Gordon Johnston
Congenital Heart Disease and Nkx2-5: The Search for Modifier Genes
Mentor:  Dr. Patrick Jay, Departments of Pediatrics and Genetics, Washington University Schoolof Medicine 

Congenital heart disease affects 1 of every 100 newborns and is the leading non-infectious cause of death in children. Nkx2-5 is an evolutionarily conserved cardiac transcription factor that is essential for normal heart development. Heterozygous mutations in man lead to a variety of heart defects. The molecular mechanisms that lead to specific defects or that cause variable penetrance are not understood, however.  Discovery of the modifier genes that contribute to these defects may suggest novel strategies for diagnosis and prevention. We are using genetic linkage analysis with an Nkx2-5 mouse knockout model and inbred strain crosses to find these genes. My summer project entailed the development of PCR-based markers to map genomic loci that are associated with the heart defect phenotype in Nkx2-5 mutation.  Microsatellites are repetitive sequences of DNA scattered across the genome.  The length of the repeats can vary between inbred mouse strains.  Polymorphic microsatellites can therefore be used to distinguish the origin of a chromosomal region in an inbred strain cross. Using the mouse genome database, I found potential markers approximately 15-20 cM apart that spanned the entire genome.  I then tested primer pairs in PCR assays, using parental strain mouse and F1 hybrid genomic DNA as the template. I sought primer pairs that were polymorphic between the parental strains and resolvable by agarose gel electrophoresis. Among 98 primers tested, 64 were polymorphic and can be for future genetic linkage analyses