Sirnas

Manufactured by Merck Group

Sourced in United States, France

SiRNAs (small interfering RNAs) are a class of double-stranded RNA molecules that play a central role in the RNA interference (RNAi) pathway. They are used in biological research to selectively silence the expression of target genes by binding to and degrading the corresponding mRNA transcripts. SiRNAs can be used to investigate gene function and validate drug targets in cellular and animal models.

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Lab products found in correlation

112 protocols using sirnas

Antibody and siRNA Details Protocol

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Detailed information about all antibodies used in this study is shown in table S4. All small interfering RNAs (siRNAs) were synthesized from Sigma-Aldrich, and the target sequences of the siRNAs are shown in table S5.

Li L., Zhou A., Wei Y., Liu F., Li P., Fang R., Ma L., Zhang S., Wang L., Liu J., Richard H.T., Chen Y., Wang H, & Huang S. (2022). Critical role of lncEPAT in coupling dysregulated EGFR pathway and histone H2A deubiquitination during glioblastoma tumorigenesis. Science Advances, 8(40), eabn2571.

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Silencing Gene Expression in Breast Cancer Cells

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ZR-75-1, MCF-7, SK-BR-3, MDA-MB-231, MDA-MB-468, MDA-MB-436, HCC1806, HCC1937 cells were grown in RPMI 1640 (Lonza), SUM159 were grown in DMEM/F12(Sigma Aldrich), all supplemented with 10% FBS, gentamycin, penicillin and streptomycin. For RNA interference, cells were transfected with siRNAs (Sigma-Aldrich) using Lipofectamine RNAiMAX (Invitrogen) according to manufacturer’s instructions and harvested 48 h later for protein and RNA analyses. For RNA stability assay, 48 h after silencing, cells were treated with either Actinomycin D (3 μg/ml) or DMSO and collected for RNA analyses at indicated time points. Sequences of siRNAs are listed in Additional File 1: Supplemental Table 1.

Naro C., De Musso M., Delle Monache F., Panzeri V., de la Grange P, & Sette C. (2021). The oncogenic kinase NEK2 regulates an RBFOX2-dependent pro-mesenchymal splicing program in triple-negative breast cancer cells. Journal of Experimental & Clinical Cancer Research : CR, 40, 397.

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Cell Culture and Gene Silencing Protocols

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v-Src-transformed MEFs and GFP-actin v-Src-transformed MEFs were grown in high-glucose DMEM (GIBCO) with 10% fetal bovine serum (GIBCO), 100 μM non-essential amino acids, 1 mM sodium pyruvate, and 55 μM 2-mercaptoethanol and cultured at 37 °C in a humidified atmosphere of 5% CO2 and 95% air. GFP-actin v-Src-transformed MEFs were selected using Zeocin (Invitrogen). Osteosarcoma cell line U2OS was grown in low-glucose DMEM (GIBCO) with 10% fetal bovine serum (GIBCO). Breast cancer MDA-MB-231 cells were cultured in RPMI-1640 (GIBCO) with 10% fetal bovine serum (GIBCO). Two independent pairs of different siRNAs (Sigma) and a siRNA pool of three duplexes (Santa Cruz Biotechnology) targeting STIM1 or Orai1 were used in this study.

Chen Y.W., Lai C.S., Chen Y.F., Chiu W.T., Chen H.C, & Shen M.R. (2017). STIM1-dependent Ca2+ signaling regulates podosome formation to facilitate cancer cell invasion. Scientific Reports, 7, 11523.

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Silencing mTOR Pathway Regulators

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Dharmafect 2 transfection reagent (Thermo Scientific, Rockford, IL) and small interference RNAs (siRNAs) (Sigma-Aldrich, St. Louis, MO), targeting raptor (100 nM; sense, 5′CAGUUCACCGCCAUCUACA), rictor (100nM; sense, 5′ CGAUCAUGGGCAGGUAUUA), DEPTOR (SASI_1297010-H/5582, 1297011-H) or Nedd4-2 (100 nM; 5′ CCCUAUACAUUUAAGGACU) were used. Control cells were transfected with a non-coding scrambled sequence (100nM; sense: 5′GAUCAUACGUGCGAUCAGATT). siRNAs were added to cultured PHT cells (~3.75 × 10⁶ cells/well in 6 well plate; ~ 7.5 × 10⁶ cells in 60 mm dish) after 18 h in culture, incubated for 24 h [39 (link)] and subsequently removed and replaced by fresh medium. At 90 hours in culture, efficiency of target silencing was determined at the protein and functional levels using Western blot.

Rosario F.J., Dimasuay K.G., Kanai Y., Powell T.L, & Jansson T. (2015). Regulation of Amino Acid Transporter Trafficking by mTORC1 in Primary Human Trophoblast cells is Mediated by the Ubiquitin Ligase Nedd4-2. Clinical science (London, England : 1979), 130(7), 499-512.

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MAGED2 Gene Silencing Protocol

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siRNAs targeting the sequences GGGAUACAUCAUUCACUCU (MAGED2-1) or GGGCAAAUGAUUUGGUGAA (MAGED2-2) of human MAGED2 and non-silencing control siRNA were purchased from Sigma-Aldrich. Cells were transfected with siRNAs using Lipofectamine RNAiMAX Reagent (Invitrogen).

Strekalova E., Malin D., Good D.M, & Cryns V.L. (2015). Methionine Deprivation Induces a Targetable Vulnerability in Triple-negative Breast Cancer Cells by Enhancing TRAIL Receptor-2 Expression. Clinical cancer research : an official journal of the American Association for Cancer Research, 21(12), 2780-2791.

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Transfection and Knockdown of Ddx3x in HEK293T Cells

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HEK293T cells were grown in DMEM (Invitrogen) supplemented with 2 mM L-glutamine and 10% heat-inactivated fetal calf serum at 37 °C in a humidified atmosphere containing 5% CO₂. Cells were transfected with 200 ng of individual plasmids using the Nanofectin siRNA Kit, according to the manufacturer's instructions (PAA, Austria). For human Ddx3x knockdown, 20 nM of chemically synthesized (Sigma) siRNAs (I-s-5′-GAUUCACUGACCUUAGUGUdTdT-3′; I-as-5′-ACACUAAGGUCAGUGAAUCdTdT-3′; II-s-5′-GAAACAGUCGCUGGUGUGAdTdT-3′; II-as-5′-UCACACCAGCGACUGUUUCdTdT-3′) were used. In all experiments, siDdx3x-I and siDdx3x-II were co-transfected together.

Krol J., Krol I., Alvarez C.P., Fiscella M., Hierlemann A., Roska B, & Filipowicz W. (2015). A network comprising short and long noncoding RNAs and RNA helicase controls mouse retina architecture. Nature Communications, 6, 7305.

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Knocking Down Clathrin in HeLa Cells

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HeLa cells were grown on glass coverslips in RPMI 1640 media (ThermoFisher Scientific) supplemented with 10% foetal bovine serum (ThermoFisher Scientific) and 1% penicillin-streptomycin (ThermoFisher Scientific) at 37 °C in 5% CO₂. Cells were transfected with 90 nM siRNA duplexes the day after seeding using Lipofectamine 2000 (ThermoFisher Scientific) according to the manufacturer’s instructions. Fresh media was added after 6–8 h and every 12–15 h thereafter, until cells were fixed. siRNAs were purchased from SigmaAldrich and had the following sequences: luciferase, 5′-CGUACGCGGAAUACUUCGAdTdT-3′ (sense), 5′-UCGAAGUAUUCCGCGUACGdTdT-3′ (antisense) and CHC, 5′-GCAAUGAGCUGUUUGAAGA-3′, 5′-UCUUCAAACAGCUCAUUGC-3′ (antisense).

Khushi M., Dean I.M., Teber E.T., Chircop M., Arthur J.W, & Flores-Rodriguez N. (2017). Automated classification and characterization of the mitotic spindle following knockdown of a mitosis-related protein. BMC Bioinformatics, 18(Suppl 16), 566.

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Silencing cFLIP Isoforms in HCT116 Cells

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siRNAs against cFLIP_L, cFLIP_S, cFLIP_L/S, and non-targeting scrambled control oligonucleotides, were synthesized by Sigma (St. Louis, MO, USA). Cell suspensions of HCT116 cells were transfected with siRNAs using jetPRIME (Polyplus, Illkirch, France) as described by the manufacturer. After 24 h, the transfection medium was replaced with a regular medium, and cells were further incubated for 24 h before treatments.
siRNAs:

cFLIP_{L/S (DUAL)};cFLIP_L;cFLIP_S	5′-GGGACCUUCUGGAUAUUUU[dt][dt]-3′; 5′-CCUAGGAAUCUGCCUGAUA[dt][dt]-3′; 5′-CACCCUAUGCCCAUUGUCC[dt][dt]-3′
Scrambled control	5′-CUUUGGGUGAUCUACGUUA[dt][dt]-3′

Mora-Molina R., Stöhr D., Rehm M, & López-Rivas A. (2022). cFLIP downregulation is an early event required for endoplasmic reticulum stress-induced apoptosis in tumor cells. Cell Death & Disease, 13(2), 111.

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BLIMP1 Silencing in MIA PaCa-2 Cells

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siRNAs (Sigma-Aldrich) were used for BLIMP 1 silencing. Non-targeting (si-Ctrl: SIC001) or BLIMP 1-targeting siRNAs (si-Blimp 1-1: 5′CUUGGAAGAUCUGACCCGA-3′; si-Blimp 1-2: 5′CCUUUCAAAUGUCAGACUU-3′ were transfected in MIA PaCa-2 cells using Lipofectamine RNAiMAX (Invitrogen, Life technologies Corp.) following the manufacturer’s instructions. The final siRNA concentration was 30 nM. The medium was changed 8 h after transfection and the efficiency of the transfection was assessed by Western blot after 72 h.

Pasquier C., Guerlais V., Pallez D., Rapetti-Mauss R, & Soriani O. (2023). A network embedding approach to identify active modules in biological interaction networks. Life Science Alliance, 6(9), e202201550.

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Targeted AMPK knockdown protocol

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The knockdown of AMPKα expression was conducted using specific small inhibitory RNAs (siRNAs) acquired from Sigma-Aldrich, as previously described [43 (link)]. Briefly, cells were transfected with 0.1 μM siRNA against AMPKα1 (5′-AGU GAA GGU UGG CAA ACA U-3′) and AMPKα2 (5′-GGA AGG UAG UGA AUG CAU A-3′) using Lipofectamine RNAiMax (Invitrogen, Carlsbad, CA, USA) at 37 °C and 5% CO₂ for 72 h.

Huang C.W., Lin Y.C., Hung C.H., Chen H.M., Lin J.T., Wang C.J, & Kao S.H. (2021). Adenine Inhibits the Invasive Potential of DLD-1 Human Colorectal Cancer Cell via the AMPK/FAK Axis. Pharmaceuticals, 14(9), 860.

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