The goal of this research is to identify specific genetic variants that affect bone fragility. In the previous iinding period of this project, we identified an inbred strain of rats (F344) that has lower bone mineral density and more fragile bones than a second rat strain (LEW). From F344 and LEW rat strains, we created an F2 population for genetic mapping. We identified quantitative trait loci (QTLs) for skeletal phenotypes on Chromosomes (Chrs) 1, 2, 4, 5, 7, 8, 10,15 and 19. LOD scores for these QTLs ranged from 4.0 to 19.6 and were significant at p<0.05. QTLs at two of these loci, Chrs 5 and 8, are syntenic with QTLs observed in our sibling pair population (Projects 1 and 2). The QTL on rat Chr 5 is syntenic with human Chr 1p, which was shown to be linked to proximal femoral bone density. In addition, the QTL on rat Chr 8 is syntenic with human Chr 15q that was shown to be linked to hip BMD. In the next five years, we will isolate the effects of key QTLs by breeding congenic rats. Each QTL may nteract with one or more other QTLs thus complicating the genetic picture. One way to isolate the effect of a single QTL is to create a congenic line. This will be done using a backcross breeding strategy. The F344 rats will be intercrossed with LEW rats and the offspring backcrossed with LEW rats for 10 generations. Through each generation, the rats will be genotyped at the QTL to identify carriers of F344 DMA within the QTL. The resulting congenic line is over 99.9% LEW with only a small region of F344 DMA at the QTL. We plan to make congenic rat lines for the Chrs 5 and 8 QTLs to isolate the chromosomal regions in the rat that match QTLs in humans. In addition to the congenic approach, we will examine gene expression in bone to identify candidate genes. Gene expression will be measured using the Affymetrix rat microarray. We will compare skeletal gene expression in F344 and congenic lines with LEW rats to assess differential gene expression during bone growth. Our goal is to identify genes that fall within the QTLs and are also differentially expressed. These genes will be flagged as strong candidates and tested in our sibling pair population (Project by Econs). We hypothesize that numerous candidate genes will be identified using this method.
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