens genome has revealed the presence of homologs for all of the Arabidopsis glycosyltransferases including IRX9, IRX10 and IRX14 required for the synthesis of the glucuronoxylan backbone. The Physcomitrella IRX10 homolog is expressed in a variety of moss tissues which were newly formed or undergoing expansion. There is a high degree of sequence conservation between the Physcomitrella IRX10 and Arabidopsis IRX10 and IRX10-L. Despite this sequence similarity, the Physcomitrella IRX10 gene is only able to partially rescue the Arabidopsis irx10 irx10-L double mutant indicating that there has been a neo- or sub-functionalisation during the evolution of higher plants. Analysis of the monosaccharide composition of stems from the partially rescued Arabidopsis plants does not show any significant change in xylose content compared to the irx10 irx10-L double mutant. Likewise, knockout mutants of the Physcomitrella IRX10 gene do not result in any visible phenotype and there is no significant change in monosaccharide composition of the cell walls.

Conclusions

The fact that the Physcomitrella IRX10 (PpGT47A) protein can partially complement an Arabidopsis irx10 irx10-L double mutant suggests that it shares some function with the Arabidopsis proteins, but the lack of a phenotype in knockout lines shows that the function is not required for growth or development under normal conditions in Physcomitrella. In contrast, the Arabidopsis irx10 and irx10 irx10-L mutants have strong phenotypes indicating an important function in growth and development. We conclude that the evolution of vascular plants has been associated with a significant change or adaptation in the function of the IRX10 gene family.

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BMC Genomics | Abstract | Comprehensive structural annotation of Pichia pastoris transcriptome and the response to various carbon sources using deep paired-end RNA sequencing

The methylotrophic yeast Pichia pastoris is widely used as a bioengineering platform for producing industrial and biopharmaceutical proteins, studying protein expression and secretion mechanisms, and analyzing metabolite synthesis and peroxisome...
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Abstract (provisional)Background

The methylotrophic yeast Pichia pastoris is widely used as a bioengineering platform for producing industrial and biopharmaceutical proteins, studying protein expression and secretion mechanisms, and analyzing metabolite synthesis and peroxisome biogenesis. With the development of DNA microarray and mRNA sequence technology, the P. pastoris transcriptome has become a research hotspot due to its powerful capability to identify the transcript structures and gain insights into the transcriptional regulation model of cells under protein production conditions. The study of the P. pastoris transcriptome helps to annotate the P. pastoris transcript structures and provide useful information for further improvement of the production of recombinant proteins.

Results

We used a massively parallel mRNA sequencing platform (RNA-Seq), based on next-generation sequencing technology, to map and quantify the dynamic transcriptome of P. pastoris at the genome scale under growth conditions with glycerol and methanol as substrates. The results describe the transcription landscape at the whole-genome level and provide annotated transcript structures, including untranslated regions (UTRs), alternative splicing (AS) events, novel transcripts, new exons, alternative upstream initiation codons (uATGs), and upstream open reading frames (uORFs). Internal ribosome entry sites (IRESs) were first identified within the UTRs of genes from P. pastoris, encoding kinases and the proteins involved in the control of growth. We also provide a transcriptional regulation model for P. pastoris grown on different carbon sources.

Conclusions

We suggest that the IRES-dependent translation initiation mechanism also exists in P. pastoris. Retained introns (RI) are determined as the main AS event and are produced predominantly by an intron definition (ID) mechanism. Our results describe the metabolic characteristics of P. pastoris with heterologous protein production under methanol induction and provide rich information for further in-depth studies of P. pastoris protein expression and secretion mechanisms.

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Transcriptome-wide mapping of N6-methyladenosine by m6A-seq based on immunocapturing and massively parallel sequencing : Nature Protocols : Nature Publishing Group

Transcriptome-wide mapping of N6-methyladenosine by m6A-seq based on immunocapturing and massively parallel sequencing : Nature Protocols : Nature Publishing Group | Plant Genomics | Scoop.it
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Abstract

N6-methyladenosine–sequencing (m6A-seq) is an immunocapturing approach for the unbiased transcriptome-wide localization of m6A in high resolution. To our knowledge, this is the first protocol to allow a global view of this ubiquitous RNA modification, and it is based on antibody-mediated enrichment of methylated RNA fragments followed by massively parallel sequencing. Building on principles of chromatin immunoprecipitation–sequencing (ChIP-seq) and methylated DNA immunoprecipitation (MeDIP), read densities of immunoprecipitated RNA relative to untreated input control are used to identify methylated sites. A consensus motif is deduced, and its distance to the point of maximal enrichment is assessed; these measures further corroborate the success of the protocol. Identified locations are intersected in turn with gene architecture to draw conclusions regarding the distribution of m6A between and within gene transcripts. When applied to human and mouse transcriptomes, m6A-seq generated comprehensive methylation profiles revealing, for the first time, tenets governing the nonrandom distribution of m6A. The protocol can be completed within ∼9 d for four different sample pairs (each consists of an immunoprecipitation and corresponding input).

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Identification of candidate effector genes in the transcriptome of the rice root knot nematode Meloidogyne graminicola - Haegeman - 2012 - Molecular Plant Pathology - Wiley Online Library

Identification of candidate effector genes in the transcriptome of the rice root knot nematode Meloidogyne graminicola - Haegeman - 2012 - Molecular Plant Pathology - Wiley Online Library | Plant Genomics | Scoop.it
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Summary

Plant-parasitic nematodes secrete so-called effectors into their host plant which are able to suppress the plant's defence responses, alter plant signalling pathways and, in the case of root knot nematodes, induce the formation of giant cells. Putative effectors have been successfully identified by genomics, transcriptomics and proteomics approaches. In this study, we investigated the transcriptome of the rice root knot nematode Meloidogyne graminicola by 454 sequencing of second-stage juveniles as well as mRNA-seq of rice infected tissue. Over 350 000 reads derived from M. graminicola preparasitic juveniles were assembled, annotated and checked for homologues in different databases. From infected rice tissue, 1.4% of all reads generated were identified as being derived from the nematode. Using multiple strategies, several putative effector genes were identified, both pioneer genes and genes corresponding to already known effectors. To check whether these genes could be involved in the interaction with the plant, in situ hybridization was performed on a selection of genes to localize their expression in the nematode. Most were expressed in the gland cells or amphids of the nematode, confirming possible secretion of the proteins and hence a role in infection. Other putative effectors showed a different expression pattern, potentially linked with the excretory/secretory system. This transcriptome study is a good starting point to functionally investigate novel effectors derived from M. graminicola. This will lead to better insights into the interaction between these nematodes and the model plant rice. Moreover, the transcriptome can be used to identify possible target genes for RNA interference (RNAi)-based control strategies. Four genes proved to be interesting targets by showing up to 40% higher mortality relative to the control treatment when soaked in gene-specific small interfering RNAs (siRNAs).

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Parental contributions to the transcriptome of early plant embryos | Current Comments

Parental contributions to the transcriptome of early plant embryos | Current Comments | Plant Genomics | Scoop.it
Biswapriya Biswavas Misra's insight:

In plants and animals, embryo development becomes ultimately controlled by zygotic genes, but the timing of zygotic genome activation (ZGA) varies greatly between organisms[1,2]. We recently showed that the transcriptome of young Arabidopsis embryos is dominated by maternal transcripts with a progressive ZGA under the maternal control of epigenetic pathways [3]. In contrast, another study reported that both parental genomes contribute equally to the transcriptome of young embryos, suggesting that ZGA occurs immediately after fertilization[4]. How to explain such dramatic differences?We propose that the discrepancies between these two studies likely reflect genuine biological differences between the two experiments, paving the road towards exciting discoveries on ZGA mechanisms in plants.

In animals, early stages of embryo development are associated with extensive epigenetic reprogramming to coordinate zygotic genome activation (ZGA) [2]. ZGA is typically delayed, although to a varying extent depending on the species, with a gradual loss of the maternal dominance and increase of zygotic influence [1,2]. In flowering plants, maternal effects on seed development have been recognized, yet are difficult to investigate because of the intricate relationships between the embryo, the embryo-nourishing endosperm, and the maternal seed coat [5]. To understand the interaction of parental genomes following fertilization, allele-specific assays were used to distinguish paternal and maternal contributions for selected loci or at the genome-wide level in dissected embryos (reviewed in [1]), with surprisingly different results. Yet, the diversity of species (Arabidopsis, maize, tobacco) and developmental stages analyzed made it difficult to draw general conclusions. In fact, the observed differences may reflect yet undiscovered biological factors controlling ZGA in flowering plants.

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BMC Genomics | Abstract | Genome-guided analysis of physiological and morphological traits of the fermentative acetate oxidizer Thermacetogenium phaeum

Thermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to CO2 in syntrophic association with a methanogenic partner. It can also grow in pure culture, e.g., by fermentation of methanol to acetate.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Thermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to CO2 in syntrophic association with a methanogenic partner. It can also grow in pure culture, e.g., by fermentation of methanol to acetate. The key enzymes of homoacetate fermentation (Wood-Ljungdahl pathway) are used both in acetate oxidation and acetate formation. The obvious reversibility of this pathway in this organism is of specific interest since syntrophic acetate oxidation operates close to the energetic limitations of microbial life.

Results

The genome of Th. phaeum is organized on a single circular chromosome and has a total size of 2,939,057 bp. It comprises 3.215 open reading frames of which 75% could be assigned to a gene function. The G+C content is 53.88 mol%. Many CRISPR sequences were found, indicating heavy phage attack in the past. A complete gene set for a phage was found in the genome, and indications of phage action could also be observed in culture. The genome contained all genes required for CO2 reduction through the Wood-Ljungdahl pathway, including two formyl tetrahydrofolate ligases, three carbon monoxide dehydrogenases, one formate hydrogenlyase complex, three further formate dehydrogenases, and three further hydrogenases. The bacterium contains a menaquinone MQ-7. No indications of cytochromes or Rnf complexes could be found in the genome.

Conclusions

The information obtained from the genome sequence indicates that Th. phaeum differs basically from the three homoacetogenic bacteria sequenced so far, i.e., the sodium ion-dependent Acetobacterium woodii, the ethanol-producing Clostridium ljungdahlii, and the cytochrome-containing Moorella thermoacetica. The specific enzyme outfit of Th. phaeum obviously allows ATP formation both in acetate formation and acetate oxidation.

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BMC Genomics | Full text | 'Bois noir' phytoplasma induces significant reprogramming of the leaf transcriptome in the field grown grapevine

Phytoplasmas are bacteria without cell walls from the class Mollicutes. They are obligate intracellular plant pathogens which cause diseases in hundreds of economically important plants including the grapevine (Vitis vinifera).

Via Knapco
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Knapco's curator insight, December 26, 2012 7:07 AM

This study revealed some fundamental aspects of grapevine interactions with the stolbur 'Bois noir' phytoplasma in particular and some plant interactions with phytoplasmas in general. In addition, the results of the study will likely have an impact on grape improvement by yielding marker genes that can be used in new diagnostic assays for phytoplasmas or by identifying candidate genes that contribute to the improved properties of grape.

By: Hren M, Nikolić P, Rotter A, Blejec A, Terrier, Ravnikar M, Dermastia M, Gruden K; BMC Genomics 2009, 10:460 doi:10.1186/1471-2164-10-460

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Genomic Selection in Wheat Breeding using Genotyping-by-Sequencing

Genomic Selection in Wheat Breeding using Genotyping-by-Sequencing | Plant Genomics | Scoop.it
Biswapriya Biswavas Misra's insight:

Genomic selection (GS) uses genomewide molecular markers to predict breeding values and make selections of individuals or breeding lines prior to phenotyping. Here we show that genotyping-by-sequencing (GBS) can be used for de novo genotyping of breeding panels and to develop accurate GS models, even for the large, complex, and polyploid wheat (Triticum aestivum L.) genome. With GBS we discovered 41,371 single nucleotide polymorphisms (SNPs) in a set of 254 advanced breeding lines from CIMMYT’s semiarid wheat breeding program. Four different methods were evaluated for imputing missing marker scores in this set of unmapped markers, including random forest regression and a newly developed multivariate-normal expectation-maximization algorithm, which gave more accurate imputation than heterozygous or mean imputation at the marker level, although no significant differences were observed in the accuracy of genomic-estimated breeding values (GEBVs) among imputation methods. Genomic-estimated breeding value prediction accuracies with GBS were 0.28 to 0.45 for grain yield, an improvement of 0.1 to 0.2 over an established marker platform for wheat. Genotyping-by-sequencing combines marker discovery and genotyping of large populations, making it an excellent marker platform for breeding applications even in the absence of a reference genome sequence or previous polymorphism discovery. In addition, the flexibility and low cost of GBS make this an ideal approach for genomics-assisted breeding.

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BMC Genomics | Abstract | Transcriptome analysis of Sacha Inchi (Plukenetia volubilis L.) seeds at two developmental stages

Sacha Inchi (Plukenetia volubilis L., Euphorbiaceae) is a potential oilseed crop because the seeds of this plant are rich in unsaturated fatty acids (FAs).
Biswapriya Biswavas Misra's insight:

Abstract (provisional)Background

Sacha Inchi (Plukenetia volubilis L., Euphorbiaceae) is a potential oilseed crop because the seeds of this plant are rich in unsaturated fatty acids (FAs). In particular, the fatty acid composition of its seed oil differs markedly in containing large quantities of alpha-linolenic acid (18C:3, a kind of omega-3 FAs). However, little is known about the molecular mechanisms responsible for biosynthesis of unsaturated fatty acids in the developing seeds of this species. Transcriptome data are needed to better understand these mechanisms.

Results

In this study, de novo transcriptome assembly and gene expression analysis were performed using Illumina sequencing technology. A total of 52.6 million 90-bp paired-end reads were generated from two libraries constructed at the initial stage and fast oil accumulation stage of seed development. These reads were assembled into 70,392 unigenes; 22,179 unigenes showed a 2-fold or greater expression difference between the two libraries. Using this data we identified unigenes that may be involved in de novo FA and triacylglycerol biosynthesis. In particular, a number of unigenes encoding desaturase for formation of unsaturated fatty acids with high expression levels in the fast oil accumulation stage compared with the initial stage of seed development were identified.

Conclusions

This study provides the first comprehensive dataset characterizing Sacha Inchi gene expression at the transcriptional level. These data provide the foundation for further studies on molecular mechanisms underlying oil accumulation and PUFA biosynthesis in Sacha Inchi seeds. Our analyses facilitate understanding of the molecular mechanisms responsible for the high unsaturated fatty acids (especially alpha-linolenic acid) accumulation in Sacha Inchi seeds.

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BMC Genomics | Abstract | Transcriptomic analysis of 'Suli' pear (Pyrus pyrifolia white pear group) buds during the dormancy by RNA-Seq

Bud dormancy is a critical developmental process that allows perennial plants to survive unfavorable environmental conditions.
Biswapriya Biswavas Misra's insight:
Abstract (provisional)Background

Bud dormancy is a critical developmental process that allows perennial plants to survive unfavorable environmental conditions. Pear is one of the most important deciduous fruit trees in the world, but the mechanisms regulating bud dormancy in this species are unknown. Because genomic information for pear is currently unavailable, transcriptome and digital gene expression data for this species would be valuable resources to better understand the molecular and biological mechanisms regulating its bud dormancy.

Results

We performed de novo transcriptome assembly and digital gene expression (DGE) profiling analyses of for 'Suli' pear (Pyrus pyrifolia white pear group) using the Illumina RNA-seq system. RNA-Seq generated approximately 100 M high-quality reads that were assembled into 69,393 unigenes (mean length = 853 bp), including 14,531 clusters and 34,194 singletons. A total of 51,448 (74.1%) unigenes were annotated using public protein databases with a cut-off E-value above 10-5. We mainly compared gene expression levels at four time-points during bud dormancy. Between Nov. 15 and Dec. 15, Dec. 15 and Jan. 15, and Jan. 15 and Feb. 15, 1,978, 1,024, and 3,468 genes were differentially expressed, respectively. Hierarchical clustering analysis arranged 190 significantly differentially-expressed genes into seven groups. Seven genes were randomly selected to confirm their expression levels using quantitative real-time PCR.

Conclusions

The new transcriptomes offer comprehensive sequence and DGE profiling data for a dynamic view of transcriptomic variation during bud dormancy in pear. These data provided a basis for future studies of metabolism during bud dormancy in non-model but economically-important perennial species.

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