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Fig. 6. PLC{zeta} quantitation in cRNA-microinjected eggs and mouse sperm. (A) Ca2+ changes in a fura-red loaded mouse egg microinjected with cRNA encoding Myc-PLC{zeta} at 0.02 mg/ml. (B) Immunoblot analysis of Myc immunoreactive protein in uninjected (U) and Myc-PLC{zeta} cRNA-injected (I) mouse eggs (240 eggs/lane, 2 mg/ml, 5 hours post-injection). Molecular weight markers in kDa are on right. (C) Immunoblot and relative mobility analysis of native PLC{zeta} in mouse sperm (Sp, left panel, anti-PLC{zeta} antibody) and of Myc-PLC{zeta} in mouse eggs (right panel, anti-Myc antibody) microinjected with 1.0, 0.3 and 0.1 mg/ml Myc-PLC{zeta} cRNA (100 eggs/lane, 5 hours post-injection). 80 kDa protein marker is on the left. (D) Densitometric calibration plot of E. coli-purified Myc-PLC{zeta}-Histag protein using anti-PLC{zeta} antibody. Correlation coefficient, r=0.99. Broken line indicates interpolation of PLC{zeta} protein content corresponding to sperm extract derived from 4x105 mouse sperm.