Establishment of oct4:egfp transgenic and oct4:egfp /β-actin:DsRed double transgenic medaka lines
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As a model to examine cellular multipotency in fish, we established a medaka transgenic (Tg) Tru.oct4:egfp line carrying the green fluorescence protein (GFP) cDNA under control of the Takifugu rubripes oct4 promoter. In this Tg line, GFP could be used to examine both maternal and zygotic oct4 expression during embryogenesis. In addition, while adult Tg fish did not express GFP in any somatic cells, activation of GFP expression was initiated in regenerating fins after amputation. In vitro, some of the cell populations that migrated from fin explants expressed GFP, implying that GFP could be used to monitor oct4 expression in both embryos and in regenerating tissues in the Tru.oct4:egfp Tg line. Next, crossing with β-actin:DsRed Tg line in which all cells emit red fluorescence by expression of red fluorescent protein (RFP) under the β-actin promoter, we prepared a Tru.oct4:egfp /β-actin:DsRed double Tg line. In the double Tg line, early embryonic cells were +GFP/+RFP double positive. In vitro fin cell culture, a small number of +GFP/+RFP double positive cells could be discriminated from other −GFP/+RFP cells. Thus, when transplanted into wild-type medaka, this double Tg line can be used to trace the fate of the transplanted cells using RFP fluorescence after the loss of GFP expression.
KeywordsTransgenic fish Medaka oct4 GFP RFP Embryo Fin regeneration
We thank the National BioResource Project NBRP Medaka for kindly providing medaka strains, d-rR and Tg(β-actin:DsRed). We also thank Dr Joachim Wittbrodt for kindly providing the I-SceI vector. This research was supported by Grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan to TS (Chosenteki-Hoga: 25660151, Kiban-B: 26292110), to HY (Kiban C: 15K07571) and by The Towa Foundation for Food Research.
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