Project description:This project aimed at identifying developmental stage specific transcript profiles for catecholaminergic neurons in embryos and early larvae of zebrafish (Danio rerio). Catecholaminergic neurons were labeled using transgenic zebrafish strains to drive expression of GFP. At stages 24, 36, 72 and 96 hrs post fertilization, embryos were dissociated and GFP expressing cells sorted by FACS. Isolated RNAs were processed using either polyA selection and libray generation or NanoCAGE. This is the first effort to determine stage specific mRNA profiles of catecholaminergic neurons in zebrafish.
Project description:All vertebrates have multiple genes encoding for different CASQ isoforms. Increasing interest has been focused on mammalian and human CASQ genes since mutations of both cardiac (CASQ2) and skeletal (CASQ1) isoforms cause different, and sometime severe, human pathologies Danio rerio (zebrafish) is a powerful model for studying function and mutations of human proteins. In this work expression, biochemical properties and cellular and sub-cellular localization of Danio rerio native CASQ isoforms are investigated. By quantitative PCR three mRNAs were detected in skeletal muscle and one mRNA in heart. Three zebrafish CASQs were identified by mass spectrometry and they share properties with mammalian skeletal and cardiac CASQs. Skeletal calsequestrins were found primarily, but not exclusively, at the sarcomere Z-line level where Terminal Cisternae of Sarcoplasmic reticulum are located.
Project description:In triplicate for each condition, 12 WT and acbd6 F0 crispant Danio rerio (zebrafish) embryos were incubated with 20 μM YnMyr for 24 h, either between 48-72 hpf or 96-120 hpf. After labelling, zebrafish were washed twice with fresh egg water, deyolked, flash frozen in liquid nitrogen and stored at -80°C until further analysis.
Project description:In triplicate for each condition, 12 WT and acbd6 F0 crispant Danio rerio (zebrafish) embryos were incubated with 20 μM YnMyr for 24 h, either between 48-72 hpf or 96-120 hpf. After labelling, zebrafish were washed twice with fresh egg water, deyolked, flash frozen in liquid nitrogen and stored at -80°C until further analysis.
Project description:Chlorothalonil (2,4,5,6-tetrachloroisophthalonitrile) is a broad spectrum fungicide used extensively in agricultural crops . The aim of this study is to analyse the effects of Chorothalonil on the gene expression profiles in zebrafish (Danio rerio), exposed to two concentrations of the fungicide in the water. Nominal concentrations were 1) Low 0.007mg/l (environmentally relevent) and 2) High 0.035mg/ml . A commercial third generation microarray for Danio rerio (Agielnt V3, 4x44k) was used to identify patterns of gene expression in male livers during a 96h toxicological assay.
Project description:Histidine phosphorylation is a reversible post-translational modification that is known to regulate signal transduction in prokaryotes. In an effort to help elucidate the heretofore hidden vertebrate phosphoproteome, this report presents a global phosphorylation analysis of Danio rerio (zebrafish) larvae. Phosphopeptide enrichment was performed using a TiO2 affinity technique. A total of 68 unique phosphohistidine sites were detected on 63 proteins among 1076 unique phosphosites on 708 proteins. This report provides the first phosphohistidine dataset obtained from zebrafish.
Project description:This project aimed at identifying developmental stage specific transcript profiles for catecholaminergic neurons in embryos and early larvae of zebrafish (Danio rerio). Catecholaminergic neurons were labeled using transgenic zebrafish strains to drive expression of GFP. At stages 24, 36, 72 and 96 hrs post fertilization, embryos were dissociated and GFP expressing cells sorted by FACS. Isolated RNAs were processed using either polyA selection and libray generation or NanoCAGE. This is the first effort to determine stage specific mRNA profiles of catecholaminergic neurons in zebrafish. Catecholaminergic neurons were labeled by four different strategies: (1) 24 hrs old embryos: we used the ETvmat2:GFP transgenic line (Wen et al. 2007). Visualization of monoaminergic neurons and neurotoxicity of MPTP in live transgenic zebrafish. Dev Biol. 2008 Vol 314 p84-92) which at this early stage labels catecholaminergic neurons in posterior tuberculum and locus coeruleus; (2) 24 hrs old embryos: we used Tg(otpb.A:egfp)zc48 transgenic line (Fujimoto et al. Identification of a dopaminergic enhancer indicates complexity in vertebrate dopamine neuron phenotype specification. Dev Biol 2011, Vol 352, p393–404) which at this stage label ventral diencephalic dopaminergic neurons and some preoptic neurons. (3) For 72 and 96 hrs old zebrafish larvae we used a th:GFP BAC transgenic lines that labels catecholaminergic neurons (Tay et al., Comprehensive catecholaminergic projectome analysis reveals single-neuron integration of zebrafish ascending and descending dopaminergic systems. Nat Comms 2011 Vol 2, 171; also: T. Leng and W. Driever, unpublished). (4) for the 36 and 48 hrs old zebrafish larvae we used a th:Gal4VP16 driver and UAS:EGFP responder transgenic line system to label catecholaminergic cells (Fernandes et al., Deep brain photoreceptors control light-seeking behavior in zebrafish larvae. Curr Biol. 2012 Vol 22 DOI 10.1016/j.cub.2012.08.016). We used the different transgenic lines, because lines (3) and (4) do not efficiently label catecholaminergic neurons at early stages, while lines (1) and (2) also have GFP expression in several other non-catecholaminergic populations at later stages of development. Embryos were dissociated and catecholaminergic neurons were FACS sorted from GFP-tagged zebrafish (Manoli and Driever, 2012, Cold Spring Harbor Protoc. DOI 10.1101/pdb.prot069633). RNA was either processed for NanoCAGE, or mRNA was isolated and amplified. cDNA was sequenced by Illumina technique. This data submission is a series of data files consisting of three independent experiments with diffrent RNA-Seq depth: Samples 1-4 (NanoCage): Samples 5-8 (RNA-Seq high read numbers), and SAmples 9-12 (RNA-Seq low read numbers).