IRF-1 siRNA (m)

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	B16F10 murine melanoma cells and A375 human melanoma cells were obtained from the National Centre for Cell Sciences, (Pune, India) and were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal calf serum (FCS), 120 mg/ml penicillin, 200 mg/ml streptomycin and 2 mM L-glutamine. The cells were cultured in a humidified 5% CO2 incubator at 37°C. Cells were regularly cultured to maintain exponential growth of cells. Twenty-four hrs before transfection, cells were diluted in fresh medium without antibiotics and transferred to 24-well plates. Transient transfection of PKCδ siRNA was performed using a Lipofectamine PLUS reagent (Invitrogen, CA, USA) according to the manufacturer’s recommendations. The efficiency of the transfection was monitored every 12 hrs. At 36 hrs, PKCδ expression was totally inhibited. Specific silencing was confirmed by at least three independent experiments. The cells were also treated with transfection reagent alone or with the non silencing scrambled PKCδ siRNA.

Protocol tips

B16F10 murine melanoma cells and A375 human melanoma cells were obtained from the National Centre for Cell Sciences, (Pune, India) and were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal calf serum (FCS), 120 mg/ml penicillin, 200 mg/ml streptomycin and 2 mM L-glutamine. The cells were cultured in a humidified 5% CO2 incubator at 37°C. Cells were regularly cultured to maintain exponential growth of cells. Twenty-four hrs before transfection, cells were diluted in fresh medium without antibiotics and transferred to 24-well plates. Transient transfection of PKCδ siRNA was performed using a Lipofectamine PLUS reagent (Invitrogen, CA, USA) according to the manufacturer’s recommendations. The efficiency of the transfection was monitored every 12 hrs. At 36 hrs, PKCδ expression was totally inhibited. Specific silencing was confirmed by at least three independent experiments. The cells were also treated with transfection reagent alone or with the non silencing scrambled PKCδ siRNA.

Publication protocol

B16F10 murine melanoma cells and A375 human melanoma cells were obtained from the National Centre for Cell Sciences, (Pune, India) and were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal calf serum (FCS), 120 mg/ml penicillin, 200 mg/ml streptomycin and 2 mM L-glutamine. The cells were cultured in a humidified 5% CO2 incubator at 37°C. Cells were regularly cultured to maintain exponential growth of cells. Twenty-four hrs before transfection, cells were diluted in fresh medium without antibiotics and transferred to 24-well plates. Transient transfection of PKCδ siRNA was performed using a Lipofectamine PLUS reagent (Invitrogen, CA, USA) according to the manufacturer’s recommendations. The efficiency of the transfection was monitored every 12 hrs. At 36 hrs, PKCδ expression was totally inhibited. Specific silencing was confirmed by at least three independent experiments. The cells were also treated with transfection reagent alone or with the non silencing scrambled PKCδ siRNA.

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