mirVana™ miRNA Inhibitor, Negative Control #1

Using TaqMan miRNA assays, you can measure the level of miRNA of interest. If the miRNA is bound to the synthetic anti-miRs and/or mirVana inhibitors transfected, the duplex should not be detected, indicating efficient miRNA inactivation. Please note, this only works when total RNA sample isolated is not heat denatured when RT primer is added, as heat denaturation will result in complex dissociation and subsequent detection of microRNA by TaqMan miRNA assay even when cells have been transfected properly. Please see the protocol for the TaqMan MicroRNA Cells-to-CT kit (Cat. No. 4391848) for more details.

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Mimics and inhibitors can be transfected or electroporated into cells using parameters similar to those used for siRNA.

Using mirVana miRNA Mimics- For the following applications, use a cell line that expresses relatively low levels of the mature miRNA of interest.
- Cell density at the time of transfection. In general, 30-70% confluency is recommended.
- Transfection method (traditional or reverse transfection).
- Length of exposure of cells to transfection agent-RNA oligonucleotides complexes.
For many miRNA mimics, maximal activity is achieved after 24 hours, but biological assays may be performed 24-72 hours posttransfection. If you maintain your transfected cells longer than 24 hours, we suggest replacing the existing medium with fresh medium at 24 hours posttransfection, for greater viability of the cells.

Using mirVana miRNA Inhibitors to analyze endogenous miRNA function
For the following applications, use a cell line that expresses relatively high levels of the mature miRNA of interest.
General transfection starting points
mirVana miRNA Mimics and Inhibitors typically work best when transfected at a final concentration of 3-30 nM. Optimization experiments might include a concentration range of 1-100 nM. To increase accuracy and reproducibility when preparing transfection complexes, prepare a dilution of your stock oligonucleotides, and pipet a higher volume of diluted stock.

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The turnaround time varies depending on the level of purification and whether it is a custom order or not. Please see the approximate timetable below:

Standard purification (5 and 20 nmol), 4 business days
HPLC/in vivo ready (250 nmol), 15 business days
Custom mirVana Mimics and Inhibitors, 15 business days

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Yes, please provide us with your unpublished miRNA sequence and we can synthesize a corresponding mirVana mimic or inhibitor by clicking on this link (www5.invitrogen.com/custom-genomic-products/tools/mirna/).

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Negative control sequences require a confidential disclosure agreement while target sequences require prior proof of purchase for release.

Find additional tips, troubleshooting help, and resources within our RNAi Support Center.