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Sirna duplex

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The SiRNA duplex is a laboratory tool used for gene silencing experiments. It functions by targeting and binding to specific mRNA sequences, preventing their translation into proteins. This technology allows for the study of gene function and the exploration of potential therapeutic targets.

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Lab products found in correlation

Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions) Opti mem medium, by Thermo Fisher Scientific (2 mentions) Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions) Fluorescein conjugate a sirna, by Santa Cruz Biotechnology (1 mentions) Fetal bovine serum (fbs), by Corning (1 mentions) Lipo2000, by Thermo Fisher Scientific (1 mentions) Sirna, by Sangon (1 mentions)

7 protocols using sirna duplex

1

NM23-H2 Knockdown in HeLa Cells

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A commercially available synthetic siRNA duplex (SantaCruz: sc 40774) against NM23-H2 was used to knock down NM23-H2 in HeLa cells. An siRNA duplex (SantaCruz: sc 37007) with a scrambled sequence was used as a negative control. Transfection of siRNA duplexes was accomplished using Lipo2000 (Invitrogen) based on the manufacturer's protocol. Cells were harvested for various experiments 72 h after transfection with the siRNA.
Shan C., Lin J., Hou J.Q., Liu H.Y., Chen S.B., Chen A.C., Ou T.M., Tan J.H., Li D., Gu L.Q, & Huang Z.S. (2015). Chemical intervention of the NM23-H2 transcriptional programme on c-MYC via a novel small molecule. Nucleic Acids Research, 43(14), 6677-6691.
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2

Knockdown of HSF1, NRF2 and SQSTM1 in PEL Cells

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HSF1, NRF2 and SQSTM1 knockdown was performed in PEL cell lines using specific small interfering RNA. Subsequently, 3 × 105 cells were seeded in 12-wells culture plate in RPMI medium supplemented with 10% fetal bovine serum (FBS) (Corning, NY, USA; 35-079), with L-glutamine and without antibiotics. Subsequently, 30 pmoli of siRNA duplex (siRNAHSF1, siRNANRF2 and siRNASQSTM1) (Santa Cruz Biotechnology Inc., Dallas, TX, USA; sc-35611, sc-37030 and sc-29679) and 7 µL of Lipofectamine 2000 Transfection Reagent (Life Technologies, 11668027) were diluted in Optimem medium (Thermo Fisher Scientific, Waltham, MA, USA; 31985062) for 20 min at room temperature. The mixture was added to the cell culture for 48 h. After this, the cells were treated either with TPA (20 ng/mL) (T) and sodium butyrate (0.3 mM), or with bortezomib (20 nM) for the last 24 h.
Transfection efficiency was evaluated by Fluorescein Conjugate-A siRNA (Santa Cruz Biotechnology Inc., Dallas, TX, USA; sc-36869). A control siRNA-A (siRNASC) (Santa Cruz Biotechnology Inc., Dallas, TX, USA; sc-37007) was also used [21 (link)].
Granato M., Gilardini Montani M.S., Angiolillo C., D’Orazi G., Faggioni A, & Cirone M. (2018). Cytotoxic Drugs Activate KSHV Lytic Cycle in Latently Infected PEL Cells by Inducing a Moderate ROS Increase Controlled by HSF1, NRF2 and p62/SQSTM1. Viruses, 11(1), 8.
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3

Transfection of Podocytes with GSK3β and RelA/p65

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The eukaryotic expression vectors encoding the WT GSK3β or S9A or KD mutant of GSK3β were provided by Dr Johnson (Birmingham, AL) and those encoding the HA-hRelA/p65 wt and HA-hRelA/p65 S468A were used previously.28 (link) The murine RelA/p65-specific siRNA duplex was provided by Santa Cruz Biotechnology (Santa Cruz, CA, USA) together with a scrambled siRNA control. Podocyte transfection was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) as described before.59 (link), 60 (link) In brief, podocytes were cultured under permissive conditions at 50% to 70% confluence in the absence of antibiotics. The DNA or siRNA oligomers-Lipofectamine 2000 complexes were prepared and applied to proliferating podocytes for transfection. The ratio of Lipofectamine 2000 to vectors or siRNA oligomers was optimized by a series of pilot experiments for each study until the best transfection efficiency or the best gene-silencing efficiency was achieved. After transfection, the cells were cultured under nonpermissive conditions in normal growth medium for 48 h before transfection efficiency or gene-silencing efficiency was assessed by immunocytochemistry staining or immunoblot analysis for target molecules. The cells were then subjected to LPS or vehicle treatments.
Bao H., Ge Y., Peng A, & Gong R. (2015). Fine-tuning of NFκB by Glycogen Synthase Kinase 3β directs the fate of glomerular podocytes upon injury. Kidney international, 87(6), 1176-1190.
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4

HRG Gene Silencing via siRNA

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An siRNA duplex used for silencing the human HRGwas purchased from Santa Cruz Biotech. Transfections were performed as described in the technical bulletin.
Menendez J.A., Benboudjema L., Vellon L., Rubio M.A., Espinoza I., Campisi J, & Lupu R. (2015). Heregulin, a new interactor of the telosome/shelterin complex in human telomeres. Oncotarget, 6(37), 39408-39421.
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5

Transfection of Podocytes with GSK3β and RelA/p65

Cited 1 time
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The eukaryotic expression vectors encoding the WT GSK3β or S9A or KD mutant of GSK3β were provided by Dr Johnson (Birmingham, AL) and those encoding the HA-hRelA/p65 wt and HA-hRelA/p65 S468A were used previously.28 (link) The murine RelA/p65-specific siRNA duplex was provided by Santa Cruz Biotechnology (Santa Cruz, CA, USA) together with a scrambled siRNA control. Podocyte transfection was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) as described before.59 (link), 60 (link) In brief, podocytes were cultured under permissive conditions at 50% to 70% confluence in the absence of antibiotics. The DNA or siRNA oligomers-Lipofectamine 2000 complexes were prepared and applied to proliferating podocytes for transfection. The ratio of Lipofectamine 2000 to vectors or siRNA oligomers was optimized by a series of pilot experiments for each study until the best transfection efficiency or the best gene-silencing efficiency was achieved. After transfection, the cells were cultured under nonpermissive conditions in normal growth medium for 48 h before transfection efficiency or gene-silencing efficiency was assessed by immunocytochemistry staining or immunoblot analysis for target molecules. The cells were then subjected to LPS or vehicle treatments.
Bao H., Ge Y., Peng A, & Gong R. (2015). Fine-tuning of NFκB by Glycogen Synthase Kinase 3β directs the fate of glomerular podocytes upon injury. Kidney international, 87(6), 1176-1190.
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6

LGR5 Knockdown in SH-SY5Y Cells

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SH-SY5Y cells were seeded at a density of 2.5×106 cells/cm2 in two six-well culture plates and cultured in antibiotic-free normal growth medium supplemented with FBS. Cells were incubated up to 60~80% confluency for 18~24 h and LGR5 knockdown was performed via LGR5 siRNA transfection. First, 10 μM of LGR5 siRNA duplex (Santa Cruz Biotechnology, Santa Cruz, CA, USA) diluted in opti-MEM medium (Gibco) was mixed with lipofectamine 2000 transfection reagent (Invitrogen, Carlsbad, CA, USA) diluted in opti-MEM medium (Gibco) at a ratio of 1:1 according to the manufacturer’s instructions. After 5 m incubation, the mixed solution was added to cells at different concentrations (30, 40, 50, 80, and 160 μM) with culture medium. The transfected cells were analyzed after 24 h at 37°C in a CO2 incubator. A control using 10 μM siRNA duplex (Santa Cruz) was produced by the same method. LGR5 siRNA is a pool of 3 different siRNA duplexes. A, 5′-GGAUGACAAUGCGUUAACAtt-3′ (sense), 5′-UGUUAACGCAUUGUCAUCCtt-3′ (antisense); B, 5′-CUCCAACCUUAAAGAAC-UAtt-3′ (sense ), 5′-UAGUUCUUUAAGGUUGGAGtt-3′ (anti-sense); and C, 5′-GAA-AGAUGCUGGAAUGUUUtt-3′ (sense), 5′-AAACAUUCCAGCAUCUUUCtt-3′ (anti-sense). Control siRNA duplex: 5′-UUCUCCGAACGUGUCACGUtt-3′ (sense), 5′-ACGUGACACGUUCGGAGAAtt-3′ (anti-sense).
Hwang M., Han M.H., Park H.H., Choi H., Lee K.Y., Lee Y.J., Kim J.M., Cheong J.H., Ryu J.I., Min K.W., Oh Y.H., Ko Y, & Koh S.H. (2019). LGR5 and Downstream Intracellular Signaling Proteins Play Critical Roles in the Cell Proliferation of Neuroblastoma, Meningioma and Pituitary Adenoma. Experimental Neurobiology, 28(5), 628-641.
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7

Silencing TRPV1 in Cardiomyocytes

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To silence the Trpv1 gene in cardiomyocytes, we performed transfection with small interfering RNA (siRNA) duplex, as described by the manufacturer (Santa Cruz Biotechnology). A pool of 3 different Trpv1 siRNAs was synthesized by Sangon Biotech (Shanghai, People's Republic of China): siRNA 1, sense GAAGACCUGUCUGCUGAAAtt, antisense UUUCAGCAGACAGGUCUUCtt; siRNA 2, sense CGAGCAUGUACAAUGAGAUtt, antisense AUCUCAUUGUACAUGCUCGtt; siRNA 3, sense CGCAUCUUCUACUUCAACUtt, antisense AGUUGAAGUAGAAGAUGCGtt. A nonrelated scrambled siRNA was used as a control. All measurements were performed 72 hours after transfection, at which time TRPV1 levels were decreased by 60% to 65%.
Chen M., Xin J., Liu B., Luo L., Li J., Yin W, & Li M. (2016). Mitogen‐Activated Protein Kinase and Intracellular Polyamine Signaling Is Involved in TRPV1 Activation–Induced Cardiac Hypertrophy. Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease, 5(8), e003718.
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