Project description:Transcriptional profiling of mouse lung tumors comparing Dnmt3a KO/K-ras G12D mutant with Dnmt3a WT/K-ras G12D mutant. The goal is to search for the difference of mRNA abundance between Dnmt3a KO and WT tumors. Two-condition experiment, KO vs. WT tissure. Biological replicates: 12.
Project description:Transcriptional profiling of mouse lung tumors comparing Dnmt3a KO/K-ras G12D mutant with Dnmt3a WT/K-ras G12D mutant. The goal is to search for the difference of mRNA abundance between Dnmt3a KO and WT tumors.
Project description:Gains and losses in DNA methylation are prominent genomic features of all mammalian cell types. To gain insight into mechanisms that could promote shifts in DNA 5-hydroxymethylation (5hmc) patterns and thus contribute to cell fate, including malignant transformation, we performed genome-wide mapping of 5hmc in purified wild type and Dnmt3a conditional knockout hematopoietic stem cells (HSCs) using cytosine-5-methylenesulphonate sequencing (CMS-seq). Comparing anti-CMS pulldown to input control, we identified 107,549 and 175,682 peaks of 5hmc enrichment in control and Dnmt3a knockout HSCs, respectively. Whole genome CMS-enriched bisulfite sequencing of secondarily-transplanted wild-type and Dnmt3a conditional knockout hematopoietic stem cells using Illumina HiSeq 2000
Project description:Transcriptional profiling of mouse postnatal SVZ NSCs comparing WT NSCs with KO NSCs under proliferating/undifferentiated states as well as differentiating conditions. Goal was to determine Dnmt3a-dependent gene expression changes in postnatal SVZ NSCs Two-condition experiment with a dye-swap design, WT NSCs vs. KO NSCs. Biological replicates: 4 replicates under proliferating/undifferentiation conditions, 2 replicates under differentiating conditions.
Project description:ra05-11_cmsrapeseed - rfppr - -To understand how/if fertility and the mitochondrial background are interlinked. -To understand how the mitochondrial background influences the nuclear gene expression. -Compare and describe the total nuclear gene expression of CMS vs. fertile and CMS vs. Restored. -Describe and analyse genes that differ in expression (of special interest are genes that differs in both comparisons). -Group genes e.g. floral genes, highly expressed genes, transcription factors, nuclear encoded genes targeted for the mitochondrion. -Comparing two CMS-systems to elucidate differences and similarities between them. - Flower buds from three B. napus lines (Pactol, CMS, Rfo PPR) at one developmental stage (stage 8). Keywords: wt vs mutant comparison 6 dye-swap - CATMA arrays
Project description:ra05-11_cmsrapeseed - rfppr - -To understand how/if fertility and the mitochondrial background are interlinked. -To understand how the mitochondrial background influences the nuclear gene expression. -Compare and describe the total nuclear gene expression of CMS vs. fertile and CMS vs. Restored. -Describe and analyse genes that differ in expression (of special interest are genes that differs in both comparisons). -Group genes e.g. floral genes, highly expressed genes, transcription factors, nuclear encoded genes targeted for the mitochondrion. -Comparing two CMS-systems to elucidate differences and similarities between them. - Flower buds from three B. napus lines (Pactol, CMS, Rfo PPR) at one developmental stage (stage 8). Keywords: wt vs mutant comparison
Project description:This SuperSeries is composed of the following subset Series: GSE32415: Mouse lung tumor tissue: Dnmt3a KO vs. Dnmt3a WT GSE32484: DNA methylation of dnmt3a deficient Kras lung tumors and dnmt3a wt Kras lung tumors Refer to individual Series
Project description:Gains and losses in DNA methylation are prominent genomic features of all mammalian cell types. To gain insight into mechanisms that could promote shifts in DNA 5-hydroxymethylation (5hmc) patterns and thus contribute to cell fate, including malignant transformation, we performed genome-wide mapping of 5hmc in purified wild type and Dnmt3a conditional knockout hematopoietic stem cells (HSCs) using cytosine-5-methylenesulphonate sequencing (CMS-seq). Comparing anti-CMS pulldown to input control, we identified 107,549 and 175,682 peaks of 5hmc enrichment in control and Dnmt3a knockout HSCs, respectively.
Project description:Since DNMT3A is a de novo DNA methyltransferase, transcriptional differences in WT versus Dnmt3a-/- HSCs upon infection suggest a possible role for epigenetic regulation in the HSC transcriptional response to inflammation. Therefore, we wanted to investigate if differences in stress and presence of Dnmt3a affect the methylome overall, and specific IFNg responses genes. 100000 LT-HSCs (LK CD150+ CD48- were sorted into lysis buffer from the pools of naive or 1-month (M. avium) infected WT/ Dnmt3-/-_x0001_ mice (n = 7-8 per group). Around 300 ng of DNA per group was extracted with AllPrep DNA/RNA Mini Kit. NEBNext Ultra II DNA library prep kit was used for library preparation. Bisulfite conversion step was added after ‘‘methylated’’ adaptor ligation and USER excision. Then, methylated adaptor ligated DNA fragment was treated with bisulfite conversion using EZ DNA methylationlightning kit. These modified DNA fragments were then amplified with index primers with Kapa HiFi urasil+ specialized polymerase for bisulfite converted DNA. For WGBS library sequencing, 150-high output kit from illumine was used. Samples (400 million reads/ sample (15-20X coverage)) were run in NovaSeq S1 FC (2, lanes 1,600 Million reads). Global methylation of Dnmt3a-/- cells was more divergent upon infection compared to WT. There were vastly more hyper- and hypomethylated regions in infected versus naive Dnmt3a-/- HSCs compared to WT. Among the altered regions, both WT and Dnmt3a-/- HSCs showed slightly more hypomethylated than hypermethylated regions, although changes in both directions were present. In the WT background, regions of hyper- and hypomethylation were mostly restricted to CpG islands. By contrast, highly significant differences in hypomethylation were found in the Dnmt3a-/- HSCs in CpG islands, shores, enhancers, and promoters DNMT3A is highly active in HSCs during infection and that the loss of DNMT3A function results in significant global changes in methylation, including both hyper- and hypomethylation, in genome regions that likely affect gene transcription. we examined the methylation of Batf2, Jun, and Fos. All 3 of these prodifferentiation genes showed increased methylation in the promoter region in the Dnmt3a-/- background as compared to the WT background. Hypermethylation of these promoter regions is consistent with their transcriptional repression.
Project description:To identify soluble factors released from Dnmt3a KO macrophages that might drive differences in osteoclast differentiation, we performed RNA sequencing on unstimulated BMDMs from WT and Dnmt3a KO mice. To investigate the mechanistic basis for the inflammatory phenotype. of myeloid cells with loss of Dnmt3a,we performed reduced representation bisulfite sequencing (RRBS) on Dnmt3a KO and WT BMDMs to assess methylation changes. Concurrently, we performed ATAC-sequencing (ATAC-seq) to assess differences in chromatin accessibility between WT and Dnmt3a KO BMDMs in the context of changes in methylation. We performed chromatin immunoprecipitation sequencing (CHIP-seq) for Irf3 and Rela based on ATAC-seq analysis.