Construction of BAC Libraries of Metazoans Occupying Key Phylogenetic Positions

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Large-insert libraries of genomic DNA are a valuable resource for comparative studies indevelopment evolution and genetics for addressing such fundamental questions in biology ashow genomes have evolved and how evolution of genomes has resulted in evolution of bodyplans. The goal of the proposed research is to construct 11 BAC libraries representative of thebreadth of the Metazoa. The focus is on animals that occupy phylogenetic positions key to anunderstanding of animal evolution. Preference is given to organisms with demonstrated potentialas developmental model organisms. Therefore, 2 deuterostomes have been chosen: lamprey,basal in the craniates and pivotal for understanding the pattern of gene and chromosomalduplications during vertebrate evolution, and the cephalochordate amphioxus, the closest livinginvertebrate relative of the vertebrates. On the basis of molecular phylogenetics, the protostomeshave recently been divided into two groups, the Ecdysozoa and the Lophotrochozoa. However,branching patterns of the phyla within each of these groups are poorly resolved. Becauserepresentatives of the Ecdysozoa, Drosophila and Caenorhabditis have been well-studied, wehave not chosen any Ecdysozoan. For the Lophotrochozoa, which have, in general, received littleattention in regard to genes, development and evolution, seven organisms have been chosen.These include 2 annelids, one of which, the polychaete Platynereis dumerilii, is relatively basalwithin the annelids and the other of which, the leech Helobdella robusta, is more highly derived.Platynereis is rapidly becoming a lophotrochozoan model developmental organism, and whilethe leech is quite derived, it has been the object of developmental genetic studies for some years.Two molluscs are included-- one bivalve, (Spisula solidissima) and one gastropod (Ilyanassaobsoleta). The other Lophotrochozoans include 1 nemertean (Cerebratulus lacteus), 1 sipunculid(Temiste langiformis), and 1 acoel flatworm (Neochildea fusca). The phylogenetic position ofthe acoels is controversial. On the basis of morphology, they were considered basal Bilateria.However, most, but not all, molecular phylogenetic analyses place acoels within the protostomes.To address bilaterian origins, we have chosen a ctenophore (Mnemiopsis leidyi). Finally, toaddress the origin of the metazoa, a sponge, Callyspongia diffusa, a representative of the mostbasal Metazoa, was selected. BAC libraries will be constructed and distributed by P. de Jong(Children's Hospital Research Institute, Oakland), who has a long track-record in construction ofBAC libraries, including making the BAC library for the human genome project. For organismsfor which interest is high to sequence the entire genome, 15X coverage is proposed. For others,10X coverage is proposed.