Effect of Humanin treatment on brain tumor cell lines
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ABSTRACT: RNA sequencing analysis was performed on mouse cell samples. Quality control of total RNA samples after RNA isolation with the Agilent Bioanalyzer revealed acceptable RNA quality for provided samples. Libraries were successfully prepared from these RNA samples using the Illumina TruSeq® Stranded mRNA technology. This approach uses fragmentation and sequencing adapter ligation in combination with a poly-T oligo pulldown. RNA sequencing was performed on the Illumina NextSeq500 next generation sequencing system (Illumina) and its high output mode with two runs of 1x75 bp single-end read chemistry and a high amount of high quality reads was generated.
Project description:Amplicon-based targeted re-sequencing analysis was performed in the patient-derived gliobastoma cell culture samples. For this purpose, genomic DNA (gDNA) was isolated and DNA libraries were prepared using the TruSeq Custom Amplicon Low Input (Illumina, Inc.) technology. By this, a pool of 375 amplicons was generated for each single sample in order to enrich for the target genes ATRX1, EGFR, IDH1, NF1, PDGFRA, PIK3CG, PIK3R1, PTEN, RB1 and TP53. Sequencing was performed on the Illumina MiSeq® next generation sequencing system (Illumina Inc.) and its 2 x 250 bp paired-end v2 read chemistry. The resulting reads were quality controlled and mapped against the human reference genome (hg19). For all samples, sequence variations of the amplified regions of interest in comparison to the human reference sequence were identified and filtered based on reliability.
Project description:RNA sequencing analysis was performed on mouse cell samples. Quality control of total RNA samples after RNA isolation with the Agilent Bioanalyzer revealed acceptable RNA quality for 12 out of 30 provided samples. Libraries were successfully prepared from these RNA samples using the Illumina TruSeq® Stranded mRNA technology. This approach uses fragmentation and sequencing adapter ligation in combination with a poly-T oligo pulldown. RNA sequencing was performed on the Illumina NextSeq500 next generation sequencing system (Illumina) and its high output mode with two runs of 1x75 bp single-end read chemistry and a high amount of high quality reads was generated.
Project description:Genome-wide copy number variation profiles were analyzed in the patient-derived gliobastoma cell culture samples. For this purpose, gDNA was analyzed using the CytoScan® assay in combination with a one-color based labeling and hybridization protocol. Signals on the CytoScan® HD microarrays were detected using the Affymetrix GeneChip® 3000 Scanner.
Project description:Genome-wide copy number variation profiles were analyzed in the patient-derived gliobastoma cell culture samples. For this purpose, gDNA was analyzed using the CytoScan® assay in combination with a one-color based labeling and hybridization protocol. Signals on the CytoScan® HD microarrays were detected using the Affymetrix GeneChip® 3000 Scanner.
Project description:MeRIP-Seq data aligned to the genome (GRCh38) for cells with IDH1-Mut or IDH1-WT genotypes. Aligned data (BAM) are separated into input RNA and m6A immunoprecipitated RNA for each cell sample.
Project description:Eukaryotic topoisomerase 1 (TOP1) regulates DNA topology to ensure efficient DNA replication and transcription. TOP1 is also a major driver of endogenous genome instability, particularly when its catalytic intermediate - a covalent TOP1-DNA adduct known as a TOP1 cleavage complex (TOP1cc) - is stabilised. TOP1ccs are highly cytotoxic and a failure to resolve them underlies the pathology of neurological disorders but is also exploited in cancer therapy where TOP1ccs are the target of widely used frontline anti-cancer drugs. A critical enzyme for TOP1cc resolution is the tyrosyl-DNA phosphodiesterase, TDP1, which hydrolyses the bond that links a tyrosine in the active site of TOP1 to a 3’ phosphate group on a single-stranded (ss)DNA break. However, TDP1 can only process small peptide fragments from ssDNA ends, raising the question of how the ~90 kDa TOP1 protein is processed upstream of TDP1. We find that TEX264 fulfils this role by forming a complex with the p97 ATPase and the SPRTN metalloprotease. We show that TEX264 recognises both unmodified and SUMO1-modifed TOP1 and initiates TOP1cc repair by recruiting p97 and SPRTN. TEX264 localises to the nuclear periphery, associates with DNA replication forks, and counteracts TOP1ccs during DNA replication. Altogether, our study elucidates the existence of a specialised repair complex required for upstream proteolysis of TOP1ccs and their subsequent resolution.
Project description:Bovine tuberculosis (bTB), caused by Mycobacterium bovis, represents a significant issue for the global agriculture industry as well as for human health. Epigenetic modifications can alter the course of the immune response and differentially methylated regions (DMRs) could contribute to the failure of current generation tests to detect all TB-infected cattle. Whole Genome Bisulphite Sequencing (WGBS) was used to profile DNA methylation levels from peripheral blood of cattle naturally infected with M. bovis (positive for the single intradermal comparative tuberculin test (SICTT) and/or the interferon-γ release assay (IGRA) compared to negative controls [n=8/group, total of 16 WGBS libraries]. Although overall methylation profiles were similar across the genome, 224 DMRs and 159 Differentially Methylated Promoter Gene (DMPGs) were identified between groups with an excess of hypermethylated sites in bTB+ infected cattle (threshold >15% differential methylation). Genes located within these DMRs included the Interleukin 1 receptor (IL1R1) and MHC related genes (BOLA and BOLA-DQB) which may influence effective immunity. KEGG pathway analysis identified enrichment of genes involved in Calcium and MAPK signalling, as well as metabolism pathways. Analysis of DMRs in a subset of SICTT- cattle (n=4 group) which were IGRA+, and thereby potentially represent a risk for disease recurrence on farms, showed differential methylation of genes including Interleukin 10 Receptor, alpha (IL10RA), Interleukin 17F (IL17F) and host defence peptides (DEFB and BDEF109). This study has identified a number of immune gene loci at which differential methylation could impact on the host immune response and the ability of cattle to clear infection. Differential methylation of immune gene loci is an informative avenue to improve our understanding of the regulation of host immunity as well as the role of methylation on diagnostic test performance.
Project description:Bulk RNA-seq data from differentiating embryoid bodies made from wild-type and MEIS1/2 knockout HES3 hESCs, collected at day 5 and 7 of differentiation. The aim of the experiments was to identity differentially expressed genes and therefore putative MEIS targets.
Project description:Bulk RNA-seq data of HES3 hESCs sampled through a differentiation timecourse. Sample collection was at day 0, 3.75, 4.75, 5.75 and 12. There are additional day 5.75 samples with MYC transgene overexpression (activated from day 4.75).
Project description:The aim of the experiment was to identify HAND1 target genes and its impact on chromatin accessibility in relation to cardiac development. A HAND1-null hESC line was used, in which a doxycycline-inducible HAND1-T2A-BFP transgene had been integrated in approximately half of the cells for HAND1 rescue / overexpression. The hESCs were differentiated with BMP4, Activin A and CHIR. On day 2.5, doxycycline was added. On day 3, cells were dissociated and sorted by BFP level using FACS. Samples were immediately processed for RNA-seq and ATAC-seq.