Detection of amplified sequences in mammalian DNA by in-gel renaturation and SINE hybridization

Manabu Fukumoto, Igor B. Roninson

Research output: Contribution to journalArticlepeer-review

5 Citations (Scopus)

Abstract

The presence of amplified sequences in mammalian DNA can be determined by in-gel renaturation of labeled restriction digests of total genomic DNA, provided that such sequences comprise at least 20-30 copies per haploid human or hamster genome or 40-50 copies per mouse genome. To detect amplified DNA in mouse cells at a lower level of amplification, a new procedure has been developed. This procedure combines in-gel DNA renaturation with Southern hybridization using a cloned probe containing a short interspersed repeated element (SINE). Using mouse cell lines containing amplified dihydrofolate reductase (DHFR)or c-Ki-ras genes as a model system, and the cloned B2 repeated sequence as a SINE probe, we have shown that this technique can detect gene amplification at a level as low as 10-15 copies of amplified DNA per haploid mouse genome. In-gel renaturation-SINE hybridization was used to assay for DNA amplification in different tissues and in different mouse strains. No tissue-specific DNA amplification has been detected, but comparison of B2-containing repeated fragments between three inbred mouse lines revealed strain-specific polymorphism.

Original languageEnglish
Pages (from-to)611-623
Number of pages13
JournalSomatic Cell and Molecular Genetics
Volume12
Issue number6
DOIs
Publication statusPublished - 1986 Nov

ASJC Scopus subject areas

  • Genetics
  • Cell Biology

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