KRAS siRNA

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	Human epidermoid carcinoma cell line A431 (DSMZ, Braunschweig, Germany) was kept in RPMI 1640 medium. Cell culture media were supplemented with 10% (vol/vol) heat-inactivated fetal calf serum, 100 U/ml penicillin, and 100 µg/ml streptomycin. Cells were seeded at a density of 1 x 105/well in six-well plates. Next day, transfection of small interfering RNA (siRNA) was carried out using Lipofectamine2000 (Invitrogen, Carlsbad, CA). For all procedures, standard protocols were used according to the manufacturers' manuals. Synthetic siRNAs targeting KRAS4b was purchased from Applied Biosystems/Ambion (Carlsbad, CA). Target sequences were as follows: KRAS siRNA 1 (ID s7939) sense 5′-CUAUGGUCCUAGUAGGAAAtt-3′ and antisense 5′-UUUCCUACUAGGACCAUAGgt-3′; KRAS siRNA 2 (ID s7940) sense 5′-GCCUUGACGAUACAGCUAAtt-3′ and antisense 5′-UUAGCUGUAUCGUCAAGGCac-3′; As a control, unspecific siRNA Negative Control 1 (Applied Biosystems/Ambion) was used. Cells were transfected with 25 nM KRAS4b-specific or control siRNA for 72 hours.

Protocol tips

Human epidermoid carcinoma cell line A431 (DSMZ, Braunschweig, Germany) was kept in RPMI 1640 medium. Cell culture media were supplemented with 10% (vol/vol) heat-inactivated fetal calf serum, 100 U/ml penicillin, and 100 µg/ml streptomycin. Cells were seeded at a density of 1 x 105/well in six-well plates. Next day, transfection of small interfering RNA (siRNA) was carried out using Lipofectamine2000 (Invitrogen, Carlsbad, CA). For all procedures, standard protocols were used according to the manufacturers' manuals. Synthetic siRNAs targeting KRAS4b was purchased from Applied Biosystems/Ambion (Carlsbad, CA). Target sequences were as follows: KRAS siRNA 1 (ID s7939) sense 5′-CUAUGGUCCUAGUAGGAAAtt-3′ and antisense 5′-UUUCCUACUAGGACCAUAGgt-3′; KRAS siRNA 2 (ID s7940) sense 5′-GCCUUGACGAUACAGCUAAtt-3′ and antisense 5′-UUAGCUGUAUCGUCAAGGCac-3′; As a control, unspecific siRNA Negative Control 1 (Applied Biosystems/Ambion) was used. Cells were transfected with 25 nM KRAS4b-specific or control siRNA for 72 hours.

Publication protocol

Human epidermoid carcinoma cell line A431 (DSMZ, Braunschweig, Germany) was kept in RPMI 1640 medium. Cell culture media were supplemented with 10% (vol/vol) heat-inactivated fetal calf serum, 100 U/ml penicillin, and 100 µg/ml streptomycin. Cells were seeded at a density of 1 x 105/well in six-well plates. Next day, transfection of small interfering RNA (siRNA) was carried out using Lipofectamine2000 (Invitrogen, Carlsbad, CA). For all procedures, standard protocols were used according to the manufacturers' manuals. Synthetic siRNAs targeting KRAS4b was purchased from Applied Biosystems/Ambion (Carlsbad, CA). Target sequences were as follows: KRAS siRNA 1 (ID s7939) sense 5′-CUAUGGUCCUAGUAGGAAAtt-3′ and antisense 5′-UUUCCUACUAGGACCAUAGgt-3′; KRAS siRNA 2 (ID s7940) sense 5′-GCCUUGACGAUACAGCUAAtt-3′ and antisense 5′-UUAGCUGUAUCGUCAAGGCac-3′; As a control, unspecific siRNA Negative Control 1 (Applied Biosystems/Ambion) was used. Cells were transfected with 25 nM KRAS4b-specific or control siRNA for 72 hours.

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