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Cellular DNA rearrangements and early developmental arrest caused by DNA insertion in transgenic mouse embryos
Abstract:
Insertional mutagenesis was investigated in a transgenic mouse strain (HUGH/4) derived from a fertilized egg injected with plasmid DNA containing the human growth hormone gene. Lethality occurred in homozygous embryos and was traced to the egg cylinder stage on days 4 to 5 of gestation, shortly after implantation. The mutation is on chromosome 12 and is distinct in location and integration pattern from another mutation also leading to lethality of homozygotes in the egg cylinder stage. Based on this and other evidence, relatively many genes may be recruited to activity near the time of implantation and may therefore present a large target of vulnerability to mutagenesis. The single insert in HUGH/4, consisting of approximately three tandem copies of plasmid sequences, is flanked by mouse cellular sequences that have undergone rearrangements, including a probable deletion. The data suggest the hypothesis that DNA rearrangements, which appear to be commonplace in transgenic mice, may arise because the initial insertional complex is unstable; stepwise changes may then be generated until a more stable conformation is achieved.
Insights
Insertional mutagenesis in transgenic mice (HUGH/4) caused embryonic lethality at implantation. DNA rearrangements at the insertion site suggest instability may drive mutations during early development.
Area of Science:
- Developmental Biology
- Genetics
- Transgenic Models
Background:
- Insertional mutagenesis is a key tool for studying gene function.
- Transgenic mice are valuable models for investigating genetic mutations and developmental processes.
- Early embryonic lethality presents challenges in understanding gene function during implantation.
Purpose of the Study:
- To investigate the effects of insertional mutagenesis in a novel transgenic mouse strain (HUGH/4).
- To characterize the genetic basis and developmental timing of lethality in HUGH/4 homozygotes.
- To explore the relationship between DNA rearrangements and insertional mutagenesis in transgenic models.
Main Methods:
- Generation of transgenic mice (HUGH/4) via pronuclear injection of plasmid DNA containing the human growth hormone gene.
- Phenotypic analysis of homozygous embryos to determine the stage of lethality.
- Genetic mapping to identify the chromosome location of the insertional mutation.
- Molecular characterization of the insertional site, including flanking mouse DNA sequences.
Main Results:
- Homozygous HUGH/4 embryos exhibited lethality at the egg cylinder stage (days 4-5 of gestation), shortly after implantation.
- The mutation was mapped to chromosome 12, with a distinct integration pattern from other known lethal mutations.
- The insertional site featured tandem copies of plasmid DNA flanked by rearranged mouse genomic DNA, including a probable deletion.
Conclusions:
- Insertional mutagenesis in the HUGH/4 strain leads to early embryonic lethality, highlighting the vulnerability of genes active around implantation.
- The observed DNA rearrangements suggest that the initial insertional complex in transgenic mice may be unstable.
- Stepwise DNA rearrangements could be a common mechanism contributing to phenotypic variation and mutations in transgenic models.