CDK5RAP1 siRNA (h)

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	The human malignant melanoma A375 cell line (A375-P) was purchased from American Type Culture Collection (Manassas, VA, USA). In accordance with experimental guidelines and ethical approval of Harbin Medical University (Harbin, China), the study was performed in Harbin Medical University. A375 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 mg/ml streptomycin (all from Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) with 5% CO2 at 37°C in a humidified incubator (Sanyo Electric Co., Ltd., Tokyo, Japan).Control siRNA and the CDK5RAP1 siRNA were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). A375 cells were seeded at a density of 1×105 cells/well onto six-well plates. After cells obtained 60–80% confluency, siRNAs were transfected into A375 cells according to the manufacturer's protocol. A375 cells were incubated for another 48 h prior to use in subsequent experiments. The transfection efficiency was analyzed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), according to the protocol outlined below.

Protocol tips

The human malignant melanoma A375 cell line (A375-P) was purchased from American Type Culture Collection (Manassas, VA, USA). In accordance with experimental guidelines and ethical approval of Harbin Medical University (Harbin, China), the study was performed in Harbin Medical University. A375 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 mg/ml streptomycin (all from Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) with 5% CO2 at 37°C in a humidified incubator (Sanyo Electric Co., Ltd., Tokyo, Japan).Control siRNA and the CDK5RAP1 siRNA were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). A375 cells were seeded at a density of 1×105 cells/well onto six-well plates. After cells obtained 60–80% confluency, siRNAs were transfected into A375 cells according to the manufacturer's protocol. A375 cells were incubated for another 48 h prior to use in subsequent experiments. The transfection efficiency was analyzed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), according to the protocol outlined below.

Publication protocol

The human malignant melanoma A375 cell line (A375-P) was purchased from American Type Culture Collection (Manassas, VA, USA). In accordance with experimental guidelines and ethical approval of Harbin Medical University (Harbin, China), the study was performed in Harbin Medical University. A375 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 mg/ml streptomycin (all from Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) with 5% CO2 at 37°C in a humidified incubator (Sanyo Electric Co., Ltd., Tokyo, Japan).Control siRNA and the CDK5RAP1 siRNA were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). A375 cells were seeded at a density of 1×105 cells/well onto six-well plates. After cells obtained 60–80% confluency, siRNAs were transfected into A375 cells according to the manufacturer's protocol. A375 cells were incubated for another 48 h prior to use in subsequent experiments. The transfection efficiency was analyzed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), according to the protocol outlined below.

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