The development of left-right asymmetry of the heart and viscera is a fundamental process in vertebrate embryogenesis. The left-right axis is established at a later stage of development than either the antero- posterior or dorsoventral axes and is first apparent in the formation of the asymmetric cardiac loop from the previously symmetric midline cardiac tube. The mechanism of determination of left-right asymmetry and normal cardiac and visceral situs remains unknown. This project takes a genetic approach to the problem by analyzing the recessively inherited mouse mutation, iv, that leads to loss of control over the development of normal cardiac and visceral situs: 50% of mice that are homozygous for the iv mutation have dextrocardia and situs inversus, 50% have levocardia nad situs solitus. To study the normal function of Iv, we are taking a """"""""reverse genetic"""""""" approach. We have mapped Iv to a location on distal mouse chromosome 12 by its segregation in a backcross with respect on distal mouse chromosome 12 by its segregation in a backcross with respect to DNA markers defined by RFLPs (Restriction Fragment Length Polymorphisms). We are now planning to isolate and identify DNA sequences containing Iv. A physical map of the region is being generated by pulsed field gel analysis. Further clones that map into the region will be isolated by strategies that include production of a radiation fusion cell line, chromosomal microdissection followed by PCR amplification, and the cloning of fragments isolated from pulsed field gels. DNA clones encompassing the region will then be isolated from phage, cosmid or YAC libraries. THe clone containing Iv will be identified by functional analysis in transgenic mice, enabling the gene and its product to be characterized.
