Functional genomics in ascidians through insertional mutagenesis and homologous recombination

Ascidians, or sea squirts, are marine invertebrate chordates. Their critical evolutionary position as basal chordates and the simplicity of their embryogenesis have attracted great attention from developmental and evolutionary biologists. Recently, ascid ians have become one of the most promising animal models for functional genomics and system biology. Extensive resources, including fully sequenced genomes and bioinformatic platforms on gene expression and regulation, have become available. Advanced mo lecular techniques, such as transient and stable transgenesis, have been developed. However, efficient mutagenesis methods, which are essential for probing developmental gene function, are still absent. In order to appreciate the full potential of asc idians as a functional genomic tool, this application seeks to develop effective mutagenesis strategies in ascidians. We will improve upon a transposon-mediated gene-breaking vector, already successful in zebrafish, for a large scale insertional mutagene sis in ascidians. In addition, we will adapt homologous recombination (knockout) technique for targeted mutagenesis. Mutant animals generated by both methods will be used to study essential function of ascidian genes in development and physiology. The results of these efforts will be made available to international research community through construction of an online database that is integrated with existing ascidian genomic and developmental biology databases.