The RNA-binding protein FUS is implicated in transcription, alternative splicing of neuronal genes and DNA repair. Mutations in FUS have been linked to human neurodegenerative diseases such as ALS (amyotrophic lateral sclerosis). We genetically disrupted fus in zebrafish (Danio rerio) using the CRISPR-Cas9 system. The fus knockout animals are fertile and did not show any distinctive phenotype. Mutation of fus induces mild changes in gene expression on the transcriptome and proteome level in the adult brain. We observed a significant influence of genetic background on gene expression and 3’UTR usage, which could mask the effects of loss of Fus. Unlike published fus morphants, maternal zygotic fus mutants do not show motoneuronal degeneration and exhibit normal locomotor activity. Overall design: We performed paired-end sequencing (100bp reads) of the polyA+ transcriptome from brains of five individuals with Fus-/- genotype and four with Fus wild type genotype. Note on RNA-Seq replicates: after performing first RNA sequencing on four replicates of Fus-/- and WT (labeled with the prefix "Sample_imb_ketting_2014_13_") we received a notice from Illumina stating a problem with the library preparation kit lot that was used to prepare the libraries. Due to that, we performed RNA sequencing a second time, using the same input RNA, except for the Fus knockout replicate #3, because there was not enough input RNA left. Instead, a different Fus knockout replicate (#1) was sequenced. However, we compared the mapped reads from sequencing run 1 and sequencing run 2 using plotCorrelaction from DeepTools, and the samples are highly correlated (at least 0.97 and 0.95, Spearman and Pearson correlation respectively). Therefore, we considered first ("Sample_imb_ketting_2014_13_") and second sequencing runs as technical replicates.
Characterization of genetic loss-of-function of Fus in zebrafish.
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View SamplesThe aim of this work was to access the early immune response triggered by R. microplus larvae attachment in previously selected resistant and susceptible animals in a bovine F2 population derived from Gyr (Bos indicus) Holstein (Bos taurus) crosses.
Microarray analysis of tick-infested skin in resistant and susceptible cattle confirms the role of inflammatory pathways in immune activation and larval rejection.
Specimen part, Time
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Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response.
Age, Specimen part, Time
View SamplesMicroarray analyses of sweet orange epicotyls transiently transfected with the pthA2, pthA4 or pthC1 gene, relative to epicotyls transfected with the uid gene (GUS)
Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response.
Age, Specimen part, Time
View SamplesMicroarray analyses of sweet orange leaves infiltrated with Xc in the presence or absence of Ch, or Ch alone
Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response.
Specimen part, Time
View SamplesCurrent pipelines used to map genetrap insertion sites are based on inverse- or splinkerette-PCR methods, which despite their efficacy are prone to artifacts and do not provide information on the impact of the genetrap on the expression of the targeted gene. We developed a new method, which we named TrapSeq, for the mapping of genetrap insertions based on paired-end RNA sequencing. By recognizing chimeric mRNAs containing genetrap sequences spliced to an endogenous exon, our method identifies insertions that lead to productive trapping. Overall design: We conducted two independent screenings for sensitivity against 6-thioguanine (6TG) and an ATR inhibitor (ATRi). We applied our RNAseq-based pipeline (TrapSeq) to identify mutations that provide resistance to these reagents. Importantly, and besides its use for screenings, when applied to individual clones our method provides a fast and cost-effective way that not only identifies the insertion site of the genetrap but also reveals the impact of the insertion on the expression of the trapped gene. Please note that HAP1, haploid for all chromosomes, derives from near-haploid KBM7 parent line which was in turn obtained from a chronic myeloid leukemia patient in blast crisis phase (Carette et al. Nature 477:340-343, 2011).
Trap<sup>Seq</sup>: An RNA Sequencing-Based Pipeline for the Identification of Gene-Trap Insertions in Mammalian Cells.
Specimen part, Cell line, Subject
View SamplesBackground: Arrhythmogenic cardiomyopathy (ACM) is a genetic autosomal disease characterized by abnormal cell-cell adhesion, cardiomyocyte death, progressive fibro-adipose replacement of the myocardium, arrhythmias and sudden death. Several different cell types contribute to the pathogenesis of ACM, including, as recently described, cardiac stromal cells (CStCs). In the present study, we aim to identify ACM-specific expression profiles of human CStCs derived from endomyocardial biopsies of ACM patients and healthy individuals employing TaqMan Low Density Arrays for miRNA expression profiling, and high throughput sequencing for gene expression quantification. Results: We identified 5 miRNAs and 272 genes as significantly differentially expressed. Both the differentially expressed genes as well as the target genes of the ACM-specific miRNAs were found to be enriched in cell adhesion related biological processes. Functional similarity and protein interaction based network analyses performed on the identified deregulated genes, miRNA targets and known ACM-causative genes revealed clusters of highly related genes involved in cell adhesion, extracellular matrix organization, lipid transport and ephrin receptor signaling. Conclusions: We determined for the first time the coding and non-coding transcriptome characteristic of ACM cardiac stromal cells, finding evidence for a potential contribution of miRNAs to ACM pathogenesis or phenotype maintenance. Besides known pathways, we identified also deregulation of genes encoding ephrin receptors and ephrins, thus suggesting a potential involvement of Eph-ephrin signaling in CStCs from ACM hearts. Overall design: Expression profiles of cardiac stromal cells from 3 ACM patients were compared against those of cardiac stromal cells from 3 healthy individuals.
The arrhythmogenic cardiomyopathy-specific coding and non-coding transcriptome in human cardiac stromal cells.
Sex, Disease, Subject
View SamplesGenetic factors contribute to the development of ischemic stroke but their identity remains largely unknown. We tested the association with ischemic stroke of 210 single nucleotide polymorphisms (SNPs) associated with pathways functionally related to stroke. We observed an association between the rs7956957 SNP in LRP1 and next performed microarrays analysis in healthy individuals to investigate possible associations of LRP genotypes with the expression of other genes.
Brain perihematoma genomic profile following spontaneous human intracerebral hemorrhage.
Sex, Age, Specimen part
View SamplesSpontaneous intracerebral hemorrhage (ICH) represents about 15% of all strokes and is associated with high mortality rates. Our aim was to identify the gene expression changes and biological pathways altered in the brain following ICH.
Brain perihematoma genomic profile following spontaneous human intracerebral hemorrhage.
Sex, Age, Specimen part
View SamplesWe performed a microarray experiment to assess the global changes in transcription occurring in leaves and roots of the vitamin B6 deficient pdx1.3 knockout mutant in comparison to WT. Vitamin B6 (pyridoxal 5-phosphate) is an essential cofactor of many metabolic enzymes. Plants biosynthesize the vitamin de novo employing two enzymes, pyridoxine synthase1 (PDX1) and PDX2. In Arabidopsis (Arabidopsis thaliana), there are two catalytically active paralogs of PDX1 (PDX1.1 and PDX1.3) producing the vitamin at comparable rates. Since single mutants are viable but the pdx1.1 pdx1.3 double mutant is lethal, the corresponding enzymes seem redundant.
Consequences of a deficit in vitamin B6 biosynthesis de novo for hormone homeostasis and root development in Arabidopsis.
Specimen part
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