System issue: Spontaneous and ethylnitrosourea-induced mutation fixation and molecular spectra at the lacl transgene in the Big Blue® Rat-2 embryo cell line
✍ Scribed by D. M. Zimmer; X. B. Zhang; P. R. Harbach; J. K. Mayo; C. S. Aaron
- Publisher
- John Wiley and Sons
- Year
- 1996
- Tongue
- English
- Weight
- 672 KB
- Volume
- 28
- Category
- Article
- ISSN
- 0893-6692
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✦ Synopsis
Big BlueTM Rat-2 cells were evaluated for mutagenesis and mutational spectra (spontaneous and ethylnitrosourea [ENUI-induced). Survival, mutant frequency, population doubling time, and kinetics of mutant increase (to 120 hr) were determined. Exposures were 100, 200, 400, 600, and 1,000 Fg ENU/ml. The spontaneous mutant frequency was similar to that previously reported in vivo, i.e., 5 x 1 O-'. Dose-related increases in mutant frequency were observed following ENU treatment. Kinetics (time course) of mutant frequency increase, population doubling, and mutational spectra were investigated following treatment at 1,000 k g ENU/ml. Among 39 spontaneous mutants, 26 independent mutations were found as follows: nine (34.6%) G:C + A:T transitions (five at CpG sites), six (23%) G:C --$ T A transversions, three (1 1.5%) G:C + C:G transversions (two at CpG sites), iwo (7.7%) frameshihs, five (1 9%) deletions or insertions, and one (3.8%) complex (deletion + insertion) mutation. Among 46 ENU-induced mutants, 37 independent mutations (all base substitutions) were found as follows: 15 (40.5%) G:C + A:T transitions (four at CpG sites), five (1 3.5%) A:T + G:C transitions, four (1 0.8%) G:C + T:A transversions, 1 1 (30%) A:T + T:A transversions, and two (5.4%) A:T + C:G transversions. Nearly 50% of the base substitutions in the ENU-treated cells were at A:T base pairs, in contrast to the spontaneous mutants where none was found. Both the spontaneous and the ENUinduced mutational spectra were similar to that reported in vivo and for other cells. An important aspect of the experiment is that all mutations sequenced following ENU treatment (1,000 Fg/ml) occurred under conditions which our experiments show corresponded to very little mitotic activity.