Cells were incubated in Dulbecco’s modified Eagle medium/F-12 culture medium with 10% fetal bovine serum (Gibco, 16,000–044) and 100 U/ml penicillin and streptomycin. All cells were cultured in a humidified incubator containing 5% CO2 at 37 °C.
For knockdown of FASN mRNA, negative control siRNA (5′- UUCUCCGAACGUGUCACGUTT-3′), FASN siRNA (5′-CAGAGUCGGAGAACUUGCAGGAGUU-3′), α-tubulin siRNA (5′-AAAGATGTCAATGCTGCCATT-3′), and β-tubulin siRNA (5′-CCCAGCGGCAACTACGTGGG-3′) were designed [1 (link), 16 (link), 24 (link)], SREBP1 siRNA was purchased from Santa Cruz Biotech (sc-36558). Cells were plated in six-well plates at a density of 1 × 106/well and cultured for 24 h. Then the cells were transfected with the FASN and α-tubulin siRNA, respectively (30 pmol/well) by LipofectamineTM RNAiMAX (Invitrogen, 13778075) when cells reach ~80% confluency.
For knockdown of FASN mRNA, negative control siRNA (5′- UUCUCCGAACGUGUCACGUTT-3′), FASN siRNA (5′-CAGAGUCGGAGAACUUGCAGGAGUU-3′), α-tubulin siRNA (5′-AAAGATGTCAATGCTGCCATT-3′), and β-tubulin siRNA (5′-CCCAGCGGCAACTACGTGGG-3′) were designed [1 (link), 16 (link), 24 (link)], SREBP1 siRNA was purchased from Santa Cruz Biotech (sc-36558). Cells were plated in six-well plates at a density of 1 × 106/well and cultured for 24 h. Then the cells were transfected with the FASN and α-tubulin siRNA, respectively (30 pmol/well) by LipofectamineTM RNAiMAX (Invitrogen, 13778075) when cells reach ~80% confluency.
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