A NEW REPORTER GENE ASSAY FOR DIOXINS USING GREEN FLUORESCENT PROTEIN: INCREASED RESPONSIVENESS USING AMPLIFICATION OF THE DIOXIN RESPONSIVE ELEMENT
A highly responsive green fluorescent protein (GFP) reporter gene assay for dioxins has been newly developed. Initially, we constructed three aryl hydrocarbon receptor (AhR)-responsive GFP reporter vectors containing an increasing number of dioxin responsive elements (DREs) to confer high responsiveness to dioxins. The 5-36 cell line stably transfected with a pZs7.5 vector containing the maximum 20 DREs had the highest GFP responsiveness to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Induction of GFP expression at a low concentration of TCDD (10 pM) in this cell line was nearly 4-fold higher than that in the 1-34 cell line transfected with pZs7.1 containing the minimum 4 DREs. In addition, as compared with the H4G1.1c2 cell line (currently the most widely used AhR-responsive GFP reporter gene assay), the 5-36 cell line had nearly 4-fold higher GFP induction at 10 pM TCDD. The 5-36 cell line generally responded to selected dioxins in line with the level of their toxic equivalent factors, indicating that our assay may be suitable to determine toxic equivalent concentrations of dioxins. This newly developed assay has advantages including rapidity and ease of use; as the result, it will provide a useful high-throughput screen for relatively low concentrations of dioxins.
Tsutsumi T Ishizuka N Denison MS Watanabe T Matsuda R
National Institute of Health Sciences,Kamiyoga 1-18-1,Setagaya-ku,Tokyo 158-8501,Japan Department of Environmental Toxicology,Meyer Hall,University of California,Davis,CA 95616
国际会议
北京
英文
1-4
2009-08-24（万方平台首次上网日期，不代表论文的发表时间）