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    Measurement of phosphoinositides in the zebrafish Danio rerio.

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    Authors
    Jones, David R
    Ramirez, I
    Lowe, M
    Divecha, Nullin
    Affiliation
    Cancer Research UK Inositide Laboratory, Paterson Institute for Cancer Research, University of Manchester, Manchester, UK.
    Issue Date
    2013-05-09
    
    Metadata
    Show full item record
    Abstract
    Phosphoinositides represent a minor fraction of the total glycerolipids in cells. Despite the fact that phosphoinositides are present in small quantities, they have crucial roles during cell signaling and in regulating numerous intracellular processes. Measuring changes in the levels of different phosphoinositides in animals is difficult, but it is essential in order to define the important functions of specific members of the phosphoinositide family. Here we detail procedures for measuring phosphoinositides in 2-days-postfertilization (2-d.p.f.) embryos in zebrafish (Danio rerio). Both in vivo radiolabeling (using [(32)P]orthophosphate) followed by thin-layer or high-performance liquid chromatography (TLC or HPLC) analysis and specific in vitro phosphorylation assays (using [(32)P]γATP) permit the quantitative measurement of phosphoinositides. Normalization of both measurements can be achieved by the determination of total lipid phosphate in embryos. All the techniques described are relatively inexpensive and accessible to most laboratories with an interest in studying the effect of gene manipulation on phosphoinositide metabolism in zebrafish. All the procedures described herein will take up to 10 working days.
    Citation
    Measurement of phosphoinositides in the zebrafish Danio rerio. 2013, 8 (6):1058-72 Nat Protoc
    Journal
    Nature Protocols
    URI
    http://hdl.handle.net/10541/294008
    DOI
    10.1038/nprot.2013.040
    PubMed ID
    23660755
    Type
    Article
    Language
    en
    ISSN
    1750-2799
    ae974a485f413a2113503eed53cd6c53
    10.1038/nprot.2013.040
    Scopus Count
    Collections
    All Paterson Institute for Cancer Research

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