Chimeric gene constructs for generation of fluorescent transgenic ornamental fish

Abstract

Four zebrafish gene promoters, which are skin specific, muscle specific, skeletal muscle specific and ubiquitously expressed respectively, were isolated and ligated to the 5′ end of the EGFP gene. When the resulting chimeric gene constructs were introduced into zebrafish, the transgenic zebrafish emit green fluorescence under a blue light or ultraviolet light according to the specificity of the promoters used. Thus, new varieties of ornamental fish of different fluorescence patterns, e.g., skin fluorescence, muscle fluorescence, skeletal muscle-specific and/or ubiquitous fluorescence, are developed.

1 - 41 . (canceled) 
     
     
         42 . A transgenic fish comprising a chimeric gene comprising a cytokeratin (CK) gene promoter operably linked to a fluorescence gene encoding a fluorescent protein, wherein the transgenic fish expresses the fluorescent protein at a level sufficient that the transgenic fish fluoresces upon exposure to sunlight. 
     
     
         43 . The transgenic fish of  claim 42 , wherein the CK gene promoter is expressed in skin epithelia of the fish, 
     
     
         44 . The transgenic fish of  claim 42 , wherein the CK gene promoter comprises the gene sequence set forth in SEQ ID NO: 7. 
     
     
         45 . The transgenic fish of  claim 42 , wherein the fluorescence gene is under control of the CK gene promoter. 
     
     
         46 . The transgenic fish of  claim 42 , wherein the transgenic fish contains the promoter in germ cells and/or in somatic cells and which is capable of breeding with either a the transgenic fish or a non-transgenic fish to produce viable and fertile transgenic progeny. 
     
     
         47 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof emits green fluorescence when the whole transgenic fish or the progeny thereof is exposed to a blue or ultraviolet light. 
     
     
         48 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses a Green Fluorescent Protein. 
     
     
         49 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses an Enhanced Green Fluorescent Protein. 
     
     
         50 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses a Blue Fluorescent Protein. 
     
     
         51 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses an Enhanced Blue Fluorescent Protein. 
     
     
         52 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses an Enhanced Yellow Fluorescent Protein. 
     
     
         53 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses a Cyan Fluorescent Protein. 
     
     
         54 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses an Enhanced Cyan Fluorescent Protein. 
     
     
         55 . The transgenic fish of  claim 42 , wherein the transgenic fish or a progeny thereof expresses more than one fluorescent proteins. 
     
     
         56 . The transgenic fish of  claim 42 , wherein the transgenic fish is selected from a group consisting of medaka, goldfish, carp, koi, loach, tilapia, glassfish, catfish, angel fish, discus, eel, tetra, goby, gourami, guppy,  Xiphophorus  (swordtail), hatchet fish, Molly fish and  Pangasius.    
     
     
         57 . The transgenic fish of  claim 42 , wherein the transgenic fish is a zebrafish. 
     
     
         58 . A transgenic fish whose genome comprises a transgene comprising a promoter consisting of the nucleic acid sequence set forth by SEQ ID NO: 7, wherein the promoter is operably linked to a fluorescence gene encoding a fluorescent protein, wherein the transgene fish is capable of breeding to produce viable and fertile transgenic progeny that express the fluorescence protein when exposed to sunlight.