Dataset Information

Heritable and lineage-specific gene knockdown in zebrafish embryo.

ABSTRACT: BACKGROUND: Reduced expression of developmentally important genes and tumor suppressors due to haploinsufficiency or epigenetic suppression has been shown to contribute to the pathogenesis of various malignancies. However, methodology that allows spatio-temporally knockdown of gene expression in various model organisms such as zebrafish has not been well established, which largely limits the potential of zebrafish as a vertebrate model of human malignant disorders. PRINCIPAL FINDING: Here, we report that multiple copies of small hairpin RNA (shRNA) are expressed from a single transcript that mimics the natural microRNA-30e precursor (mir-shRNA). The mir-shRNA, when microinjected into zebrafish embryos, induced an efficient knockdown of two developmentally essential genes chordin and alpha-catenin in a dose-controllable fashion. Furthermore, we designed a novel cassette vector to simultaneously express an intronic mir-shRNA and a chimeric red fluorescent protein driven by lineage-specific promoter, which efficiently reduced the expression of a chromosomally integrated reporter gene and an endogenously expressed gata-1 gene in the developing erythroid progenitors and hemangioblasts, respectively. SIGNIFICANCE: This methodology provides an invaluable tool to knockdown developmental important genes in a tissue-specific manner or to establish animal models, in which the gene dosage is critically important in the pathogenesis of human disorders. The strategy should be also applicable to other model organisms.

SUBMITTER: Dong M

PROVIDER: S-EPMC2702085 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Heritable and lineage-specific gene knockdown in zebrafish embryo.

Dong Mei M Fu Yan-Fang YF Du Ting-Ting TT Jing Chang-Bin CB Fu Chun-Tang CT Chen Yi Y Jin Yi Y Deng Min M Liu Ting Xi TX

PloS one 20090703 7

<h4>Background</h4>Reduced expression of developmentally important genes and tumor suppressors due to haploinsufficiency or epigenetic suppression has been shown to contribute to the pathogenesis of various malignancies. However, methodology that allows spatio-temporally knockdown of gene expression in various model organisms such as zebrafish has not been well established, which largely limits the potential of zebrafish as a vertebrate model of human malignant disorders.<h4>Principal finding</h ...[more]

PMID: 19582161

Similar Datasets

Project description:Sensitivity and throughput of transcriptomics and proteomomics technologies have advanced tremendously in recent years. With the use of deep sequencing of RNA samples (RNAseq) and mass spectrometry technology for protein identification and quantitation, it is now feasible to compare gene and protein expression on a massive scale and on any organism for which genomic data is available. Although these technologies are currently applied to many research questions in various model systems ranging from cell cultures to the entire organism level, there are few comparative studies of these technologies in the same system, let alone on the same samples. Here we present a comparison between gene and protein expression in zebrafish embryos, which is an upcoming model in disease studies. We compared Agilent custom made expression microarrays with Illumina deep sequencing for RNA analysis, showing as expected a high degree of correlation of expression of a common set of 15,927 genes. Gene expression was also found to correlate with the abundance of 1,017 quantified proteins, with several categories of genes as exceptions. These exceptions include ribosomal proteins, histones and vitellogenins, for which biological and technical explanations are discussed. By comparing state of the art transcriptomic and proteomic technologies on samples derived from the same group of organisms we have for the first time benchmarked the differences in these technologies with regard to sensitivity and bias towards detection of particular gene categories. Our datasets submitted to public repositories are a good starting point for researchers interested in disease progression in zebrafish at a stage of development highly suited for high throughput screening technologies. The transcriptomics data is deposited in NCBI GEO. Proteomics informatics: Spectra were identified by Mascot and validated by PeptideProphet and ProteinProphet in the Trans-Proteomic Pipeline (TPP). The main search parameters are: full tryptic specificity, precursor mass searched within -0.5 to +2.5 Da, product ions within [-0.5,0.5] Da and allowing for one missed cleavage. Carbamidomethylation was assumed as the only and fixed PTM.

Dataset Information

Heritable and lineage-specific gene knockdown in zebrafish embryo.

Publications

Heritable and lineage-specific gene knockdown in zebrafish embryo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets