Alterations of Gene Structure in Ethyl Methane Sulfonate-Induced Mutants of Mammalian Cells

Meuth, Mark; Arrand, Janet E.

doi:10.1128/mcb.2.11.1459

Cited by 44 publications

(22 citation statements)

References 20 publications

(19 reference statements)

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“…Breimer et al (1986), Grosovsky et al (1986fFuscoe et al (1983), Vrieling et al (1985, Breimer et al (1986), Fuscoe et al (1986), 3Skulimowski et al (1986), Liber et al (1987hYandell et al (1986hYandell et al ( , 1987 aprt locus The endogenous genetic locus coding for the non-essential enzyme adenine phosphoribosyl transferase (aprt) in cultured Chinese hamster ovary cells is particularly attractive for analysis of mutational events because it offers a much greater resolution than any other mammalian locus used so far (Meuth & Arrand, 1982;Nalbantoglu et al, 1983). The locus is small (3.9 kb) and contains a large number of restriction endonuclease recognition sites closely spaced, allowing detection and mapping of alterations as small as 25bp.…”

mentioning

confidence: 99%

Ionizing radiation-induced mutagenesis

Breimer

1988

Br J Cancer

150

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confidence: 99%

Ionizing radiation-induced mutagenesis

Breimer

1988

Br J Cancer

150

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“…Other endpoints have, by necessity, been used to construct a preliminary view of mutation in mammalian cells. These include comparative mutation induction at several selectable loci (4,5), twodimensional gel electrophoresis analysis of mutant proteins (6), and Southern blot analysis of restriction site polymorphisms (7)(8)(9)(10). Recently, recombinant shuttle vectors, which can be propagated in both prokaryotic and eukaryotic cells, have been developed for the analysis of mutant DNA sequences in mammalian cells (reviewed in ref.…”

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confidence: 99%

Spectrum of spontaneous mutation at the APRT locus of Chinese hamster ovary cells: an analysis at the DNA sequence level.

Jong

Grosovsky

Glickman

1988

Proc. Natl. Acad. Sci. U.S.A.

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The spectrum of spontaneous mutation of an endogenous mammalian cell gene has been determined at the DNA sequence level. Thirty independent spontaneous APRTmutations were cloned and subsequently completely sequenced. Twenty-seven contained single base substitutions. Of these, 22 were G-C to A-T transitions, suggesting a major role for the deamination of cytosine in spontaneous mutagenesis of Chinese hamster ovary cells. The remaining mutants included a tandem double substitution, a -1 frameshift, and a 17-base-pair deletion flanked by a 2-base-pair direct repeat. Many of the independently recovered mutants were clustered at sites of multiple occurrence (hot spots). One site accounted for >25% of all independently recovered events. Mutations were generally located within the coding sequence, although two mutations occurred within the consensus sequence for a 3' splice site.A fundamental understanding of mutagenesis in mammalian cells ultimately requires specific knowledge of the DNA sequence alterations involved. A complete analysis would describe the spectrum of mutational alterations for a random collection of independent mutants at a given locus. Such analyses have recently been reported in bacteria (1-3), but logistical difficulties have prevented such an analysis of an endogenous mammalian cell gene. Other endpoints have, by necessity, been used to construct a preliminary view of mutation in mammalian cells. These include comparative mutation induction at several selectable loci (4, 5), twodimensional gel electrophoresis analysis of mutant proteins (6), and Southern blot analysis of restriction site polymorphisms (7-10). Recently, recombinant shuttle vectors, which can be propagated in both prokaryotic and eukaryotic cells, have been developed for the analysis of mutant DNA sequences in mammalian cells (reviewed in ref. 11). Although the use of shuttle vectors facilitates the recovery and sequence analysis of mutations, this experimental approach has several drawbacks, often including an extraordinarily high mutation frequency and a high percentage of deletions and rearrangements among the recovered plasmids (12-16). This is true even under conditions in which the shuttle vector has been incorporated into the chromosome, although some inserts appear to approach the stability expected for natural genes (17, 18). These technical problems and the artificial nature of such constructs make it questionable whether these systems accurately represent mutagenesis at endogenous cellular genes.Our efforts have therefore concentrated on the development of a practical approach to the cloning and sequencing of mutant alleles of an endogenous gene. Our system involves the analysis of mutants at the adenine phosphoribosyltransferase (APRT) locus of Chinese hamster ovary (CHO) cells. This locus is well suited for mutational studies. It encodes an enzyme in the nucleotide salvage pathway and is thus nonessential for cell survival in ordinary growth medium. Consequently, all classes of mutational events can in principl...

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“…Ethyl methane sulphonate is a monofunctional alkylating agent and is considered as a potent mutagen and carcinogen (Meuth and Arrand 1982). In plants, EMS chemically alters DNA through the modification of oxygen present in nucleotide bases as well as in DNA phosphate groups (Sega 1984).…”

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Nuclear Blebs Assay for Genome Instability in <i>Phaseolus vulgaris</i> L.

Kumar

Chaudhary

2015

CYTOLOGIA

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Summary Micronuclei are extra nuclear structures, formed by the exclusion of chromosomes during cell division. The genotoxicity of a compound and its impact on the genome can be assessed via micronuclei formation. The present study has been undertaken to evaluate the mutagenic effect of the alkylating agent EMS on the meiotic behavior of pollen mother cells of Phaseolus vulgaris L. The cytological analysis revealed the induction of micronuclei at all the three administered concentrations of EMS, viz. 0.1, 0.3 and 0.5%. Besides micronuclei formation, other meiotic abnormalities associated with chromosomal segregation, such as stickiness, laggard, bridge, multivalent, etc., were also reported at metaphase I/II and anaphase I/II. The micronuclei formed would have two fates either it remains until the tetrad stage or is expelled out in the form of microcytes which will give rise to sterile pollen grains affecting the pollen fertility. The present study also documents the first instance of nuclear budding in Phaseolus vulgaris L. It is contemplated that these nuclear buds will later on convert into micronuclei. The micronuclei formation results in partial elimination of the genome, which could be efficiently utilized in breeding programmes for the production of addition and substitution lines. Similarly, haploid lines could also be produced by the complete loss of genome. It is therefore essential to understand the biological aspects of micronuclei formation, its consequences on the cell carrying it and other factors related to it so that in the future, it could be used as a marker of genetic damage and also be utilized in breeding programmes.

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Alterations of Gene Structure in Ethyl Methane Sulfonate-Induced Mutants of Mammalian Cells

Cited by 44 publications

References 20 publications

Ionizing radiation-induced mutagenesis

Ionizing radiation-induced mutagenesis

Spectrum of spontaneous mutation at the APRT locus of Chinese hamster ovary cells: an analysis at the DNA sequence level.

Nuclear Blebs Assay for Genome Instability in <i>Phaseolus vulgaris</i> L.

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