Project description:The aim of our study is to identify the role of FUS in shaping the transcriptome. RNA-seq of two FUS KO clones was performed and compared to wt; for each, four replicates were sequenced. RNA molecules associated with the FUS protein were determined by means of a RNA immuno-precipitation, followed by high-throughput sequencing. Total RNA was used as a control. SH-SY5Y cells were used for both experiments. RNA-seq: 4 wt samples, 4 A4 KO samples, 4 A5 KO samples. RIP-seq: 1 input control sample, 3 anti-FUS IP replicates.
Project description:We compared the epigenetic status of the mutant and disease-free iPSCs at the whole genome level. Whole epigenome profiling based on trimethylated H3K4 (H3K4me3) showed concordant epigenetic remodeling in the two corrected clones when compared with two mutant iPSC clones. Examination of genome-wide gene expression in Fanconi anemia patient iPSCs before and after gene correction
Project description:We compared the epigenetic status of the mutant and disease-free iPSCs at the whole genome level. Whole epigenome profiling based on trimethylated H3K4 (H3K4me3) showed concordant epigenetic remodeling in the two corrected clones when compared with two mutant iPSC clones. Examination of the trimethylated H3K4 histone modification in Fanconi anemia patient iPSCs before and after gene correction
Project description:To address how Csf3r and RUNX1 mutations in combination with CSF3 administration affect hematopoiesis in vivo, we performed serial transplantation experiments. Lineage-negative Csfr-d715 BM cell cells were lentivirally transduced with the patient specific RUNX1-D171N (RHD) mutant or an empty vector control. Primary recipients showed a pre-leukemic condition characterised by myeloblasts in the peripheral blood, but not overt AML. Upon serial transplantation, one of the Csfr-d715/RUNX1-D171N mutant mice, treated with CSF3, could repopulate secondary and tertiary recipients. Whole exome sequencing on these mice were performed to investigate whether these mice acquired an additional mutation. Enzymatically fragmented genomic DNA was used to construct sample libraries following the SeqCap EZ HyperPlusCap workflow User’s Guide version 1.0 (Roche). Unique, dual index adapters (Integrated DNA technologies) were used for ligation. After ligation of adapters and an amplification step, exome target sequences were captured using in-solution oligonucleotide baits (SeqCap EZ Developer Library mm9_exome_L2R_D02). Amplified captured sample libraries were paired-end sequenced on the HiSeq 2500 platform (Illumina).
Project description:Homologous recombination (HR) is a Rad51-mediated evolutionary conserved process that plays a unique role in genome plasticity, controlling the balance genetic stability/diversity. In this study we compared the induction of genetic modifications by whole exome sequencing in primary mouse embryonic fibroblasts after the expression of a dominant negative form of Rad (SMRad51). We used a transgenic mouse embryonic fibroblasts in which SMRad51 expression was induced under doxycycline (Dox) control. We compared SMRad51 primary mouse embryonic fibroblasts treated with Dox (transgene expression induced) with control mouse embryonic fibroblasts treated with Dox.
Project description:In the present study we created and analyzed cardiomyocytes from two separate iPSC clones from the fibroblasts of five different female individuals and two male individuals, using footprint-free Sendai virus RNA-seq of iPSC cardiomyocytes, Ampli-seq of heart left ventricle and iPSC cardiomyocytes (with and without drug treatment) and Exome-seq of patient fibroblasts.
Project description:CSF-1R is recruited on EGR1 motifs in monocytes where it colocalizes with EGR1. To address if EGR1 required for CSF-1R recruitment on chromatin, THP-1 monocytic cell line has been deleted for EGR1 by CRISPR-Cas9 approach. 3 clones were generated by single cell cloning and CSF-1R localization on chromatin was compared to two unmodified THP-1 clones by ChIP-sequencing. Since the read number was strongly decreased in the EGR1-deleted clones, the three clones were pooled for the comparison with wild-type clones. ChIPseq of CSF-1R (Nter Antibody) in monocytes of two CMML patients (CMML2130 and CMML2609)
Project description:We have earlier reported the establishment of a comprehensive in vitro panel of 19 isogenic cell lines from a patient with Grade IV serous adenocarcinoma (Bapat et al. Cancer Research. 2005). In this study, one of the cell lines termed as A4 at early passes (less than ~20) was assessed as being non-tumorigenic since it failed to exhibit anchorage independence growth in soft agar and form tumors in nude mice. After further culture (more than 20 passages in vitro), the cells expressed all features suggestive of transformation including capabilities of anchorage independence growth, clonogenecity and reproducibility of the human disease in mice models as assessed through tumor and ascites formation in nude mice, and serial transplantion of tumor-derived cells over 3-4 mouse generations. The tumors generated in nude mice were also identified to be histologically similar to the primary tumor of the patient. A4 cells thus represent a single tumorigenic clone that underwent transformation in vitro, and hence may be considered to be an ovarian tumor progression model. Overall design: Two-condition experiment, Samples for hybridization included – (i) Non-tumorigenic A4 cells at passage 15 that were designated A4p15 – RNA labeled with Cy3; and (ii) Transformed A4 cells at passage 84 that were designated A4p84 – RNA labeled with Cy5. Three samples of each were chosen for hybridization.