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Screening and validation of siRNAs in mouse primary hepatocytes.

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posted on 20.02.2013, 04:19 by Huma Safdar, Ka Lei Cheung, Hans L. Vos, Frank J. Gonzalez, Pieter H. Reitsma, Yusuke Inoue, Bart J. M. van Vlijmen

Screening of siRNA in mouse primary hepatocytes 24 hours after transfection with 0.3, 3 and 30nM of three Hnf4a-specific (#1–3, panel A), three Cebpa -specific siRNAs (#1–3, panel B) or a control siRNA (siNEG, panel A and B, open bars). Hnf4a and Cebpa transcript levels were determined by quantitative real-time PCR. β-actin was used as internal control for quantification and normalization. The ΔCt values of the individual samples were related to the mean ΔCt of the reference group (siNEG). On the x-axis siRNA concentrations are indicated. Data are expressed as mean with error bars representing the difference between 2 POWER of upper and lower range of the mean ΔΔCt (difference 2∧ΔΔCt+SEM and 2∧ΔΔCt−SEM). Individual experiments were performed in triplicate. Data were statistically analyzed using the Student's t-test. P-values <0.05 were regarded as statistically significant. *P<0.05, **P<0.01, ***P<0.001.