Cloning, characterization, and transcriptional activity of β-actin promoter of African catfish (Clarias gariepinus).

Abstract

Selection of suitable promoters is crucial for the efficient expression of exogenous genes in transgenic animals. Although one of the most effective promoters, the β-actin promoter, has been widely studied in fish species, it still remains unknown in the economical important African catfish (Clarias gariepinus). In this study, the β-actin promoter of African catfish (cgβ-actinP) was cloned and characterized. In addition, recombinant plasmid pcgβ-actinP-EGFP with enhanced green fluorescent protein (GFP) gene as the reporter gene was constructed to verify the transcriptional activity. We obtained a cgβ-actinP fragment length of 1405bp, consisting 104bp of the 5' proximal promoter, 96bp of the first exon, and 1205bp of the first intron. Similar to those of other fish species, cgβ-actinP contains three key transcription regulatory elements (CAAT box, CArG motif, and TATA box). GFP-specific fluorescent signals were detected in chicken embryonic fibroblasts cells (DF-1 cells) transfected with pcgβ-actinP-EGFP, which was approximately 1.11 times of the positive control. In addition, GFP was effectively expressed in zebrafish larvae microinjected with linearized cgβ-actinP-EGFP, with expression rate reaching approximately 49.84%. Our data indicate that cgβ-actinP could be a potential candidate promoter in the practice of constructing "all fish" transgenic fish.