The cells were transfected with mirVana™ hsa‐miR‐218‐5p mimic (Invitrogen) in a final concentration of 10 nM and 50 nM at 70%–80% confluency. Lipofectamine™ 2000 (Invitrogen) was used as a transfection agent according to the manufacturer’s protocols. A non‐specific scrambled siRNA (Sigma‐Aldrich) was used as a control in all transfection experiments. The cells were processed after 24 h for the quantification of transcript levels using qPCR, Western blot, cellular assays or AFM analysis.
Non specific scrambled sirna
Manufactured by Merck Group
Non‐specific scrambled siRNA is a laboratory reagent used as a control in RNA interference (RNAi) experiments. It serves as a negative control to help determine the specificity of gene silencing effects observed with target-specific siRNAs. The non‐specific scrambled siRNA does not target any known gene and is used to assess off‐target effects.
Automatically generated - may contain errors
3 protocols using non specific scrambled sirna
1
Transfection of miR-218-5p Mimic in Cells
2
Modulating Apoptosis Pathways in T24 Cells
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The T24 human BC cell line was chosen for the knockdown of mRNA encoding PKM2 (143814), Bcl-2 (214532), and PUMA (134322) using specific siRNAs from Thermo Fisher Scientific. Non-specific, scrambled siRNA from Sigma was used as a negative control. The pCEP4-HA-Bax (#16587) plasmid was purchase from Addgene. The control plasmid pCEP4 (V04450) was purchase from Thermofisher Scientific. The transfection was conducted by lipofetamin 2000 according to the manufacturer's instruction.
3
Silencing Autophagy Genes in Osteosarcoma
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Small interfering RNAs (siRNAs) against ATG5 (sense: 5′-GUGAGAUAUGGUUUGAAUAdTdT-3′ and antisense: 3′-UAUUCAAACCAUAUCUCACdTdT-5′), Beclin 1 (sense: 5′-GUUUGGAGAUCUUAGAGCAdTdT-3′ and antisense: 3′-UGCUCUAAGAUCUCCAAACdTdT-5′) and a nonspecific scrambled siRNA were purchased from Sigma-Aldrich (Merck KGaA). HOS and U2OS cells (5 × 105) were seeded in six-well plates and transfected with Lipofectamine 3000 (Invitrogen, Rockville, MD, USA) mixed with a serum-free medium containing ATG5 siRNA, Beclin 1 siRNA, and scrambled siRNA. Transfection was performed under 5%CO2 at 37 °C for 24 h.
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