Project description:MBL-1 is an RNA-binding protein important for the differentiation and development of many animal tissues. Our aim was to identify the binding targets of the MBL-1 protein in C. elegans. RNA from a CRISPR/CAS9-generated strain expressing a 3xFLAG-tagged MBL-1 protein was immunopreciprecipitated with an anti-FLAG antibody. RNA-seq was then performed to identify binding targets of the RNA-binding protein MBL-1. Experiments were performed at the L4 stage.
Project description:To identify Smaug’s target mRNAs on a genome-wide scale we used ribonucleoprotein (RNP) co-immunoprecipitation followed by microarray analysis of the co-purifying mRNAs (RIP-Chip). Extracts, prepared from wild-type embryos collected 0-3 hours post-egglaying, were immunoprecipitated with an anti-Smaug antibody (Smaug RIPs) while control immunoprecipitations using non-immune serum served as a negative control (control RIPs). Genome-wide transcript expression in wild-type embryos were also assessed and used as reference.
Project description:Homozygous masterblind (mbl-/-) zebrafish exhibit reduced or absent eyes and telecephalon, and the expansion of the diencephalic fates to the front of the brain. A missense mutation in the GSK3-binding domain of zebrafish axin1, a scaffolding protein in the Wnt signaling pathway, results in the mbl phenotype. In an effort to identify and study the genes affected by Wnt signaling, we used a 14,000-oligonucleotide-gene microarray to determine differentially expressed genes in mbl/axin1 (-/-) and wild type control zebrafish embryos and larvae. Keywords: zebrafish, Danio rerio, wild-type mbl, axin1, development
Project description:To identify SmaugM-bM-^@M-^Ys target mRNAs on a genome-wide scale we used ribonucleoprotein (RNP) co-immunoprecipitation followed by microarray analysis of the co-purifying mRNAs (RIP-Chip). Extracts, prepared from wild-type embryos collected 0-3 hours post-egglaying, were immunoprecipitated with an anti-Smaug antibody (Smaug RIPs) while control immunoprecipitations using non-immune serum served as a negative control (control RIPs). Genome-wide transcript expression in wild-type embryos were also assessed and used as reference. There are 11 array experiments presented here: 1. gene expression profiling of total mRNA sample extracted from wild-type 0-3 hour embryos (3 technical replicates performed using a pooled input reference sample); 2. RNA co-immunoprecipitations of endogenous Smaug (Smaug RIPs) from wild-type 0-3 hour embryos (3 biological replicates and 1 technical replicates); 3. control RNA co-immunoprecipitations (control RIPs) from wild-type 0-3 hour embryos (3 biological replicates and 1 technical replicates).
Project description:To identify targets of miR-203a we coupled RNA-seq and AGO2-RNA immunoprecipitation (RIP-seq) in MCF10A cells expressing a control pre-miR-203a or control, empty vector
Project description:Ozone is a common pollutant and a potent oxidant in industrialized nations. The mechanisms of ozone-induced lung injury and differential susceptibility are not fully understood. Ozone-induced lung inflammation is mediated, in part, by the innate immune system. We hypothesized that mannose binding lectin (MBL), which has a central role in the activation of the complement pathway of innate immunity, is a necessary component of the pro-inflammatory events caused by ozone-mediated activation of the innate immune system. Wild-type (Mbl+/+) and MBL deficient (Mbl-/-) mice were exposed to ozone (0.3 ppm) for 24, 48, and 72 hours, and bronchoalveolar lavage fluid (BALF) was examined for inflammatory markers. Compared to Mbl+/+ mice, significantly greater mean BALF eosinophils, neutrophils and neutrophil attractants CXCL2 (MIP-2) and CXCL5 (LIX) were found in Mbl-/- mice exposed to ozone. Using genome-wide mRNA microarray analyses, we identified significant differences in expression response profiles and networks at baseline (e.g. NRF2 mediated oxidative stress response) and after exposure (e.g. humoral immune response) between Mbl+/+ and Mbl-/- mice. The microarray data were further analyzed using a pattern recognition analysis for Extracting Patterns and Identifying co-expressed Genes (EPIG), and discovered several informative differential response patterns and subsequent gene sets, including antimicrobial response and inflammatory response. These novel findings demonstrate that targeted deletion of Mbl caused differential expression of inflammation-related gene sets basally and after exposure to ozone, and significantly reduced pulmonary inflammation thus indicating an important innate immunomodulatory role of the gene in this model.
Project description:IMP1 is a RNA binding protein shown to regulate translation, localization, or stability of their target RNAs. However, whether IMP1 can widely bind to lncRNAs and regulate their expression in Breast cancer cells remains to be confirmed. In this study, we used RNA binding protein immunoprecipitation-high throughput sequencing (RIP-Seq) technique to identify LncRNAs which bind to IMP1 protein in breast cancer cell line T47D cells.