Selection of targeted mutants from a library of randomly mutagenized ES cells

Methods Mol Biol. 2011;693:283-94. doi: 10.1007/978-1-60761-974-1_17.

Abstract

A method for random relatively unbiased mutagenesis of ES cells with a mutagenic retroviral vector is described. An orderly assembly of mutant ES cells in multi-well plates is generated. 3D pooling of the wells of the assembly allows quick PCR search for insertions in genes of interest. Mutant ES cell clones are then isolated from the positive wells and used to produce mutant animals using conventional techniques.

MeSH terms

  • Animals
  • Embryonic Stem Cells / metabolism*
  • Gene Targeting / methods*
  • Genetic Vectors / genetics
  • Mice
  • Mice, Knockout
  • Mutagenesis / genetics
  • Polymerase Chain Reaction
  • Retroviridae / genetics