Generation and application of a Tg(cyp1a:egfp) transgenic marine medaka (Oryzias melastigma) line as an in vivo assay to sensitively detect dioxin-like compounds in the environment

Abstract

Large-range environmental pollution by dioxin and dioxin-like compounds (DLCs) is becoming a serious problem. To establish an in vivo method for the detection of DLCs in seawater, a Tg(cyp1a-12DRE:egfp) transgenic marine medaka (Oryzias melastigma) line was first developed with the modified cyp1a-12DRE promoter driving enhanced green fluorescent protein (EGFP) expression using Tol2 transgenesis technology. With increasing concentrations of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs), the EGFP fluorescence intensity increased significantly. The Tg(cyp1a-12DRE:egfp) medaka possessed high sensitivity (limit of detection of 1 ng/L TCDD) and specificity and low background. This transgenic line is capable of detecting DLCs in environmental seawater in which the concentration of DLCs is at least 0.12207 ng/L TCDD after sample enrichment. The fluorescence-toxic equivalency (TEQ) values from EGFP intensity were closely correlated with the chemical-TEQ values obtained from chemical analyses. Furthermore, the Tg(cyp1a-12DRE:egfp) medaka can directly detect DLCs in seawater samples after a serious pollution accident and screen unknown aryl hydrocarbon receptor (AhR) agonists for risk assessment. For the first time, a convenient method has been established that sensitively and specifically responds to DLCs using the Tg(cyp1a-12DRE:egfp) marine medaka, which could be a highly efficient tool for detecting seawater DLCs in the future.

Keywords: AhR agonist; Cyp1a promoter; Dioxin-like compounds; Oryzias melastigma; TEQ.