A large-scale gene-trap screen for insertional mutations in developmentally regulated genes in mice

Area of Science:

• Developmental Biology
• Genetics
• Molecular Biology

Background:

• Gene-trap screens are valuable for identifying insertional mutations.
• Understanding gene regulation during embryogenesis is crucial for developmental biology.

Purpose of the Study:

• To screen for insertional mutations in developmentally regulated genes using a gene-trap vector and mouse embryonic stem cells.
• To assess the temporal and spatial expression patterns of disrupted genes during mouse embryogenesis.

Main Methods:

• Utilized a gene-trap vector in mouse embryonic stem (ES) cells to generate insertional mutations.
• Assayed for disrupted transcription units via beta-galactosidase reporter gene expression.
• Injected modified ES cells into blastocysts to create chimeric mouse embryos for expression analysis at 8.5 and 12.5 days post-coitum (dpc).

Main Results:

• Screened 38,730 ES cell lines, identifying 393 with disrupted transcription units.
• Analyzed 279 chimeric mouse embryos, with 36 (13%) showing restricted expression, 88 (32%) widespread expression, and 155 (55%) no detectable expression at 8.5 dpc.
• Approximately one-third of non-expressing clones at 8.5 dpc showed reporter gene activity at 12.5 dpc, indicating temporal regulation.

Conclusions:

• Gene-trap mutagenesis in ES cells is an effective strategy for isolating mutations in developmentally regulated genes.
• A significant proportion of genes expressed in ES cells exhibit temporal or spatial regulation during embryogenesis.
• This approach facilitates the discovery of novel genes involved in embryonic development.