Co-immunoprecipitation was performed with postnatal day 20 mouse testes using affinity-purified MORC2B antibodies as previously described [34 (link)]. Immunoprecipitated proteins were resolved by SDS-PAGE. The protein bands specific to the wild type testis sample were subjected to mass spectrometry for protein identification.
The full-length coding sequences of mouse Morc2a and Morc2b were cloned into pcDNA3.1/myc-His vector and pcDNA3.1/V5-His-TOPO vector respectively. Plasmids were transfected into HEK 293T cells. Forty-eight hours after transfection, cells were collected and lysed in whole cell lysis buffer (50 mM HEPES, pH 7.5, 140 mM NaCl,1 mM DTT, 10% glycerol, 0.5% NP-40, 1 mM PMSF). Immunoprecipitation on protein lysate was performed with anti-V5 antibody (Catalogue number R96025, Invitrogen), followed by Western blotting with anti-Myc antibody (Catalogue number 631206, Clontech).
The full-length coding sequences of mouse Morc2a and Morc2b were cloned into pcDNA3.1/myc-His vector and pcDNA3.1/V5-His-TOPO vector respectively. Plasmids were transfected into HEK 293T cells. Forty-eight hours after transfection, cells were collected and lysed in whole cell lysis buffer (50 mM HEPES, pH 7.5, 140 mM NaCl,1 mM DTT, 10% glycerol, 0.5% NP-40, 1 mM PMSF). Immunoprecipitation on protein lysate was performed with anti-V5 antibody (Catalogue number R96025, Invitrogen), followed by Western blotting with anti-Myc antibody (Catalogue number 631206, Clontech).
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