A large-scale screening of Minos-inserted mutants and analysis of genes involved in the mutation in the chordate Clone intestinalis

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Ciona intestinalis offer the promise of forward-genetics since their life cycle is relatively short, 8-12 weeks from fertilization to Fl embryos, and it is possible to induce self-fertilization within hermaphrodites. The compact genome may also offer opportunity to accomplish mutations leading to visible phenotypes. In Ciona, successful germ-line transgenesis with the Tc1/mariner superfamily transposon, Minos, has been established by ourselves. The aim of the present study is to isolated Minos-inserted mutants as many as possible, and to identify genes that are responsible for the mutation. The present study is also intended to introduce various transgenic techniques into the Ciona system. During the period of 2005-2008, we have isolated approximately 50 mutation lines and 20 transgenic lines. The mutation lines include swimming juveniles (sj) 1-4, balloon (bal), and tail regression failed (trf). All these mutants showed abnormal process of metamorphosis. Interestingly, the sj- I mutant phenotype arises form the insertion of Minos into the promoter region of Ci-CesA, a gene encoding cellulose synthase. We are now analyzing molecular mechanisms involved in the mutations using microarray technique. Transgenic lines include those with GFP expression in embryonic muscle, notochord, or nervous system. These provide useful marker lines for further studies of the expression and function of chordate genes.