Silencer®_Faslg siRNA (r)

Protocol tips

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	Predesigned siRNAs directed against rat FasL mRNA (GenBank: NM_012908) were purchased from Ambion (Austin, Texas; siRNA IDs 197271, 48716, and 197272). SiRNAs were transiently transfected into wild-type 9L and F98 glioma cells and screened by immunoblotting for optimal knock-down 48 hours after transfection. A scramble siRNA was used as negative control (Ambion product # AM4636). Transient transfections were performed using 100 nM siRNA combined with siPORT lipid transfection reagent (Ambion, Inc.) in optiMEM media (Gibco) according to the manufacturers' recommendations. Briefly, glioma cells were plated in 100-mm-diameter tissue culture dishes at 5 × 105 cells/plate in medium supplemented with 10% FBS. The following day medium was removed and replaced with medium containing 0.1% FBS. Approximately 1 hour later, siRNA:lipid complexes were added to the cells and allowed to incubate for 48 hours before cells were harvested as described above.

Protocol tips

Predesigned siRNAs directed against rat FasL mRNA (GenBank: NM_012908) were purchased from Ambion (Austin, Texas; siRNA IDs 197271, 48716, and 197272). SiRNAs were transiently transfected into wild-type 9L and F98 glioma cells and screened by immunoblotting for optimal knock-down 48 hours after transfection. A scramble siRNA was used as negative control (Ambion product # AM4636). Transient transfections were performed using 100 nM siRNA combined with siPORT lipid transfection reagent (Ambion, Inc.) in optiMEM media (Gibco) according to the manufacturers' recommendations. Briefly, glioma cells were plated in 100-mm-diameter tissue culture dishes at 5 × 105 cells/plate in medium supplemented with 10% FBS. The following day medium was removed and replaced with medium containing 0.1% FBS. Approximately 1 hour later, siRNA:lipid complexes were added to the cells and allowed to incubate for 48 hours before cells were harvested as described above.

Publication protocol

Predesigned siRNAs directed against rat FasL mRNA (GenBank: NM_012908) were purchased from Ambion (Austin, Texas; siRNA IDs 197271, 48716, and 197272). SiRNAs were transiently transfected into wild-type 9L and F98 glioma cells and screened by immunoblotting for optimal knock-down 48 hours after transfection. A scramble siRNA was used as negative control (Ambion product # AM4636). Transient transfections were performed using 100 nM siRNA combined with siPORT lipid transfection reagent (Ambion, Inc.) in optiMEM media (Gibco) according to the manufacturers' recommendations. Briefly, glioma cells were plated in 100-mm-diameter tissue culture dishes at 5 × 105 cells/plate in medium supplemented with 10% FBS. The following day medium was removed and replaced with medium containing 0.1% FBS. Approximately 1 hour later, siRNA:lipid complexes were added to the cells and allowed to incubate for 48 hours before cells were harvested as described above.

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