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. 2015 Jul 6;112(29):9034–9039. doi: 10.1073/pnas.1501032112

Fig. 3.

Fig. 3.

Neurofibromin depletion with siRNA in mel-WT and mel-NF1 cells. (A) qRT-PCR analysis of NF1 transcript expression in mel-WT-1 and mel-WT-2 cells transfected with either siRNA targeting NF1 (siNF1) or control siRNA (siCtrl). qRT-PCR levels are presented as fold change relative to siCtrl-mel-WT cells. Results are expressed as mean ± SD (n = 3). (B) Western blot analysis of neurofibromin expression in mel-WT-1 and mel-WT-2 cells transfected with siNF1 or siCtrl. β-tubulin served as a loading control (n = 1). (C) Melanin content in siCtrl-mel-WT-1 and WT-2 and siNF1-mel-WT-1 and WT-2 cells. Measurements were performed using 105 cells of each cell type. Data are presented as mean ± SD (n = 3) normalized to the expression in siCtrl-mel-WT-1 cells. (D) Western blot analysis of tyrosinase expression in siNF1-mel-WT-1 and WT-2 and siCtrl-mel-WT-1 and WT-2 cells. β actin served as a loading control. Densitometry measurement of protein levels was relative to siCtrl-mel-WT-1 cells. Results are expressed as mean ± SD (n = 3). (E) qRT-PCR analysis of NF1 transcript expression in mel-NF1-1 and NF1-2 cells transfected with siNF1 and siCtrl. qRT-PCR values are presented as fold change relative to siCtrl-mel-NF1 cells. Results are expressed as mean ± SD (n = 3). (F) Western blot analysis of neurofibromin expression in mel-NF1-1 and mel-NF1-2 cells transfected with siCtrl or siNF1. β-tubulin served as a loading control (n = 1). (G) Melanin content in siCtrl-mel-NF1-1 and NF1-2 and siNF1-mel-NF1-1 and NF1-2 cells. Measurements were performed using 105 cells of each cell type. The results are expressed as a relative level normalized to siCtrl-mel-NF1-1 cells. Data are presented as mean ± SD (n = 3). (H) Western blot analysis of tyrosinase expression in siCtrl-mel-NF1and siNF1-mel-NF1 cells. β-actin served as a loading control. Results are expressed as mean ± SD (n = 3), *P < 0.05, **P < 0.01, ***P < 0.001, ANOVA followed by Dunnett’s multiple-comparison test with siCtrl-mel-WT cells (A, C, and D) or siCtrl-mel-NF1-1 cells (E, G, and H).