[Experimental study of use of shuttle vector to develop a system of detecting mutagenesis in mammalian cells].
The paper presents a series of experimental studies of the use of shuttle vector to develop a short-time mutagenesis system of detecting gene mutation in the mammalian cell at DNA level. The system is based on the finding that in the EB virus the shuttle vector pMCi5 carries LacI gene. The LacI is used as a target of induced mutagenesis in mouse cells. Use pMCi5 plasmid to transform the 3T3 mouse cell, expose it to a chemical mutagen, ethyl methanesulfonate (EMS); transfer the treated plasmid DNA to E. coli strain MC1061F' 150Kan; and then plate the transformation on a special culture medium containing X-gal for rapid detection and analysis of LacI mutation.


RESULTS
The spontaneous mutant frequency of the system is less than 1.21 x 10(-4). After the treatment with 300 micrograms/ml EMS, the induced mutant frequency increases to 3.8 x 10(-3). No gross alteration has been discovered in seven LacI mutants at DNA level. No point mutation has occurred in the EcoRI site by analysis with restriction enzyme EcoRI.
Lin
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Fu
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Zhai
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D016475:3T3 Cells / Q000187:drug effects; D000818:Animals; D002478:Cells, Cultured; D005020:Ethyl Methanesulfonate / Q000633:toxicity*; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics; D051379:Mice; D009152:Mutagenicity Tests / Q000379:methods; D009154:Mutation / Q000187:drug effects*; D010957:Plasmids / Q000235:genetics*; D014170:Transformation, Genetic
D005020:Ethyl Methanesulfonate