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Fig. 6 | Breast Cancer Research

Fig. 6

From: A method of producing genetically manipulated mouse mammary gland

Fig. 6

Establishment of PyMT oncogene-introduced mammary glands. a Vector construct of piggyBac transposon donor vector expressing mCherry, firefly luciferase (FLuc), and Tet3G under the control of the mouse mammary tumor virus (MMTV) promoter. Polyoma-virus middle T antigen (PyMT) was introduced downstream of TRE3G using LR recombinase of the Gateway system. b Immunoblot analysis of indicated lysates derived from NMuMG cells not transfected or transfected with TRE-PyMT vector. Tubulin represents internal control. c (left panel) A representative image showing luciferase activity of mammary outgrowths at day 42 after mammary epithelial cell injection into cleared fat pad. (Right panels) Microscopic view of resected fat pads. Red fluorescence (lower panel) shows mCherry marker expression. Scale bar = 5 mm. d Analysis of PyMT expression by immunofluorescence staining. Frozen sections from TRE-PyMT-introduced mammary glands passaged from a primarily reconstituted gland without Dox (right panels) and with Dox (left panels) in drinking water were co-immunostained with antibodies against mCherry (red, upper panels) and PyMT (green). Scale bars = 80 μm. DAPI 4′,6-diamidino-2-phenylindole, Dex dexamethasone, Dox doxycycline

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