MiR-NC, miR-218 mimic, anti-MiR-NC, and anti-miR-218 were purchased from Ambion (Austin, TX, USA). Small interfering RNA-218 (siRNA-218) targeting survivin and the siRNA control were purchased from GenePharma (Shanghai, People's Republic of China). The miR-218 mimic, inhibitor, or control were diluted in Opti-MEM medium (Life Technologies, CA, USA) at room temperature (RT) for 15 mins, then transfected human CC cells with miR-218 mimic or inhibitor and cultured for 48 hrs. The expression of miR-218 was examined by qRT-PCR assay. siRNA transfection was performed with Lipofectamine RNAiMAX reagent (Invitrogen, CA, USA) following the manufacturer’s instructions. The expression of survivin was examined by qRT-PCR and western assay.
Anti mir 218
Manufactured by Thermo Fisher Scientific
Sourced in United States, Canada
Anti-miR-218 is a synthetic oligonucleotide designed to inhibit the function of microRNA-218 (miR-218). miR-218 is a small, non-coding RNA molecule that plays a role in regulating gene expression. Anti-miR-218 can be used in research applications to study the biological functions and cellular pathways associated with miR-218.
Automatically generated - may contain errors
Lab products found in correlation
Opti mem medium, by Thermo Fisher Scientific (1 mentions)
Anti mir nc, by Thermo Fisher Scientific (1 mentions)
Mir nc, by Thermo Fisher Scientific (1 mentions)
Mir 218 mimic, by Thermo Fisher Scientific (1 mentions)
Lipofectamine rnaimax reagent, by Thermo Fisher Scientific (1 mentions)
Pmir report luciferase vector, by Thermo Fisher Scientific (1 mentions)
β gal vector, by Thermo Fisher Scientific (1 mentions)
Lipofectamine 2000 reagent, by Thermo Fisher Scientific (1 mentions)
Luciferase reporter assay kit, by Promega (1 mentions)
Siport neofx transfection agent, by Thermo Fisher Scientific (1 mentions)
3 protocols using anti mir 218
1
Investigating miR-218 and Survivin in Cancer
2
Validating miR-218 Targeting of UTRs
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The 3′UTR of COL10A1 and MEF2C containing a seed region for miR-218 were amplified from genomic DNA by PCR. The 3′UTR fragment was cloned into SpeI and HindIII restriction sites of pMIR-REPORT Luciferase vector (ThermoFisher Scientific). The 3′UTR of RUNX2 containing a seed region for miR-218, as well as all mutated sequences, were chemically synthesized (GeneArt, Germany). Primers used for this assay are listed in Table S2 .
For miR gain-of-function experiments, microRNA mimics and the miR-218 inhibitor (anti-miR-218) were purchased from Ambion (mirVana: MC10328, MH10328, 4464058).
HEK293T cells, seeded a day before transfection at a density 5 × 104 cells/well, were cultured on 24 well plates. Co-transfection of 50 nM of a selected miR mimic combined with 250 ng of a corresponding tested reporter construct, together with 250 ng of a normalization control β-Gal vector (ThermoFisher Scientific), was carried out using Lipofectamine 2000 reagent (Invitrogen, Germany). Luciferase activity was measured 48 h after transfection with Victor3 Multilabel Counter 1420–042 using the Luciferase Reporter Assay Kit (Promega, USA). Luciferase intensity signals were normalized to the β-Galactosidase activity in the same sample. Three independent experiments with six biological replicates were performed for this assay.
For miR gain-of-function experiments, microRNA mimics and the miR-218 inhibitor (anti-miR-218) were purchased from Ambion (mirVana: MC10328, MH10328, 4464058).
HEK293T cells, seeded a day before transfection at a density 5 × 104 cells/well, were cultured on 24 well plates. Co-transfection of 50 nM of a selected miR mimic combined with 250 ng of a corresponding tested reporter construct, together with 250 ng of a normalization control β-Gal vector (ThermoFisher Scientific), was carried out using Lipofectamine 2000 reagent (Invitrogen, Germany). Luciferase activity was measured 48 h after transfection with Victor3 Multilabel Counter 1420–042 using the Luciferase Reporter Assay Kit (Promega, USA). Luciferase intensity signals were normalized to the β-Galactosidase activity in the same sample. Three independent experiments with six biological replicates were performed for this assay.
3
miR-218 and cezanne regulation in fibroblasts
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Pre–miR-218 and pre-miRNA negative control, anti–miR-218, and anti-miRNA negative control were purchased from Ambion, Burlington, Canada. SMARTpool ON-TARGET cezanne siRNA (si-cezanne) and negative control were purchased from Thermo Scientific, Ottawa, Canada. For all the assays based on miR-218 overexpression, gingival fibroblasts were cultured in DMEM/10% FBS and then transfected using siPORT NeoFX transfection agent (Invitrogen) with pre-miRNA or negative control at a final concentration of 10 nM. Cells were used for further treatment 48 h after transfection. For all the assays based on the inhibition of miR-218 expression, human dermal fibroblasts were transfected with anti-miRNA or negative control with the same method. For all the assays based on the inhibition of cezanne expression, human gingival fibroblasts were transfected with si-cezanne or negative control with the same method.
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