Arachis hypogaea is a widely cultivated crop both as an oilseed and protein source. The genomic analysis of Arachis species hitherto has been limited to the construction of genetic maps; the most comprehensive one contains 370 loci over 2,210 cM in length. However, no attempt has been made to analyze the physical structure of the peanut genome. To investigate the practicality of physical mapping in peanut, we applied a total of 117 oligonucleotide-based probes ("overgos") derived from genetically mapped RFLP probes onto peanut BAC filters containing 182,784 peanut large-insert DNA clones in a multiplex experimental design; 91.5% of the overgos identified at least one BAC clone. In order to gain insights into the potential value of Arabidopsis genome sequence for studies in divergent species with complex genomes such as peanut, we employed 576 Arabidopsis-derived overgos selected on the basis of maximum homology to orthologous sequences in other plant taxa to screen the peanut BAC library. A total of 353 (61.3%) overgos detected at least one peanut BAC clone. This experiment represents the first steps toward the creation of a physical map in peanut and illustrates the potential value of leveraging information from distantly related species such as Arabidopsis for both practical applications such as comparative map-based cloning and shedding light on evolutionary relationships. We also evaluated the possible correlation between functional categories of Arabidopsis overgos and their success rates in hybridization to the peanut BAC library.