Visualizing loss of heterozygosity in living mouse cells and tissues

Jared M. Fischer, James R. Stringer

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

Loss of heterozygosity (LOH) in somatic cells can contribute to the genesis of cancer, but little is known about the frequency with which LOH occurs in normal cells of the body. To detect LOH in situ, we studied mouse shYFP embryonic stem (ES) cells and cells of the intestinal epithelia derived from these ES cells. shYFP ES cells are heterozygous at the ROSA26 locus. One copy of the locus carries a gene encoding a yellow fluorescent protein (YFP), while the other copy harbors an shRNA gene that produces a short hairpin RNA (shRNA) molecule that causes degradation of YFP mRNA. Nearly all cells in shYFP populations were faintly fluorescent, but brightly fluorescent cells arose at a rate of approximately 10-5 bright cells/generation. Bright cells lacked the gene encoding the shRNA and contained two copies of the YFP gene. Comparison of these results to previous data on LOH in ES cells that lacked interfering shRNA showed that LOH in shYFP cells was not influenced by the presence of the shRNA. Bright cells were also seen in intestinal villi of chimeric mice made by injecting blastocysts with shYFP cells. These data demonstrate that this approach can detect LOH and suggest that it will allow detection of LOH in a broad array of tissues and cell types in transgenic mice made from shYFP cells.

Original languageEnglish (US)
Pages (from-to)1-8
Number of pages8
JournalMutation Research - Fundamental and Molecular Mechanisms of Mutagenesis
Volume645
Issue number1-2
DOIs
StatePublished - Oct 14 2008

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Keywords

  • Embryonic stem cells
  • Intestinal epithelia
  • Loss of heterozygosity
  • Mutation
  • Short hairpin RNA
  • Yellow fluorescent protein

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Health, Toxicology and Mutagenesis

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