Scrambled scr sirna, by Horizon Discovery - Product details

1

Gene Silencing by siRNA Transfection

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Cells were transfected with siRNA by nucleofection using the Amaxa Cell-line Nucleofector Kit V (Cat# VCA-1003; Lonza AG). Controls were Scrambled siRNA (SCR) (#D-001960-01-05, Dharmacon, Lafayette, CO, USA) or EGFP oligonucleotides (NS; 5′-CAAGCTGACCCTGAAGTTC-3′) [68 (link)]. Transfected cells were re-seeded 48 h post-transfection and allowed to attach overnight before starting treatments.
siRNA sequences for CEBPD were as follows:
CEBPD siRNA-1-sense 5′-rUrCrGrCrCrGrArCrCrUrCrUrUrCrArArCrArGTT-3′
CEBPD siRNA-1-antisense 5′-rCrUrGrUrUrGrArArGrArGrGrUrCrGrGrCrGrATT-3′
CEBPD siRNA2-sense 5′-rCrCrArCrUrArArArCrUrGrCrGrArGrArGrArATT-3′
CEBPD siRNA2-antisense 5′-rCrUrGrUrUrGrArArGrArGrGrUrCrGrGrCrGrATT-3′
siRNA pools for ATF6A/ATF6α (sc-37699), ERN1/IRE1α (sc-40705), EIF2AK3/PERK (sc-36213), NRF2 (sc-37030) and STAT3 (sc-29493) were purchased from Santa Cruz Biotechnology.

Sheshadri N., Poria D.K., Sharan S., Hu Y., Yan C., Koparde V.N., Balamurugan K, & Sterneck E. (2021). PERK signaling through C/EBPδ contributes to ER stress-induced expression of immunomodulatory and tumor promoting chemokines by cancer cells. Cell Death & Disease, 12(11), 1038.

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2

Optimize siRNA Knockdown in Ramos B Cells

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X 10⁶ Ramos B cells were suspended in 300 μL Amaxa buffer SG (246 μL SG buffer + 54 μL Supplement 1, Lonza: #V4XC-3024). 100 μL of resuspended cells was added to three separate Amaxa cuvettes. 0.5 μL of either ddH₂O (Mock), 100 μM Scrambled siRNA (Scr) (GE Dharmacon Catalog#: D-001810-10-05), or 100 μM IRF5 targeting siRNA (IRF5KD) (GE Dharmacon Catalog#: L-011706-00-0010) was added to each cuvette. Cells were then nucleofected on the Amaxa 4D Nucleofector using program CA-137. After nucleofection, cells were immediately added to 1 mL of RPMI-1640 (+10% FBS, 1x Glutamine, 1X Non-essential Amino Acids), cultured for 24 h, and then pelleted and re-nucleofected with siRNA as before. Cells were then lysed 24 h after the second nucleofection for immunoblot analysis.

Li D., De S., Li D., Song S., Matta B, & Barnes B.J. (2016). Specific detection of interferon regulatory factor 5 (IRF5): A case of antibody inequality. Scientific Reports, 6, 31002.

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3

Atg5 Knockdown in RAW Cells

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Scrambled siRNA (scr) and siRNA against mouse Atg5 were purchased from Dharmacon. Thirty five nM siRNA was used for transfection of RAW cells using lipofectamine 2000 (ThermoFisher Scientific: 52758) using manufacturer’s protocol. At 24 hours post- transfection, cells were removed and plated into 6-well plates. After an additional 24 hours, cells were treated with the IAP or vehicle for 24 hrs followed by addition of LPS (25 ng/ml) for the next 24 hrs. IAP was removed from the medium prior to adding LPS.

Singh S.B., Carroll-Portillo A., Coffman C., Ritz N.L, & Lin H.C. (2020). Intestinal Alkaline Phosphatase Exerts Anti-Inflammatory Effects Against Lipopolysaccharide by Inducing Autophagy. Scientific Reports, 10, 3107.

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4

Pharmacological and Genetic Modulation of PLA2g7 in Pain

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All drugs were prepared in coded syringes on the day of injection by an individual not performing behavioral testing in order to blind the experimenter. Animals were initially acclimated to observation boxes atop thermal or mechanical platforms for a minimum of 30 min prior to collection of baseline recordings. After baselines, animals were administered vehicle or testing compound. Darapladib (Selleckchem; SB-480848) was dissolved in dimethyl sulfoxide (DMSO) and Tween-20 then diluted in phosphate buffered saline (PBS) to 2% of each for final concentrations of 300 nmol and 30 nmol. Control animals received equivalent volume of the Darapladib vehicle.
Knockdown of PLA2g7 was performed using Dharmacon SMARTpool small interfering RNA (siRNA) against four separate PLA2g7 target sequences. Control groups received non-targeting scrambled (scr) siRNA (Dharmacon). The PLA2g7 or scrambled siRNA was injected intrathecally (i.t., 2 µg) for three consecutive days for a cumulative dose of 6 µg. Behavior was tested on 2 days after final injection by blinded observers, followed by collection of DRGs for protein and transcript quantification.

Boyd J.T., LoCoco P.M., Furr A.R., Bendele M.R., Tram M., Li Q., Chang F.M., Colley M.E., Samenuk G.M., Arris D.A., Locke E.E., Bach S.B., Tobon A., Ruparel S.B, & Hargreaves K.M. (2021). Elevated dietary ω-6 polyunsaturated fatty acids induce reversible peripheral nerve dysfunction that exacerbates comorbid pain conditions. Nature metabolism, 3(6), 762-773.

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5

Pharmacological and Genetic Modulation of PLA2g7 in Pain

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All drugs were prepared in coded syringes on the day of injection by an individual not performing behavioral testing in order to blind the experimenter. Animals were initially acclimated to observation boxes atop thermal or mechanical platforms for a minimum of 30 min prior to collection of baseline recordings. After baselines, animals were administered vehicle or testing compound. Darapladib (Selleckchem; SB-480848) was dissolved in dimethyl sulfoxide (DMSO) and Tween-20 then diluted in phosphate buffered saline (PBS) to 2% of each for final concentrations of 300 nmol and 30 nmol. Control animals received equivalent volume of the Darapladib vehicle.
Knockdown of PLA2g7 was performed using Dharmacon SMARTpool small interfering RNA (siRNA) against four separate PLA2g7 target sequences. Control groups received non-targeting scrambled (scr) siRNA (Dharmacon). The PLA2g7 or scrambled siRNA was injected intrathecally (i.t., 2 µg) for three consecutive days for a cumulative dose of 6 µg. Behavior was tested on 2 days after final injection by blinded observers, followed by collection of DRGs for protein and transcript quantification.

Boyd J.T., LoCoco P.M., Furr A.R., Bendele M.R., Tram M., Li Q., Chang F.M., Colley M.E., Samenuk G.M., Arris D.A., Locke E.E., Bach S.B., Tobon A., Ruparel S.B, & Hargreaves K.M. (2021). Elevated dietary ω-6 polyunsaturated fatty acids induce reversible peripheral nerve dysfunction that exacerbates comorbid pain conditions. Nature metabolism, 3(6), 762-773.

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6

Gastric Cancer Cell Lines Transfection Protocol

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Gastric cancer cell lines (SNU1, SNU16, SNU216, SNU620, SNU638, and N87) were purchased from the Korean Cell Line Bank (Seoul). Cells were cultured in RPMI1640 supplemented with 25 mM HEPES, 10% fetal bovine serum (FBS), and 100 μg/ml of penicillin/streptomycin (1 × P/S) in 5%CO₂/95% air at 37°C. Cells were transfected with siRNA using DharmaFECT reagent 1 or 3 (Dharmacon, Lafayette, CO), according to the manufacturer’s instructions. The sequences of siRNA used were as follows: MED30 siRNA (Bioneer, Daejeon, Korea), 5’-CGA GCA ACU UAU UCC AUA U(dTdT)-3’, 5’-GCU GCC AAA UGG UGU CAC U(dTdT)-3’, and 5’-CGA GAA AUU GCU GAA GUA A(dTdT)-3’; scrambled (SCR) siRNA (Dharmacon, Lafayette, CO), 5’- GAU CCG CAA AAG AGC GAA A(dTdT)-3’.

Lee Y.J., Han M.E., Baek S.J., Kim S.Y, & Oh S.O. (2015). MED30 Regulates the Proliferation and Motility of Gastric Cancer Cells. PLoS ONE, 10(6), e0130826.

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7

Knockdown and Overexpression of ZHX1 in Glioblastoma

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Scrambled (SCR) siRNA and ZHX1 siRNA were purchased from Dharmacon (Lafayette, CO, USA) and Bioneer Corporation (Daejeon, Korea). The siRNA sequences used were as follows: ZHX1 siRNA, 5′-CUGACUUUUGAUG GUAGUU(dTdT)-3′, 5′-GAAAGUAAUGCAGGUAGUU (dTdT)-3′, and SCR siRNA, 5′-CAGUUCAUCAUAACU CAGU(dTdT)-3′ and 5′-GAUCCGCAAAAGAGCGAAA (dTdT)-3′. For ZHX1 knockdown, glioblastoma cell lines were seeded at 1500 cells/well in a 96-well plate or 70,000 cells/well in a 6-well plate. Cells were transfected with 100 nM of SCR or ZHX1 siRNA within 1 day of seeding. DharmaFECT 1 (Thermo Scientific, Kafayette, CO, USA) was used for the transfection as directed by the manufacturer. The ZHX1 gene was obtained from Invitrogen and was amplified using ZHX1 primers including NotI and XbaI restriction site. The amplified ZHX1 gene and the 3XFlag CMV-10 vector (Sigma, E4401) were digested by NotI and XbaI restriction enzyme, and then ZHX1 was inserted into the vector. To create a stable cell line, U373MG cells were seeded at 100,000 cells/well in a sixwell plate and transfected with empty vector as control or the vector containing the ZHX1 using ViaFect TM (Promega, Madison, WI, USA), according to the manufacturer's instructions. G418 (30 µg/ml; Roche, 04727894001) was used to choose empty control vector-expressing (Mock) and ZHX1-overexpressing (ZHX1-over) clones.

Kwon R.J., Han M.E., Kim Y.J., Kim Y.H., Kim J.Y., Liu L., Heo W, & Oh S.O. (2017). Roles of zinc-fingers and homeoboxes 1 during the proliferation, migration, and invasion of glioblastoma cells. Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine, 39(3).

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Scrambled scr sirna

Lab products found in correlation

Spelling variants of this product

7 protocols using scrambled scr sirna

Gene Silencing by siRNA Transfection

Optimize siRNA Knockdown in Ramos B Cells

Atg5 Knockdown in RAW Cells

Pharmacological and Genetic Modulation of PLA2g7 in Pain

Pharmacological and Genetic Modulation of PLA2g7 in Pain

Gastric Cancer Cell Lines Transfection Protocol

Knockdown and Overexpression of ZHX1 in Glioblastoma

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