Global changes of gene expression during scleroglucan synthesis in Sclerotium rolfsii
Ontology highlight
ABSTRACT:
ORGANISM(S): Agroathelia rolfsii
PROVIDER: GSE25254 | GEO | 2011/03/01
SECONDARY ACCESSION(S): PRJNA134679
REPOSITORIES: GEO
Similar Datasets
Project description:Two S. rolfsii transcriptomes obtained from scleroglucan-producing and scleroglucan-nonproducing conditions were pooled and sequenced using the 454 pyrosequencing technique yielding ~350,000 reads. These could be assembled into 21,937 contigs and 171,833 singletons, for which 6,951 had significant matches in public protein data bases. Sequence data were used to obtain first insights into the genomics of scleroglucan and oxalate production and to predict putative proteins involved in the synthesis of both metabolites. Using comparative transcriptomics, namely Agilent microarray hybridization and suppression subtractive hybridization, we identified ~ 800 unigenes which are differently expressed under scleroglucan-producing and non-producing conditions.
2010-05-15 | GSE21040 | GEO
Project description:Two S. rolfsii transcriptomes obtained from scleroglucan-producing and scleroglucan-nonproducing conditions were pooled and sequenced using the 454 pyrosequencing technique yielding ~350,000 reads. These could be assembled into 21,937 contigs and 171,833 singletons, for which 6,951 had significant matches in public protein data bases. Sequence data were used to obtain first insights into the genomics of scleroglucan and oxalate production and to predict putative proteins involved in the synthesis of both metabolites. Using comparative transcriptomics, namely Agilent microarray hybridization and suppression subtractive hybridization, we identified ~ 800 unigenes which are differently expressed under scleroglucan-producing and non-producing conditions. Tailor-made microarrays (44K multiplex chip, Agilent) were designed by imaGenes (Berlin, Germany) using an in-house developed method for empirical selection of best performing probes for each gene (Pre Selection Strategy). Briefly, up to ten probes were designed for each of the 454 unigenes (60 bp long oligomers). The 244K Agilent test array was hybridized with pooled Cy3-labeled cRNAs gained form EPSmax13 and EPSmin17 cultures and (in average) two of the best performing oligos were selected for each unigene. For comparative expression profiling, total RNA was isolated from S. rolfsii, cultured for 37 h in EPSmax13 (biological triplicate) and EPSmin17 media (biological triplicate). RNA quality control, synthesis of Cy3-labeled cRNA including cRNA purification and cRNA quality control, microarray hybridization, scanning and data extraction (Agilent´s feature extraction software) were performed by imaGenes GmbH. Expression data were analyzed by imaGenes GmbH using an in-house developed data analysis pipeline. After quantile normalization, genes were defined as differentially expressed if their expression levels varied at least 2 fold in EPSmax13 samples compared to EPSmin17 samples and if the difference was statistically significant (Student’s t-test, P-value cut-off of 0.05). 454 pyrosequencing cDNA study data was uploaded at SRA and assigned accession number SRP002142.
2010-05-15 | E-GEOD-21040 | biostudies-arrayexpress
Project description:The soil worm Enchytraeus crypticus (oligochaete) is an ecotoxicology model species although without genome or transcriptome sequence information. The present research aimed at studying, via high-throughput pyrosequencing, the transcriptome of Enchytraeus crypticus, sampled from multiple test conditions, and the construction of a high-density microarray for functional genomic studies. A pyrosequencing run retrieved approximately 1.5 million reads representing 645 million bases. After assembly, 27,296 contigs and 87,686 singletons were obtained. from which 44% and 25% were annotated as protein-coding genes. We show that the high amount of orphan genes is not due to poor sequence or assemble quality: 84% of the contig sequences contains an open reading frame with a start codon and E. crypticus homologs were identified for 92% of the core eukaryotic genes. Moreover, 65 and 77% of the unknown singletons and contigs, respectively, showed transcriptional activity. An Agilent 180K microarray platform was designed and validated by hybridizing cDNA from 3 day zinc- exposed E. crypticus to the concentration corresponding to 50% reduction in reproduction (EC50). Overall, 70% of all probes exerted a hybridization signal above background level. More specifically, the probes derived from contigs showed a wider range of average intensities when compared to probes derived from singletons. In total, 522 significantly regulated transcripts were identifying upon zinc exposure. Several significantly regulated genes exerted predicted functions (e.g. zinc efflux, zinc transport) associated with zinc stress. Unexpectedly, the microarray data suggest that zinc exposure alters retrotransposon activity in the E. crypticus genome. In conclusion, characterization of the presented E. crypticus transcriptome and associated microarray platform is a valuable and high quality resource that permits further functional genomics experiments examining gene expression patterns underlying distinct environmental stress conditions. We show that unknown sequences are not the result of technical errors but mostly represent functional genes that are actively transcribed.
2014-05-01 | GSE51838 | GEO
Project description:A method was developed to isolate RNA from 1 dpa fiber. ESTs derived from this and other cotton mRNAs were sequenced and assembled into contigs. Contigs composed of EST unique to libraries of interest and unique singletons were represented on a microarray. Microarrays were hybridized with labed nucleic acids derived from 10 dpa fiber and 1 dpa fiber to identify genes differentially regulated during fiber initiation and fiber elongation. Genes with known expression in fiber were included to validate the microarray. Analysis of the gene ontology (GO) of differentially expressed genes suported previously reported aspects of fiber initiation such as cell wall remodeling. Transcription factors upregulated in fiber initials and elongating fiber were identified Keywords: cell type comparison
2007-06-26 | GSE6852 | GEO
Project description:Genome-wide DNA sequence resources are expected to enhance our understanding of the molecular basis of plant development and lead to improvements in desirable traits in horticultural crops. In this study, we sequenced 643,366 ESTs from Eustoma grandiflorum flowers using a normalised cDNA library constructed for several different flower development stages, times of day, and pollinated stamens. The sequences were assembled into 63,401 contigs and 242,212 singletons. BlastX searches for all of the contigs in the GenBank database matched 65% of the contigs to registered sequences, while 35% presented no hits. GO mapping assigned 48% of the 63,401 contigs to GO terms. Microarray analysis showed that subsets of genes were up- or downregulated as the flower developed. The downregulated genes were enriched for GO terms related to 1) oligopeptide transport, 2) response to jasmonic acid stimulus, and 3) cell wall modification, whereas the upregulated genes were enriched for GO terms involved in 1) secondary metabolism, such as the flavonoid biosynthetic process and the terpenoid biosynthetic process, and 2) epidermal cell modification.
2012-09-01 | GSE34332 | GEO
Project description:Genome-wide DNA sequence resources are expected to enhance our understanding of the molecular basis of plant development and lead to improvements in desirable traits in horticultural crops. In this study, we sequenced 643,366 ESTs from Eustoma grandiflorum flowers using a normalised cDNA library constructed for several different flower development stages, times of day, and pollinated stamens. The sequences were assembled into 63,401 contigs and 242,212 singletons. BlastX searches for all of the contigs in the GenBank database matched 65% of the contigs to registered sequences, while 35% presented no hits. GO mapping assigned 48% of the 63,401 contigs to GO terms. Microarray analysis showed that subsets of genes were up- or downregulated as the flower developed. The downregulated genes were enriched for GO terms related to 1) oligopeptide transport, 2) response to jasmonic acid stimulus, and 3) cell wall modification, whereas the upregulated genes were enriched for GO terms involved in 1) secondary metabolism, such as the flavonoid biosynthetic process and the terpenoid biosynthetic process, and 2) epidermal cell modification. Two-condition experiment, loop-design among 4 developmental stages. Biological replicates: 4.
2012-09-01 | E-GEOD-34332 | biostudies-arrayexpress
Project description:The soil worm Enchytraeus crypticus (oligochaete) is an ecotoxicology model species although without genome or transcriptome sequence information. The present research aimed at studying, via high-throughput pyrosequencing, the transcriptome of Enchytraeus crypticus, sampled from multiple test conditions, and the construction of a high-density microarray for functional genomic studies. A pyrosequencing run retrieved approximately 1.5 million reads representing 645 million bases. After assembly, 27,296 contigs and 87,686 singletons were obtained. from which 44% and 25% were annotated as protein-coding genes. We show that the high amount of orphan genes is not due to poor sequence or assemble quality: 84% of the contig sequences contains an open reading frame with a start codon and E. crypticus homologs were identified for 92% of the core eukaryotic genes. Moreover, 65 and 77% of the unknown singletons and contigs, respectively, showed transcriptional activity. An Agilent 180K microarray platform was designed and validated by hybridizing cDNA from 3 day zinc- exposed E. crypticus to the concentration corresponding to 50% reduction in reproduction (EC50). Overall, 70% of all probes exerted a hybridization signal above background level. More specifically, the probes derived from contigs showed a wider range of average intensities when compared to probes derived from singletons. In total, 522 significantly regulated transcripts were identifying upon zinc exposure. Several significantly regulated genes exerted predicted functions (e.g. zinc efflux, zinc transport) associated with zinc stress. Unexpectedly, the microarray data suggest that zinc exposure alters retrotransposon activity in the E. crypticus genome. In conclusion, characterization of the presented E. crypticus transcriptome and associated microarray platform is a valuable and high quality resource that permits further functional genomics experiments examining gene expression patterns underlying distinct environmental stress conditions. We show that unknown sequences are not the result of technical errors but mostly represent functional genes that are actively transcribed. The data presented in our manuscript is part of a larger experiment which was performed in single, large loop design. The analysis presented here can be replicated only by including all raw data from the larger experiment (all raw files are included in the archive linked to this submission). A single channel, interwoven loop design was used to test animals exposed to zinc EC50 on reproduction as compared to untreated controls for 4 days. 4 biological replicates per condition were used containing 25 grams of soil and 5 - 7, adult old animals per replicate. T4_con stands for untreated control soil while T4_50 are the samples exposed to EC50 of zinc on reproduction.
2014-05-01 | E-GEOD-51838 | biostudies-arrayexpress
Project description:Identification of testis-relevant genes using in silico analysis from testes ESTs and cDNA microarray in the black tiger shrimp (Penaeus monodon) A total of 4,803 ESTs from the testis library were unidirectionally sequenced and analyzed in silico using a customizable data analysis package, ESTplus. A total of 2,702 unique sequences from 424 contigs and 2,278 singletons were identified. Approximately, 1,134 sequences from the total of 2,702 unique sequences (contigs and singletons) are homologous to genes with known functions. Some of which (mago nashi, insulin-degrading enzyme, actin-depolymerizing factor) are related to testicular development in studies of other organisms. Moreover, the sequences were further characterized according to the gene ontology categories (41% biological process, 24% molecular function, 35% cellular component). By comparing with the EST libraries of other tissues, a total of 1,579 possible testis-specific ESTs were identified. Some of these testis-specific ESTs, such as Phospholipase A2 and Matrix Metalloproteinases, have functions relevant to testicular development. In addition, novel ESTs (621 ESTs) that have never been identified in any ESTs from the Penaeidae family were found in this study. Through cDNA microarray analysis, some of testis-specific ESTs were preferentially expressed in testes to ovary; one of which was a saposin homolog. Therefore, saposin was further characterized by real-time PCR and its full-length sequence was obtained by RACE-PCR.
2010-11-04 | GSE19037 | GEO
Project description:The high level of human genome structural variation among individuals suggests that there must be portions of the genome that have yet to be discovered, annotated and characterized at the sequence level. Using clone resources developed as part of the Human Genome Structural Variation Sequencing Project, we focused on the characterization of 2,363 novel sequence contigs not present in the human reference genome. We determined that these contigs corresponded to 720 distinct loci of which 400 now have an anchored position in the reference genome. We investigated the sequence properties of these loci and determined that 37% of these novel insertions are copy-number polymorphic. We find that they are significantly enriched within the last 5 Mb of chromosomes (a 2.9-fold enrichment, p=1.0e-18, binomial test) and that most arose as a result of deletions in the human lineage after separation from the African great apes. A subset of these sites shows evidence of marked population stratification among Asian, African and European populations, including a 3.9-kb insertion within the first intron of the lactase gene. Complete sequencing of clones from 192 genomic loci, including 156 completely spanned insertions, provides a detailed and contextual view of 1.67 Mb of inserted sequence. Analysis of this sequence identified 477 elements that show evidence of sequence constraint over evolutionary time, as well as matches to 22 RefSeq gene segments. Twenty-six of the insertions contain matches against mRNA-seq data indicating the potential presence of functionally important, unannotated human sequences. Taking advantage of this high-quality sequence, we develop a method to accurately genotype these novel insertions using next-generation whole-genome sequencing datasets.
2010-04-20 | GSE20634 | GEO
Project description:We wished to determine the changes in gene expression that occurred in S. ratti parasitic females as an infection progressed and thus as these stages are exposed to an anti-S. ratti immune response. To do this we compared gene expression in parasitic females recovered 6 days p.i. (i.e. no or very low immune response) with those recovered at 15 days p.i. (i.e. high immune response); for convenience, we refer to these as parasitic females subject to "low immune pressure" and "high immune pressure", respectively. These days were chosen because previous analyses of S. ratti parasitic females have shown significant differences in the size, appearance etc. of worms at these time points. The experimental design used, was to have at least three biological replicates for each sample (i.e. three independent preparations of the relevant worm samples and their RNA) and to have at least three technical replicates (i.e. independent, separate cDNA synthesis, amplification and hybridization etc.) for each biological replicate. For each hybridisation (below) a dye-swap was used i.e. each sample to be used in a hybridisation was labelled, separately, with each of the two dyes (below).<br> <br> 21,085 ESTs were sequenced from various S. ratti stage-specific libraries, of which 14,761 resulted in sequence data above a quality threshold that were then submitted to public databases. 11,551 clones were derived from the S. ratti parasitic libraries of which 7,385 produced sequence data (above a quality threshold); all of these 7,385 clones were arrayed together with a random sample of 1,619 of clones for which no sequence data were available. These 7,385 ESTs are highly redundant since they represent 2,963 contigs and 2,125 clusters, both including 1,220 singletons (i.e. clusters or contigs containing only one EST). Notwithstanding this redundancy, they were used in the microarray construction for two reasons: (i) this approach was less error-prone than attempting to select a unique clone set and (ii) this in-built redundancy provides many replicates of individual contigs and clusters, which can be exploited in quality-control analyses. In addition to these 9,004 S. ratti clones, the following controls were included: 281 EST clones from the mixed iL3/free-living adult library (representing 173 contigs and 167 clusters, 67 of which are singletons) to ensure that gene expression in the parasitic and free-living stages could be differentiated; 230 commercially available controls (Amersham Biosciences UK, Ltd.). 12 poly-A and 457 spotting buffer-only controls. Thus, in total 9,984 spots were arrayed.
2007-04-07 | E-MEXP-1010 | biostudies-arrayexpress