Immunoblotting and Immunoprecipitation of Nuclear Proteins

Total and nuclear lysates prepared from subconfluent cells were subjected to immunoblot analysis using anti-MUC1-C (#HM-1630-P1ABX; Thermo Fisher Scientific, Waltham, MA, USA), anti-MYC (#ab32072; Abcam), anti-ERα (#ab108398; Abcam) anti-β-actin (#A5441; Sigma), anti-MTA1 (#5647), anti-MBD3 (#14540), anti-CHD4 (#11912), anti-HDAC1 (#5356), anti-SOX2 (#D6D9), anti-KLF4 (#D1F2), anti-BMI1 (#D20B7), anti-CD44 (#156-3C11), anti-OCT4 (#2750S; Cell Signaling Technology, Danvers, MA, USA). Nuclear proteins were immunoprecipitated in the absence and presence of 50 μg/ml ethidium bromide (EtBr; #15585-011, Thermo Fisher Scientific) as described (25 (link)).

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Hata T., Rajabi H., Takahashi H., Yasumizu Y., Li W., Jin C., Long M.D., Hu Q., Liu S., Fushimi A., Yamashita N., Kui L., Hong D., Yamamoto M., Miyo M., Hiraki M., Maeda T., Suzuki Y., Samur M.K, & Kufe D. (2019). MUC1-C ACTIVATES THE NuRD COMPLEX TO DRIVE DEDIFFERENTIATION OF TRIPLE-NEGATIVE BREAST CANCER CELLS. Cancer research, 79(22), 5711-5722.

Publication 2019

anti-MUC1-C (#HM-1630-P1ABX; anti-MYC anti-ERα anti-β-actin anti-MBD3 anti-CHD4 anti-HDAC1 anti-SOX2 anti-KLF4 anti-BMI1 anti-CD44 anti-OCT4

Actin Bmi1 Cd44 Cells Chd4 Etbr Ethidium bromide Immunoblot analysis Klf4 Mbd3 Mta1 Muc1 Nuclear proteins Oct4 Sox2

Corresponding Organization : Dana-Farber Cancer Institute

Other organizations : Roswell Park Comprehensive Cancer Center

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Variable analysis

independent variables

Presence of 50 μg/ml ethidium bromide (EtBr)

dependent variables

Levels of MUC1-C, MYC, ERα, β-actin, MTA1, MBD3, CHD4, HDAC1, SOX2, KLF4, BMI1, CD44, OCT4

control variables

Nuclear proteins

controls

Immunoprecipitation in the absence of 50 μg/ml ethidium bromide (EtBr)

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