HO-1 small interfering RNA (siRNA) (Cat# EHU051241, Sigma), AMPK siRNA (AMPKα1, Cat# EHU074041; AMPKα2, Cat# EHU042081, Sigma), and control siRNA (Cat# EHUEGFP, Sigma) were purchased from Sigma-Aldrich. control siRNA (Cat# SIC001, Sigma) was used as a negative control. For siRNA transfection, SH-SY5Y cells were cultured in 6-well plates with antibiotic-free medium the day before transfection. The transfection was conducted when cells reached 50–70% confluency using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) and serum-free medium according to the manufacturer’s instructions. SH-SY5Y cells were transfected with 100 pmol siRNA. After 5 hours, the Dulbecco’s modified Eagle’s medium was replaced with culture medium containing 10% fetal bovine serum, and the cells were then incubated at 37°C.
Control sirna
Manufactured by Merck Group
Sourced in United States, France
Control siRNA is a laboratory tool used to establish baseline measurements in RNA interference (RNAi) experiments. It serves as a reference point to evaluate the effects of specific target siRNAs. Control siRNA does not target any known gene and is designed to have minimal impact on cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (14 mentions)
Lipofectamine rnaimax, by Thermo Fisher Scientific (12 mentions)
Lipofectamine rnaimax reagent, by Thermo Fisher Scientific (5 mentions)
Sirnas, by Merck Group (5 mentions)
Opti mem, by Thermo Fisher Scientific (4 mentions)
Rnaimax, by Thermo Fisher Scientific (3 mentions)
Lipofectamine rnaimax transfection reagent, by Thermo Fisher Scientific (3 mentions)
Rnaimax reagent, by Thermo Fisher Scientific (2 mentions)
Neon transfection system, by Thermo Fisher Scientific (2 mentions)
Mission sirna, by Merck Group (2 mentions)
β actin, by Merck Group (2 mentions)
Lipofectamine 2000 sirna transfection reagent, by Thermo Fisher Scientific (2 mentions)
Smgm 2 medium, by Lonza (2 mentions)
Adamts9 sirna sasi hs02 00371041, by Merck Group (2 mentions)
Vcan sirna silencer select s229335, by Thermo Fisher Scientific (2 mentions)
Cc 4149, by Lonza (2 mentions)
Human usp13 sirna, by Merck Group (2 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions)
Hiperfect transfection reagent, by Qiagen (2 mentions)
Gli1 sirnas, by Merck Group (2 mentions)
99 protocols using control sirna
1
Silencing HO-1 and AMPK Pathways in SH-SY5Y Cells
2
CD26 Knockdown in Oocytes
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The siRNA was designed and commercially synthesized by Sigma-Aldrich, with duplex sequences as follows: control siRNA; 5′-ACUUCGACACAUCGACUGC[dT][dT]-3′ and cd26 siRNA; 5′-UUAAGUAAUCAGUUAGAGUGU-3′. The siRNA at three concentrations (5, 10, and 20 µg) were assessed, with each experiment replicated five times. To knockdown cd26, approximately 10 pL of siRNA prepared in RNase-free H2O was micro-injected into the cytoplasm, using an inverted Nikon Diaphot microscope (200× magnification) and an electric microinjector (IM-400, Narishige, Tokyo, Japan). Non-silencing siRNA (Sigma-Aldrich) was injected into another group of oocytes (control siRNA), and a third group received a sham injection without siRNA (uninjected).
3
Cell Line Maintenance and RNAi Knockdown
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Cell lines used were obtained from the American Type Culture Collection (ATCC). 293T, HeLa, U2OS and MCF-7 cells were maintained in DMEM supplemented with 10% FBS in a humidified incubator equilibrated with 5% CO 2 at 37°C. MDA-MB-231 cells were cultured in L-15 medium supplemented with 10% FBS without CO 2 . Transfections were carried out using Polyethyienimine (Polysciences) or Lipofectamine RNAiMAX Reagent (Invitrogen) according to the manufacturer's instructions. Each experiment was performed in triplicate and repeated at least three times. For RNAi experiment, at least three independent siRNA/shRNA sequences were tested for each gene, and the one with the best efficiency was used. The sequences of siRNA were: control siRNA, 5′-UUCUCCGAACGUGUCACGU-3′; ZNF704 siRNA-1, 5′-CAAUGGUACUAACCAGCUUGU -3′; ZNF704 siRNA-2, 5′-CCCUUUGGUUCGAAGUCCU-3′; control siRNA and siRNAs for ZNF704 were synthesized by Sigma-Aldrich. The siRNA oligonucleotides were transfected into cells using RNAiMAX with a final concentration of 20 nM.
4
Cell Culture and Transfection Protocols for hnRNPM Knockdown
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Human TC71, LAP35 and SK-N-MC cells were maintained in Iscove's modified Dulbecco's medium (IMDM, GIBCO), supplemented with 10% fetal bovine serum. Transfection of TC71 cell line was performed using RNAimax reagent (Invitrogen) according to manufacter's instructions. TC71 cell line was trasfected with control siRNA (Sigma Aldrich) and siRNA for hnRNPM (SantaCruz Biotechnology) at the final concentration of 30 nM. BEZ235 was purchased from EMD Chemical Inc./Calbiochem. Inhibitors were dissolved in dimethyl sulfoxide and aliquots were stored at -20°C. Stock solutions were diluted to the final concentrations in growth medium just before use. At the end of the incubation, cells were washed twice with ice-cold phosphate buffered saline (PBS), resuspended in RIPA lysis buffer [150 mM NaCl, 50 mM Tris–HCl pH 7.5, 2 mM EDTA, 0,1% in sodium dodecyl sulfate (SDS), 0,5% sodium deoxycolate, 1 mM dithiothreitol, 0,5 mM Na-ortovanadate, 10 mM B-glycerolphosphate, 10 mM sodium fluoride, 1% NP-40 and Protease-Inhibitor Cocktail (Sigma-Aldrich)] and kept on ice for 10 min. Soluble protein extracts were separated by centrifugation at 12 000 rpm for 10 min and diluted in sodium dodecyl sulfate (SDS) sample buffer.
5
Lipofectamine-mediated Transfection and Silencing in Adipocytes
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For expression assays, 3T3‐L1 cells, human adipocytes, and HEK-293 AD cells, were transfected with the corresponding plasmid vectors at 2.5 µg/mL using a 7.5:1000 dilution of Lipofectamine 2000 (Invitrogen), and cultured for 48 h prior to the experiments. For silencing studies, cells were transfected with a 7.5:1000 dilution of Lipofetamine RNAiMAX (Invitrogen) and mouse Rab34 siRNA (Dharmacon), mouse UBA1 siRNA (Dharmacon), or control siRNA (Sigma-Aldrich) (scrambled-transfected cells) at 25 nmol/L. Then, cells were kept in culture for 72 h. At the end of the experiments, cells were processed for confocal microscopy and/or immunoblotting as indicated in the corresponding sections. In another set of experiments, cells were collected in radioimmunoprecipitation assay (RIPA) buffer and intracellular concentration of TGs was determined using Triglyceride Reagent (Sigma-Aldrich) and Amplex UltraRed Reagent (Invitrogen), while culture media were analyzed for free glycerol content using Amplex UltraRed Reagent (Invitrogen) and Free Glycerol Reagent (Sigma-Aldrich) as previously described [24 (link)].
6
Fibroblast Isolation and CFIm25 Knockdown
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Primary human skin fibroblasts were isolated using an outgrowth model from skin punch from normal donor and SSc patients. Isolated fibroblasts were treated in DMEM (Sigma-Aldrich) containing 10% fetal bovine serum and 1% antibiotics to avoid contamination, and mycoplasma infection was tested using the MycoAlert Mycoplasma Detection Kit (Lonza). Cell culture was maintained at 37°C in a humidified 5% carbon dioxide atmosphere.
To knock down CFIm25, fibroblasts were transfected with 50 ng/ml CFIm25 or control siRNA (Sigma-Aldrich) using Lipofectamine RNAiMAX (ThermoFisher Scientific) on day 0 and day 1, and RNA and protein were collected on day 4 for analysis.
To knock down CFIm25, fibroblasts were transfected with 50 ng/ml CFIm25 or control siRNA (Sigma-Aldrich) using Lipofectamine RNAiMAX (ThermoFisher Scientific) on day 0 and day 1, and RNA and protein were collected on day 4 for analysis.
7
Silencing Snail in DU145 Cells
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DU145-shTRIM16#-transfected cells were transiently transfected with either 20 nM control siRNA (Sigma) or siRNA specific to Snail (Sigma) using Lipofectamine 2000 (Thermo Fisher Scientific) as the transfection agent. The sequence for Snail siRNA was TTGTACCTCAAAGAAGGTGGC.
8
siRNA-mediated Knockdown in Cell Coculture
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siRNA-mediated knockdown of Cx43 and Dicer was performed by electroporation with Amaxa nucelofector II (Lonza) according to the manufacturer’s instructions. Cells were harvested and resuspended in nucleofection buffer containing 90 mM Na2PO4, 90 mM NaH2PO4, 5 mM KCl, 10 mM MgCl2 and 10 mM sodium succinat. 250 nM Cx43, Dicer siRNA or control siRNA (Sigma Aldrich) were transfected using nucleofector program U-23 for MSCs or G-009 for CMs. If specified, MSCs were transfected with 3 µg EGFP Plasmid. To study miRNA exchange between CMs and MSCs, CMs were transfected with 250 nM DY-547 labeled control miRNA (Dharmacon).
9
Modulation of Prostate Cancer Regulators
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Human SREBP-2 cDNA was subcloned into p3XFLAG-myc-CMV-26 (Sigma-Aldrich) at NotI/XbaI restriction enzyme sites to generate p3XFLAG-SREBP-2 expression construct. LNCaP and LAPC4 cells were transfected with either p3XFLAG-SREBP-2 or empty vector as a control using Lipofectamine LTX Plus reagent (Life Technologies). For the SREBP-2 shRNA-mediated knockdown study, non-targeting control (pLKO.1, empty vector) or SREBP-2 shRNA lentiviral particles (Sigma-Aldrich) were used to infect CWR22Rv1 or C4-2B cells. In order to generate c-Myc-overexpressing PCa cell clones, retroviruses that carried either pWZL-Blast-Myc (Addgene) or control vector (pWZL-Blast) were used. For the silencing of c-Myc, cells were transfected with c-Myc siRNA-1, c-Myc siRNA-2 or control siRNA (Sigma-Aldrich) using DharmaFECT 2 transfection reagent (Fisher Scientific Dharmacon).
10
Knockdown of NPM1/B23 in T-cell lines
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Control siRNA (#SIC001, Sigma) and siRNAs targeting NPM1/B23 (siRNA21 (SI049500010): CACCAGT GGTCTTAAG GTTGA and siRNA22 (SI04949994): AAGGACAAG AATCCT TCAAGA) were ordered from Qiagen and transfected into MT-2 and C8166-45 cells with Neon Transfection System (Invitrogen), according to manufacturer’s instructions. After 24 h, cells were harvested for further analyses.
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