Related Experiment Video
Updated: Jul 5, 2026
Differentiating Functional Roles of Gene Expression from Immune and Non-immune Cells in Mouse Colitis by Bone Marrow Transplantation
Published on: October 1, 2012
EMS-induced mutant frequency and spectrum in bone marrow of D6-2 transgenic mice
1Department of Biology, Open Laboratory of Medical Molecular Genetics, Second Military Medical University, 200433, Shanghai, China.
Abstract:
EMS-induced mutant frequency and mutation spectrum as well as background mutant frequency have been characterized fur bone marrow of the D6-2 transgenic mice. ThelacI genes carried on pSPORT1 vectors were recovered from the treated or untreated mouse genomic DNA by excision and circularization, and analyzedin vitro for mutations that occurred in the mouse bone marrow, lacI(-) mutants were positively selected with the M9/L media. The 6 lacI(-) mutants were identified out of 11 935 vectors recovered from genomic DNA of the treated mice (mutant frequency was 50 x 10(5)), while no mutant was found in 11 649 vectors Imm untreated mice (the background mutant frequency wan lower than 8.6 x 10(-5)). Two regions oflacI for each mutant, in which the majority of sensitive sites for inactivation of thelacI gene product have been located, were sequenced and 16 mutation events were identified. The predominant mutations (14/16 or 87.5%) were base substitutions, whereas the remaining 2 mutations were single base deletions (12.5%). Of these base substitutions, transversions made up 9/14 or 64%, and transitions cornprised 5/14 or 36%, These findings were markedly different from the spontaneous spectra characterized by using Big-Blue system, as well as from the EMS-induced mutation spectra obtained within vitro assay systems, where the EMS-induced predominant mutations are CG --> AT transitions. In addition, 45% of mutations analyzed occurred at CpG dinucleotides, which was in accordance with previous studies with other systems. These data show that: (i) the D6-2 transgenic mouse lineage is a suitable mdel for studying mutagenesisin vivo; (ii) a fundamental difference in mutagenesis for EMS betweenin nitro andin vivo assay systems may exist, but more extensive sequence analyses are required to determine the possible differences in mutation spectra.
Insights
This study characterizes ethyl methanesulfonate (EMS)-induced mutations in D6-2 transgenic mice. Results show a higher mutation frequency in vivo compared to in vitro assays, suggesting distinct mutagenesis mechanisms.
Area of Science:
- Toxicology
- Genetics
- Molecular Biology
Background:
- Transgenic mouse models are crucial for studying mutagenesis in vivo.
- Understanding mutation spectra induced by chemical mutagens like ethyl methanesulfonate (EMS) is vital for risk assessment.
Purpose of the Study:
- To characterize the mutant frequency and mutation spectrum induced by EMS in the bone marrow of D6-2 transgenic mice.
- To compare in vivo EMS mutagenesis with in vitro findings.
Main Methods:
- Utilized the lacI gene in pSPORT1 vectors recovered from mouse genomic DNA.
- Employed positive selection for lacI(-) mutants and DNA sequencing to identify mutations.
- Analyzed mutation types, including base substitutions and deletions, and their distribution.
Main Results:
- Identified a mutant frequency of 50 x 10^-5 in EMS-treated mice, significantly above the background (<8.6 x 10^-5).
- Sequencing revealed 16 mutations, predominantly base substitutions (87.5%), with transversions (64%) being more common than transitions (36%).
- Observed differences in mutation spectra compared to spontaneous mutations and in vitro EMS assays, with 45% of mutations occurring at CpG dinucleotides.
Conclusions:
- The D6-2 transgenic mouse is a suitable model for in vivo mutagenesis studies.
- A fundamental difference in EMS mutagenesis may exist between in vitro and in vivo systems.
- Further sequence analysis is needed to fully elucidate in vivo EMS mutation spectra.
