ZFIN ID: ZDB-PUB-090422-22
Production of pseudotyped retrovirus and the generation of proviral transgenic zebrafish
Jao, L.E., and Burgess, S.M.
Date: 2009
Source: Methods in molecular biology (Clifton, N.J.)   546: 13-30 (Chapter)
Registered Authors: Burgess, Shawn
Keywords: Zebrafish, Retrovirus, Pseudotyped Moloney murine leukemia virus, Insertional mutagenesis, Injection
MeSH Terms:
  • Animals
  • Animals, Genetically Modified*
  • Cell Line
  • Chimerism
  • DNA/metabolism
  • Female
  • Gene Transfer Techniques
  • Genome/genetics
  • Humans
  • Male
  • Membrane Glycoproteins/biosynthesis
  • Membrane Glycoproteins/genetics
  • Moloney murine leukemia virus/genetics
  • Moloney murine leukemia virus/growth & development*
  • Moloney murine leukemia virus/metabolism*
  • Mutagenesis, Insertional/methods*
  • Reverse Transcriptase Polymerase Chain Reaction/methods
  • Viral Envelope Proteins/biosynthesis
  • Viral Envelope Proteins/genetics
  • Zebrafish/embryology
  • Zebrafish/genetics*
  • Zebrafish/virology*
PubMed: 19378095 Full text @ Meth. Mol. Biol.
This chapter describes a method for generation of the high-titer pseudotyped Moloney murine leukemia virus (MLV) that efficiently infects zebrafish embryos (i.e., more than 25 retroviral copies per cell). Injection techniques are also described for production of the retrovirus-infected mosaic "founder" fish. We describe a quantitative PCR (qPCR)-based assay as a quick way to assess the infectivity after each round of viral production and injection. Most of the required equipment is commercially available and commonly present in most research laboratories.