Plasmid mediated mutagenesis of a cellular gene in transfected eukaryotic cells

Curtis R. Brandt; Franco M. Buonaguro; James K. Mcdougall; Denise A. Galloway

doi:10.1093/nar/15.2.561

Plasmid mediated mutagenesis of a cellular gene in transfected eukaryotic cells

Curtis R. Brandt, Franco M. Buonaguro, James K. Mcdougall, Denise A. Galloway

Istituto Nazionale Tumori Fondazione G. Pascale

Research output: Contribution to journal › Article › peer-review

Abstract

NIH3T3 cells are widely used in transformation assays and readily take up transfected DNA. A system has been devised using NIH3T3 cells to measure the mutagenic effect of transfected DNA on recipient cell genes. NIH3T3 cells can be mutated to 6-thioguanine resistance at a frequency which suggests that at least a portion of the cells have only one functional copy of the HGPRT gene. They have a low spontaneous background mutation frequency (≈1×10^-7). Transfection of three different plasmids into NIH3T3 cells induced 6-thioguanine resistant mutants at frequencies ranging from 3 to 11 fold above background. The mutant phenotype is stable and reversion frequencies of several mutants are ≤1×10^-7. Southern blot analysis of the HGPRT gene in several mutants showed that 4 of 26 mutants (15.4%) had detectable alterations in the structure of the HGPRT gene. Interestingly 3 of the 4 mutants showing rearrangements were obtained by transfection of the HSV-2 morphological transforming region.

Original language	English
Pages (from-to)	561-573
Number of pages	13
Journal	Nucleic Acids Research
Volume	15
Issue number	2
DOIs	https://doi.org/10.1093/nar/15.2.561
Publication status	Published - Jan 26 1987

ASJC Scopus subject areas

Statistics, Probability and Uncertainty
Applied Mathematics
Health, Toxicology and Mutagenesis
Toxicology
Genetics(clinical)
Genetics

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Plasmid mediated mutagenesis of a cellular gene in transfected eukaryotic cells

Abstract

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