Functional screen of zebrafish deubiquitylating enzymes by morpholino knockdown and in situ hybridization

Ka Fai William Tse, Yun Jin Jiang

Research output: Chapter in Book/Report/Conference proceedingChapter

9 Citations (Scopus)

Abstract

In order to unfold the function of genes, solely performing mRNA over-expression is not enough nowadays. Traditional protein expression experiments, such as Western blotting and immunohistochemical staining, could only provide researchers the changes of expression levels and/or location of their targets. To make a more strong and convincing statement about gene function, it is necessary to perform both "gain-of-function" and "loss-of-function" studies. Both assays can be performed easily by transfecting DNA plasmid and siRNA in cell culture system; while in zebrafish, mRNA and morpholino (MO) microinjection can serve similar purposes. It is common for the zebrafish community to carry out microinjection experiments to explore a gene function. Instead of making a single knockdown/over-expression of a gene, we foresee that more and more large-scale screens on certain protein families will be performed in the future. Here, based on our previous experience in zebrafish "loss-of-function" screening on deubiquitylating enzymes, we describe a general work flow, from morpholino designation, in situ hybridization, to data analysis, as a reference for researchers who may be interested in a similar screen.

Original languageEnglish
Title of host publicationFunctional Genomics
Subtitle of host publicationMethods and Protocols
EditorsMichael Kaufmann, Claudia Klinger
Pages321-331
Number of pages11
DOIs
Publication statusPublished - Jan 2 2012
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume815
ISSN (Print)1064-3745

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics

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