Analysis of the immune response to ectopically expressed allergen gene into epidermis

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To analyze the immunological response in epidermis of the skin, and to establish the animal model system of atopic dermatitis, we conducted to generate gene-targeting mouse which manifests epidermal keratinocyte specific-expression of green fluorescent protein (GFP). To escape the induction of immunological tolerance to ectopically introduced xenoantigen at the embryonic stage, we applied tet-on system to express GFP by epidermal keratinocytes under the control of tetracycline. Following methods were applied in this project: 1) to design the construct which transduces rtTA, downstream of epidermal basal cell specific keratin 5 promoter, in the presence of doxycycline and to establish the gene targeting- mouse in which a poly A signal of human growth hormone was introduced to downstream of rtTA(K5-rtTA2S-M2), (2) to establish the gene targeting mouse in which membranous GFP construct and SV40 poly A signal construct with insertion of insulator (Tet-EGFP Thy1). We already confirmed the designed constructs were successfully introduced into the mouse by southern blot analysis. We are now under investigation to analyze immune response in these mice.