| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
Endocrine disruptors, compounds that mimic the action of natural hormones, are present in the environment and represent a public health concern due to potentially disruptive effects on physiological processes including reproduction, fetal development, and the development of cancers. The inability to reliably predict hormonal activities based on structural analysis necessitates the development of functional screening systems to safeguard human health. Therefore, in this project, I have tried to develop the transgenic medaka which can easily detect the estrogenic activity.
The GFP, green fluorescent protein, was used as a reporter protein. I constructed the reporter plasmid, carrying the estrogen response element (ERE) positioned to transcribe the GFP gene. When this reporter plasmid was transfected into HeLa cells with estrogen recertor (ER) expression plasmid, the GFP was induced with the addition of 17β-estradiol (E2). Next, I established the method of microinjection of DNA into fertilized medaka eggs and confirmed the expression of foreign gene in germ-line trasformants and F1 progeny by using constitutive active GFP vector. Finally, I injected a linear form of reporter plasmid into fertilized medaka eggs. When the transgenic medakas were treated with 10 ppb and 100 ppb E2, no GFP expression was detected. It is probably due to the silencing effect by chromosome positioning. The further study is necessary to get transgenic medaka which express GFP in response to endocrine disruptors.
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