Genomic cloning of mud loach Misgurnus mizolepis (Cypriniformes, Cobitidae) beta-actin gene and usefulness of its promoter region for fish transgenesis.
As a part of our effort to obtain a strong regulatory element for the construction of an autogenic mud loach transgenic vector, we isolated a genomic clone that contains an open reading frame encoding the beta-actin gene, then examined the transcriptional activity of the upstream sequences, including the first intron, in transiently transfected cell lines. It showed that the upstream region has substantially strong transcriptional activity, and that both the proximal promoter and distal region of intron 1 play a crucial role in the activity. A similar result, based on fish growth, was obtained with the expression vectors containing the growth hormone gene of mud loach. These were driven by the regulatory region of the mud loach beta-actin gene with various mutations, and were directly transferred into the trunk muscle of fish using an electrostimulation-mediated method. Fish weights were monitored over the next 4 weeks. These data suggest that the proximal promoter and the first intron enhancer of the mud loach beta-actin gene are useful for autogenic mud loach transgenesis.
D000199:Actins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003531:Cypriniformes / Q000235:genetics*; D005260:Female; D015870:Gene Expression / Q000235:genetics; D018014:Gene Transfer Techniques; D013006:Growth Hormone / Q000235:genetics / Q000502:physiology; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015183:Restriction Mapping; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection; D019076:Transgenes / Q000502:physiology
D000199:Actins; D013006:Growth Hormone