# MECHANISM OF MUTATION INDUCED IN MAMMALIAN GENE

> **NIH NIH R01** · COLUMBIA UNIV NEW YORK MORNINGSIDE · 1987 · $149,647

## Abstract

The proposal describes approaches to define the spectrum of lesions induced
by mutagens in a mammalian gene at the DNA sequence level.  Mutations
occurring in an endogenous eukaryotic gene differ qualitatively.  We will
examine carcinogen-induced mutations in the Chinese hamster ovary locus
encoding dihydrofolate reductase (DHFR).  In this system, we can
characterize point mutations, as well as chromosomal events (translocation,
deletion, sequence duplication, and insertion).  DHFR-deficient cells have
been isolated that are either hemozygous (dhfr+/d) or deleted for the locus
(dhfr(d/d)).  DHFR- mutants are triple auxotrophs for glycine, thymidine,
and hypoxanthine.  Mutants will be selected starting with the hemizygote.
The gene structure has been well characterized.  Southern blot analysis of
the induced dhfr-/d genomic DNAs will show changes arising from chromosomal
events.  Using dhfr-specific cosmid vectors, we will clone the locus from
DHFR-cells carrying point mutations.  We will map these mutations by marker
rescue relying on in vivo or in vitro recombination of the dhfr- cosmid
with dhfr+ fragments to correct the lesion.  After transfection of the
mutant and wild-type DNAs into dhfr(d/d) cells, prototrophic clones will be
selected.  Upon further mapping, we will sequence regions containing the
mutations.  Another approach for studying the nature of induced mutations
in animal cells similar to the Ames assay is described.  We will construct
by site-directed mutagenesis 4 different minigenes (full length cDNA fused
to the dhfr promoter).  The changes to be introduced will create
transition, transversion, +1, and -1 frameshift mutations.  We will
transfect the minigenes into dhfr(d/d) cells selecting for another marker
(XGPT).  These clones will then be mutagenized to induce reversion to
DHFR+.  If we can induce reversion, this system will be useful for
screening the mutational potential of drugs and chemicals.  Also, other
parameters influencing the spontaneous and induced frequencies of reversion
such as chromosomal location and DNA excision repair will be investigated.

## Key facts

- **NIH application ID:** 3178656
- **Project number:** 5R01CA039547-03
- **Recipient organization:** COLUMBIA UNIV NEW YORK MORNINGSIDE
- **SAM.gov UEI:** F4N1QNPB95M4
- **Principal Investigator:** DEZIDER GRUNBERGER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1987
- **Award amount:** $149,647
- **Award type:** 5
- **Project period:** 1985-04-01 → 1988-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3178656

## Citation

> US National Institutes of Health, RePORTER application 3178656, MECHANISM OF MUTATION INDUCED IN MAMMALIAN GENE (5R01CA039547-03). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3178656. Licensed CC0.

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