Over the past decade, the construction of Bacterial Artificial Chromosome (BAC) libraries has revolutionized gene mapping in marsupials and monotremes, and has been invaluable for genome sequencing, either for sequencing target regions or as part of whole genome sequencing projects, making it possible to include representatives from these two major groups of mammals in comparative genomics studies. Marsupials and monotremes bridge the gap in vertebrate phylogeny between reptile-mammal divergence 310 million years ago and the radiation of eutherian (placental) mammals 105 million years ago (Fig. 1). The inclusion of these interesting species in such studies has provided great insight and often surprising findings regarding gene and genome evolution. In this chapter, I will review the important role BACs have played in marsupial and monotreme comparative genomics studies.