Direct removal in the mouse of a floxed neo gene from a three-loxP conditional knockout allele by two novel approaches

Genesis. 2001 May;30(1):1-6. doi: 10.1002/gene.1025.

Abstract

The presence in an intron of the ploxP-neo-loxP cassette often results in severe interference with gene expression. Consequently, many investigators selectively remove the ploxP-neo-loxP cassette by transient expression of Cre in ES cells. Although effective, the added manipulation of the ES cells may reduce the likelihood that a clone will be able to transmit via the germline. Therefore, we developed two novel approaches that remove the ploxP-neo-loxP by Cre-mediated recombination in mouse. First, the ploxP-neo-loxP-containing mice were crossed with EIIa-Cre transgenic mice. Second, a Cre-expression plasmid was injected into pronuclei of fertilized eggs bearing the ploxP-neo-loxP allele. Both approaches produced mosaic mice with partial and complete excision. These mosaic mice were then mated, and the neo-less conditional knockout allele was found in the offspring after screening only a few litters. These procedures provide options for removing neo directly in the mouse in addition to the commonly used approach that deletes neo in ES cells.

MeSH terms

  • Alleles
  • Animals
  • Cell Nucleus / metabolism
  • Crosses, Genetic
  • Embryo, Mammalian / cytology
  • Embryo, Mammalian / metabolism
  • Exons
  • Female
  • Gene Transfer Techniques*
  • Genes, BRCA1 / genetics
  • Genetic Vectors*
  • Heterozygote
  • Integrases / genetics*
  • Introns
  • Male
  • Mice
  • Mice, Knockout*
  • Mice, Transgenic
  • Models, Genetic
  • Phenotype
  • Plasmids / metabolism
  • Recombination, Genetic
  • Reverse Transcriptase Polymerase Chain Reaction
  • Stem Cells / cytology
  • Viral Proteins / genetics*

Substances

  • Viral Proteins
  • Cre recombinase
  • Integrases