Project description:To identify FHY3 associated genes in floral organ. We performed ChIP-seq using 35S:3FLAG-FHY3-3HA fhy3-4 tansgenic plants. A total of 21, 24 and 37 million reads were obtained from two replicates and input library, respectively, which were uniquely mapped to the Arabidopsis genome by Bowtie2 software resulting in 1885 FHY3 binding peaks. 84% of FHY3 binding sites (1580 loci) were subsequently assigned to genic regions and grouped into 2192 genes in flower.
Project description:To identify FHY3 regulated genes in floral organ. We performed RNA-seq using Ler, ag-10, fhy3-68, ag-10 fhy3-68. Inflorescence of Ler, ag-10, fhy3-68, ag-10 fhy3-68 containing stage 8 and younger flowers were harvested forRNA-seq analysis two distinct biological replicates were subjected to ultra-high-throughput Solexa (Illumina) sequencing.
Project description:To identify FHY3 associated genes in floral organ. We performed ChIP-seq using 35S:3FLAG-FHY3-3HA fhy3-4 tansgenic plants. A total of 21, 24 and 37 million reads were obtained from two replicates and input library, respectively, which were uniquely mapped to the Arabidopsis genome by Bowtie2 software resulting in 1885 FHY3 binding peaks. 84% of FHY3 binding sites (1580 loci) were subsequently assigned to genic regions and grouped into 2192 genes in flower. Inflorescence of 35S:3FLAG-FHY3-3HA fhy3-4 containing stage 8 and younger flowers were harvested for ChIP-seq analysis with anti-FLAG antibodies. No antibody served as negative control. The chromatin DNA from two distinct biological replicates and an input DNA sample were subjected to ultra-high-throughput Solexa (Illumina) sequencing.
Project description:Retinoblastoma (pRb) is a multifunctional regulator, which was likely present in the last common ancestor of all eukaryotes. The Arabidopsis pRb homolog RETINOBLASTOMA RELATED 1 (RBR1), similar to its animal counterparts, controls not only cell proliferation but is also implicated in developmental decisions, stress responses and maintenance of genome integrity. Although most functions of pRb-type proteins involve chromatin association, a genome-wide understanding of RBR1 binding sites in Arabidopsis is still missing. Here, we present a plant chromatin immunoprecipitation protocol optimized for genome-wide studies of indirectly DNA-bound proteins like RBR1. Our analysis revealed binding of Arabidopsis RBR1 to approximately 1000 genes and roughly 500 transposable elements, preferentially MITES. The RBR1-decorated genes broadly overlap with previously identified targets of two major transcription factors controlling the cell cycle, i.e. E2F and MYB3R3 and represent a robust inventory of RBR1-targets in dividing cells. Consistently, enriched motifs in the RBR1-marked domains include sequences related to the E2F consensus site and the MSA-core element bound by MYB3R transcription factors. Following up a key role of RBR1 in DNA damage response, we performed a meta-analysis combining the information about the RBR1-binding sites with genome-wide expression studies under DNA stress. As a result, we present the identification and mutant characterization of three novel genes required for growth upon genotoxic stress.
Project description:CodY is a global transcriptional regulator that controls, directly or indirectly, the expression of dozens of genes and operons in Listeria monocytogenes. We used in vitro DNA affinity purification combined with massively parallel sequencing (IDAP-Seq) to identify genome-wide L. monocytogenes chromosomal DNA regions that CodY binds in vitro. The total number of CodY-binding regions exceeded 2,000, but they varied significantly in their strengths of binding at different CodY concentrations. The 388 strongest CodY-binding regions were chosen for further analysis. A strand-specific analysis of the data allowed pinpointing CodY-binding sites at close to single-nucleotide resolution. Gel shift and DNase I footprinting assays confirmed the presence and locations of several CodY-binding sites. Surprisingly, most of the sites were located within genes' coding regions. The binding site within the beginning of the coding sequence of the prfA gene, which encodes the master regulator of virulence genes, has been previously implicated in regulation of prfA, but this site was weaker in vitro than hundreds of other sites. The L. monocytogenes CodY protein was functionally similar to Bacillus subtilis CodY when expressed in B. subtilis cells. Based on the sequences of the CodY-binding sites, a model of CodY interaction with DNA is proposed.