Analyses of mutations affecting life span in model organisms have revealed a number of genes that regulate longevity in evolutionarily conserved signaling pathways. These studies suggest that genes involved in insulin-like signaling pathways, metabolism, stress response, and prevention of oxidative damage influence life span. However, we do not know whether functional polymorphisms at these candidate genes affect population variation in. longevity. To identify naturally occurring molecular polymorphisms that are responsible for variation in. life span, we must first map the quantitative trait gene (QTG), followed by linkage disequilibrium mapping in a large sample of alleles collected from a natural population. Genome-wide recombination mapping is a well developed approach for identifying the chromosomal regions (quantitative trait loci [QTLs]) where the QTGs affecting variation in life span between two strains map. The challenge for this approach has been to resolve the QTL to the level of individual genes. This chapter reports details of quantitative complementation tests and linkage disequilibrium mapping to identify positional genes and causative genetic polymorphisms determining variation in Drosophila longevity. © Humana Press Inc.