The Effect of Linear Versus Circular Vector on Enhanced Green Fluorescent Protein (EGFP) Expression in Transgenic Zebrafish (Danio rerio)

Abstract

Green Fluorescent Protein (GFP) has been used as an indicator of transgene expression in living cells and organisms. In this study, a transgene construct containing the Cyto-Megalo-Virus (CMV) promoter sequences, SV40 polyA signal and the Enhanced Green Fluorescent Protein reporter gene (EGFP) was microinjected into the cytoplasm of one-cell zebrafish embryos. About 65 ng mu L-1 circular and linearized pEGFP-N1 DNA was used in microinjection. Transgenic founders were detected by Polymerase Chain Reaction (PCR), Slot and Southern blots and Reverse-Transcriptase PCR (RT-PCR). EGFP gene expression was detected by inverted fluorescence microscope in F-0 transgenic zebrafish larvae. About 54 and 25 F-0 transgenic zebrafish were obtained after microinjection of linearized and circular gene constructs, respectively. This is the first study for generation of transgenic zebrafish via cytoplasmic DNA microinjection in Turkey. In conclusion, these results indicate that the gene expression efficiency of circular form was higher than the linearized form in F-0 transgenic zebrafish larvae.