The molecular mechanisms mediating morphogenesis during mammalian embryogenesis are poorly understood. In part this is related to the fact that few mammalian genes known to control specific aspects of morphogenetic development have been cloned and characterized. The recent development of techniques to insert new genetic information into the mouse genome and to create transgenic mice provides a new methodology to simultaneously mutate and molecularly tag mammalian genes. The major goal of the research proposed herein is to develop a vector system that significantly simplifies the search for, and maintenance of, transgenic mice with developmental disorders caused by insertional mutagenesis.
The specific aims are 1) to develop a visual identification system (VlS) vector that will permit hemizygous and homozygous transgenic mice to be identified and distinguished by simple visual inspection, and then to sue the vector to generate new families of transgenic mice which will be screened for insertional developmental mutations; 2) to use the VIS vector to evaluate the frequency of integration by homologous recombination in microinjected mouse embryos; and 3) to use the VIS vector to evaluate the frequency of integration by gap repair in microinjected mouse embryos. The VIS vector to be tested contains a lens-specific crystallin promotor linked to a truncated, non-tumorigenic, SV40 early region. The vector will be tested for dominant induction of cataract formation, an easily identifiable visual phenotype. The downless gene will be the target for the homologous recombination experiments and the OTCase gene will be used for the gap repair experiments. The identification by insertional mutagenesis of mammalians that play specific roles in development, and the improvement of techniques for gene-specific mutagenesis in mammalian organisms are important steps toward a better understanding of the molecular basis for mammalian embryogenesis and, morphogenesis. Such studies can have important implications for treatment of human infertility and prevention of human birth defects.
