The endogenous EF-1α promoter is highly active in driving gene overexpression in developing embryos of the Pacific oyster Crassostrea gigas

Abstract

Gain and loss-of-function analyses are powerful genetic approaches to uncover gene functions in basic and applied research. However, the genetic analysis in bivalve molluscs has been challenged by the lack of effective gene promoters for driving gene overexpression. The cosmopolitan Pacific oyster (Crassostrea gigas) is an economically important marine bivalve and a representative model species for ecological, evolutionary, and developmental studies. Here, we isolated the elongation factor1-α (EF-1α) gene promoter from the Pacific oyster and compared the promoter activity with the commonly used cytomegalovirus (CMV) and Xenopus EF-1α promoters in oyster embryos. We found that the 3465 bp DNA sequence prior to the ATG start codon in oyster EF-1α was able to drive enhanced green fluorescent protein (EGFP) expression in oyster embryos. In contrast, CMV and Xenopus EF-1α promoters failed to direct EGFP expression in oyster embryos. It appeared that the first intron sequence in oyster EF-1α gene was required for the promoter activity. DNA construct without the first intron located upstream of the ATG start codon failed to drive EGFP expression in oyster embryos. Collectively, these studies indicate that the endogenous EF-1α promoter is preferred promoter for driving gene expression in bivalves.