Isolation of cDNA Clones for Epidermis-Specific Genes of the Ascidian Embryo

The present investigation was conducted to isolate cDNA clones that correspond to epidermis-specific genes of the ascidian embryo. When cleavage of fertilized eggs of Halocynthia roretzi is blocked by treatment with cytochalasin B and the arrested eggs are reared as one-celled embryos for about 30 hr, they develop features of differentiation of the epidermis only. Translation in vitro of poly(A)⁺ RNA from cleavage-arrested embryos and analysis of the products by two-dimensional gel electrophoresis revealed several predominant polypeptides that were not detected in a similar analysis of fertilized eggs, suggesting the appearance of epidermis-specific mRNAs in cleavage-arrested embryos. A cDNA library was constructed from arrested one-celled embryos. Differential screening of the library with a total cDNA probe from cleavage-arrested embryos and with a similar probe from fertilized eggs yielded eight different cDNA clones specific for the cleavage-arrested embryos. Northern blot analysis revealed that the mRNAs that corresponded to these cDNAs were present in normal tailbud embryos. In addition, in situ hybridization of whole-mount specimens showed that the mRNAs were restricted to the epidermal cells of tailbud embryos.