Negative sirna

Manufactured by Qiagen

Sourced in Germany

Negative siRNA is a laboratory tool used to study gene function by temporarily suppressing the expression of target genes. It serves as a control to compare the effects of gene silencing experiments.

Automatically generated - may contain errors

Lab products found in correlation

Pin1 sirna, by Thermo Fisher Scientific (3 mentions) Rnaimax, by Thermo Fisher Scientific (3 mentions) Odyssey imaging system, by LI COR (2 mentions) Lipofectamine rnaimax, by Thermo Fisher Scientific (1 mentions) Pcdna3, by Thermo Fisher Scientific (1 mentions) Lipofectamine 2000 reagent, by Thermo Fisher Scientific (1 mentions) Okadaic acid, by Cell Signaling Technology (1 mentions) Odyssey blocking buffer, by LI COR (1 mentions) α tubulin, by Cell Signaling Technology (1 mentions) Dlk 1, by Abcam (1 mentions) Hyperfect transfection reagent, by Qiagen (1 mentions)

Close spelling variants of this product

AllStars Negative siRNA AF 488 Allstars negative siRNA control AllStars Negative siRNA Negative siRNA control SiRNAs

8 protocols using negative sirna

GATA2 Knockdown in Primary HUVECs

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siRNA targeting GATA2 was administered to primary HUVECs at 10 nM or a matching concentration of negative siRNA (Qiagen, Limburg, Netherlands) using lipofectamine RNAiMAX (Life Technologies). Primary HUVECs were cultured for 48 hours, with treatment of both GATA2 siRNA and negative siRNA occurring after 24 hours of incubation.
Culture media was collected for analysis by ELISA and RNA collected for analysis by RT-PCR. Knockdown efficiency of GATA2 siRNA in primary HUVECs was confirmed by qRT-PCR. (Supplementary Figure 3A). We initially confirmed no negative effect on viability using MTS (data not shown).

Whigham C.A., MacDonald T.M., Walker S.P., Pritchard N., Hannan N.J., Cannon P., Nguyen T.V., Hastie R., Tong S, & Kaitu’u-Lino T.J. (2019). Circulating GATA2 mRNA is decreased among women destined to develop preeclampsia and may be of endothelial origin. Scientific Reports, 9, 235.

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Silencing ERβ and GPR30 in T24 cells

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T24 cells were transfected using siRNA transfection regent (Qiagen, Hilden, Germany) with 10 nM ERβ or GPPR30 siRNA (Qiagen, Hilden, Germany) according to the manufacturer's instructions; negative siRNA (Qiagen, Hilden, Germany) was used as a negative control. The target sequence of the used ERβ siRNA was 5′-CAGCGATTACGCATCGGGATA-3′, and the sequence of used GPR30 siRNA was 5′-CGGCCACGTCATGTCTCTAAA-3′. After culturing in phenol-red-free RPMI 1640 medium containing 10% dextran-coated charcoal-treated FBS for 24 h, E2 was added to 6-well plates for the qPCR and western blot experiments or to 96-well plates for the MTT assays.
Mammalian expression vectors encoding ERβ or GPPR30 were constructed by inserting PCR-amplified fragments into pcDNA3 (Invitrogen). Lipofectamine 2000 reagent was used for transfections according to the standard protocols (Invitrogen).

Huang W., Chen Y., Liu Y., Zhang Q., Yu Z., Mou L., Wu H., Zhao L., Long T., Qin D, & Gui Y. (2015). Roles of ERβ and GPR30 in Proliferative Response of Human Bladder Cancer Cell to Estrogen. BioMed Research International, 2015, 251780.

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Knockdown of Pin1 and ACC1 in Cell Lines

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DU145, LNCap and HEK-293T cells were transfected with either negative siRNA (Qiagen) or Pin1 siRNA (Invitrogen) using RNAiMAX (Invitrogen) according to the manufacturer’s protocol: Pin1 siRNA1, CCG UGU UCA CGG AUU CCG GCA UCC A; Pin1 siRNA2, GCC CUG GAG CUG AUC AAC GGC UAC A; ACC1 siRNA1, GGG ACU UCA UGA AUU UGC UGA UUC U; ACC1 siRNA2, CCU UAC AAG GGA UAC AGG UAU UUA U.

Ueda K., Nakatsu Y., Yamamotoya T., Ono H., Inoue Y., Inoue M.K., Mizuno Y., Matsunaga Y., Kushiyama A., Sakoda H., Fujishiro M., Takahashi S.I., Matsubara A, & Asano T. (2019). Prolyl isomerase Pin1 binds to and stabilizes acetyl CoA carboxylase 1 protein, thereby supporting cancer cell proliferation. Oncotarget, 10(17), 1637-1648.

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Measuring FGF-2-Induced ERK Signaling

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HUVECs seeded in 6-well plates were incubated with nanocomplexes at 50:1 containing 1.0µg per well of negative siRNA (Allstars, Qiagen) or miR-126–3p for 24 hours. HUVECs were completely serum starved with EBM-2 basal medium for 16 hours and then treated with fibroblast growth factor (FGF-2, 10 ng/ml) for 15 or 45 minutes. HUVECs were then placed on ice, washed in ice cold PBS and lysed with RIPA Buffer (50mM Tris-HCl pH 8.0, 150 mM NaCl, 1% Nonidet P-40 (NP-40), 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulphate (SDS), 1 mM sodium orthovanadate, 1 mM NaF) supplemented with a protease/phosphatase inhibitor and okadaic acid (20nM; Cell Signaling). Protein lysates were resolved in a 4–12% Bis-Tris Protein Gel at 100–200V for 1.5 hours. Proteins were transferred to a PVDF membrane at 100V for 1 hour and blocked using Odyssey Blocking Buffer (Li-Cor) for 1 hour. Blots were probed with phosphoERK 42/44 (1:1000), total ERK 42/44 (1:1000) overnight at 4C. Blots were analyzed using an Odyssey Licor Imaging System (Li-COR).

Jones Buie J.N., Zhou Y., Goodwin A.J., Cook J.A., Vournakis J., Demcheva M., Broome A.M., Dixit S., Halushka P.V, & Fan H. (2019). Application of deacetylated-poly-N-acetyl glucosamine nanoparticles for the delivery of miR-126 for the treatment of cecal-ligation puncture induced sepsis. Inflammation, 42(1), 170-184.

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DLK-1 Expression Modulation in HUVECs

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For DLK-1 experiments, HUVECs seeded in 6-well plates were incubated with nanocomplexes at a 50:1 ratio containing 1.0µg of negative siRNA (Allstars, Qiagen) or miR-126–5p for 48 hours. The cellular proteins were extracted and lysates were processed as described above. Blots were probed for α-tubulin (1:1000; Cell Signaling), or DLK-1 (1:1000; Abcam) overnight at 4°C and IRDye secondary antibodies (1:10000) for 1 hour. Blots were analyzed using an Odyssey Licor Imaging System (Li-COR).

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SARS-CoV-2 Infection of VeroE6/TMPRSS2 Cells

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Yamamotoya T., Nakatsu Y., Kanna M., Hasei S., Ohata Y., Encinas J., Ito H., Okabe T., Asano T, & Sakaguchi T. (2021). Prolyl isomerase Pin1 plays an essential role in SARS-CoV-2 proliferation, indicating its possibility as a novel therapeutic target. Scientific Reports, 11, 18581.

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SARS-CoV-2 Infection in VeroE6/TMPRSS2 Cells

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VeroE6/TMPRSS2 cells (African green monkey kidney-derived cells expressing human TMPRSS2, purchased from the Japanese Collection of Research Bioresources (JCRB) Cell Bank, JCRB1819) were maintained in Dulbecco's Modi ed Eagle's Medium (DMEM) containing 10% fetal bovine serum (FBS) and 1 mg/mL G418 at 37°C in 5% CO 2 . For siRNA treatment, VeroE6/TMPRSS2 cells were transfected with either negative siRNA (QIAGEN) or Pin1 siRNA (Invitrogen) using RNAiMAX (Invitrogen) according to the manufacturer's protocol and subjected to SARS-CoV-2 infection 3 days later. Pin1 siRNA1: CCG UGU UCA CGG AUU CCG GCA UCC A. Pin1 siRNA2: GCC CUG GAG CUG AUC AAC GGC UAC A. Pin1 inhibitors Chemical structures of Pin1 inhibitors termed H-77, H-175, H-363, H-371 and H-593 are shown in Table 1. These Pin1 inhibitors inhibit isomerase activity by more than 80% at a concentration of 20 µM, based on an in vitro assay using recombinant Pin1 protein. However, it should be noted that the results of such an in vitro assay usually differ signi cantly from the results obtained by in vivo experiments. The compounds were solubilized in DMSO. Before the infection experiments, the culture medium of VeroE6/TMPRSS2 cells was changed to DMEM without FBS and G-418, and virus and/or Pin1 inhibitors were added at the indicated titer or concentrations.

Yamamotoya T., Nakatsu Y., Hasei S., Ohata Y., Encinas J., Ito H., Okabe T., Asano T., & Sakaguchi T. (2021). Prolyl isomerase Pin1 plays an essential role in SARS-CoV-2 proliferation, indicating its possibility as a novel therapeutic target.

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Endophilin A2 Gene Silencing

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The siRNA (small interfering RNA) duplexes against the rat endophilin A2 gene (Gene ID: 81922) was designed and constructed by QIAGEN and transfected with Hyperfect Transfection Reagent, according to the manufacturer's instructions (QIAGEN). The working concentration of endophilin A2 siRNA was 40 nmol/L. A negative siRNA (QIAGEN) was used as a negative control. The cells were then used for western blot and patch-clamp studies after 48 h.

Liu C.Z., Li X.Y., Du R.H., Gao M., Ma M.M., Li F.Y., Huang E.W., Sun H.S., Wang G.L, & Guan Y.Y. (2016). Endophilin A2 Influences Volume-Regulated Chloride Current by Mediating ClC-3 Trafficking in Vascular Smooth Muscle Cells. Circulation journal : official journal of the Japanese Circulation Society, 80(11).

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