Project description:An exponentially growing culture of strain R6 in AGCH at OD620nm=0.4 was either non-treated or treated with two LVX concentrations: 0.125 ug/ml LVX (0.5x MIC of R6) and 2.5 ug/ml LVX (10x MIC of R6). Samples were taken before treatment (0 min), at 15, 30 and 60 min in the non-treated culture, and at 5, 15 ,30 and 60 min in the LVX treated cultures
Project description:The oncomir microRNA-125b (miR-125b) is up-regulated in a variety of human neoplastic blood disorders and constitutive up-regulation of miR-125b in mice can promote myeloid and B cell leukemia. We found that miR-125b promotes myeloid and B cell neoplasm by inducing tumorigenesis in hematopoietic progenitor cells. Our study demonstrates that miR-125b induces myeloid leukemia by enhancing myeloid progenitor output from stem cells as well as inducing immortality, self-renewal, and tumorigenesis in myeloid progenitors. Through functional and genetic analyses, we demonstrated that miR-125b induces myeloid and B cell leukemia by inhibiting IRF4 but through distinct mechanisms; it induces myeloid leukemia through repressing IRF4 at the mRNA level without altering the genomic DNA and induces B cell leukemia via genetic deletion of the gene encoding IRF4. The cancer myeloid (Cd11b+ sorted) and B cells (CD19+ sorted) were harvested from mice that over-express miR-125b. The genomic DNA was extracted from these cells. A total of 4 cancer samples (Two myeloid cancer samples and two B cell cancer samples) were analyzed. As control, genomic DNA from cells harvested from healthy C57bl/6 mice were harvested.
Project description:Following the domestication of maize over the past ,10,000 years, breeders have exploited the extensive genetic diversity of this species to mold its phenotype to meet human needs. The extent of structural variation, including copy number variation (CNV) and presence/absence variation (PAV), which are thought to contribute to the extraordinary phenotypic diversity and plasticity of this important crop, have not been elucidated. Whole-genome, array-based, comparative genomic hybridization (CGH) revealed a level of structural diversity between the inbred lines B73 and Mo17 that is unprecedented among higher eukaryotes. A detailed analysis of altered segments of DNA conservatively estimates that there are several hundred CNV sequences among the two genotypes, as well as several thousand PAV sequences that are present in B73 but not Mo17. Haplotype-specific PAVs contain hundreds of single-copy, expressed genes that may contribute to heterosis and to the extraordinary phenotypic diversity of this important crop. In our experimental design we had seven replicates of B73 (one with Cy3 and six with Cy5) and seven replicates of Mo17 (six with Cy3 and one with Cy5). Images were processed and spatial normalization of data within the array was conducted according to Nimblegen's standard protocol. The RIL samples (M0022 and M0023) were in included because we used the probe's B73 and Mo17 signals of those samples during our analysis.
Project description:Maintenance of chromatin structure is essential to eukaryotic life; dysregulation is known to be causal for aberrant development and disease. The Mi-2/nucleosome remodeling and histone deacetylase (NuRD) complex is a multiprotein machine proposed to regulate chromatin structure by nucleosome remodeling and histone deacetylation activities. We identified the localization of MBD3, a component of Mi-2/NuRD complex, in two breast cancer cell lines (MCF7 and MDA-MB-231) using ChIP-Seq. MBD3 showed cell-type specific localization with overlap across cell lines being less than 50%. MBD3 localized across gene bodies, peaking around the transcription start site (TSS). Contrary to existing models, MBD3 preferentially associated with CpG rich promoters marked by H3K4me3. These data suggest that MBD3, and by extension the Mi-2/NuRD complex, may have roles in fine tuning expression for active genes. These data represent an important first step in defining regulatory mechanisms by which Mi-2/NuRD complex controls chromatin structure and gene expression. DamID experiment was performed in human breast cancer cell lines (MCF-7 and MDA-MB-231) in triplicate. Samples were hybridized to NimbleGen Human Whole-Genome Tiling Arrays (0701_HG18_TILE_05_HX1 and 100718_HG18_TILE_05_HX1). MBD3-Dam material was hybridized over Dam-only control.
Project description:Maintenance of chromatin structure is essential to eukaryotic life; dysregulation is known to be causal for aberrant development and disease. The Mi-2/nucleosome remodeling and histone deacetylase (NuRD) complex is a multiprotein machine proposed to regulate chromatin structure by nucleosome remodeling and histone deacetylation activities. We identified the localization of MBD3, a component of Mi-2/NuRD complex, in two breast cancer cell lines (MCF7 and MDA-MB-231) using ChIP-Seq. MBD3 showed cell-type specific localization with overlap across cell lines being less than 50%. MBD3 localized across gene bodies, peaking around the transcription start site (TSS). Contrary to existing models, MBD3 preferentially associated with CpG rich promoters marked by H3K4me3. These data suggest that MBD3, and by extension the Mi-2/NuRD complex, may have roles in fine tuning expression for active genes. These data represent an important first step in defining regulatory mechanisms by which Mi-2/NuRD complex controls chromatin structure and gene expression. DamID experiment was performed in human breast cancer cell lines (MCF-7 and MDA-MB-231) in duplicate. Samples were hybridized to Nimblegen 2.1M Deluxe promoter array. MBD3-Dam material was hybridized over Dam-only control.
Project description:The goal of the study was to identify subsets of the genes, which are up-regulated and down-regulated by CovR regulator in UA159 strain. Two independently isolated RNA samples from each of the wild-type UA159 strain and its covR mutant strain grown until mid-exponential phase were analyzed. Each sample was hybridized with 5 blocks of NimbleGen arrays.
Project description:Nuclear pores associate with active protein-coding genes in yeast and have been implicated in transcriptional regulation. Here, we show that in addition to transcriptional regulation, key components of C. elegans nuclear pores are required for processing of a subset of small nucleolar RNAs (snoRNAs) and tRNAs transcribed by RNA Polymerase (Pol) III. Chromatin immunoprecipitation of NPP-13 and NPP-3, two integral nuclear pore components, and importin-M-CM-^_ IMB-1, provides strong evidence that this requirement is direct. All three proteins associate specifically with tRNA and snoRNA genes undergoing Pol III transcription. These pore components bind immediately downstream of the Pol III pre-initiation complex, but are not required for Pol III recruitment. Instead, NPP-13 is required for cleavage of tRNA and snoRNA precursors into mature RNAs, whereas Pol II transcript processing occurs normally. Our data suggest that integral nuclear pore proteins act to coordinate transcription and processing of Pol III transcripts in C. elegans. Genome-wide ChIP-seq and ChIP-chip were performed in mixed-stage C. elegans embryos for nuclear pore proteins NPP-13, NPP-3, IMB-1 and chromatin proteins Pol III (RPC-1), TBP-1, TFC-1 (SFC-1), TFC-4 (TAG-315), and Pol II (AMA-1). For RPC-1 and TBP-1 ChIP-seq, embryos depleted for NPP-13 were also used. Total RNAs from wild-type, NPP-13 RNAi, and IMB-1 RNAi embryos were analyzed by RNA-seq.