商品名称：miRCURY LNA miRNA Power Inhibitor (15) - No Modification
miRCURY LNA miRNA Inhibitors and Power Inhibitors are exceptionally potent, enabling assessment of cellular phenotypes in miRNA loss-of-function studies, even in difficult-to-transfect cell lines. miRCURY LNA miRNA Inhibitor Controls and Power Inhibitor Controls are similar in sequence length and LNA design to the inhibitors and have no homology to any known mouse, rat or human miRNA or mRNA sequence.
Tm-normalized miRCURY LNA miRNA Inhibitors have unmatched potency against all miRNAs, regardless of their GC content (see figure Examples of miRNA silencing with miRCURY LNA miRNA inhibitors). The Power Inhibitors are so potent that they can be added directly to cell cultures without the need for transfection reagents (see figure miRNA silencing via direct uptake (gymnosis) of Power Inhibitors). In addition, simple systemic administration of LNA-modified in vivo-grade miRNA inhibitors enable analysis of miRNA function in animal models.
miRNA silencing via direct uptake (gymnosis) of Power Inhibitors.
miRCURY LNA miRNA Inhibitors are antisense oligonucleotides with perfect sequence complementary to their targets. When introduced into cells, they sequester the target miRNA in highly stable heteroduplexes, effectively preventing the miRNA from hybridizing with its normal cellular interaction partners.
The potency of miRNA antisense inhibitors is determined by their affinity for their miRNA target and their biological stability. The affinity of normal inhibitors with conventional nucleotide chemistry is a function of the GC content of the miRNA target. Since the GC content of miRNAs can vary from 5–95%, potencies of such inhibitors will vary greatly according to the miRNA sequence, with essentially no potency with AT-rich miRNAs. >
We have exploited the high affinity properties of LNA chemistry to create Tm-normalized miRCURY LNA miRNA Inhibitors and miRCURY LNA miRNA Power Inhibitors. Varying the numbers and positions of LNA nucleotides in these DNA/LNA mixmer inhibitors and carefully choosing the target sequences normalizes the melting temperatures of the oligonucleotides within a narrow window around an empirically determined optimal high temperature (see figure Overall features of the third-generation miRNA inhibitor design). These design features ensure that miRCURY LNA miRNA Inhibitors have the same high efficacy, regardless of the GC content of their miRNA targets.
We provide the following two types of miRNA inhibitors:
The combination of LNA and phosphorothioate modifications dramatically improves the stability of miRCURY LNA miRNA Power Inhibitors against enzymatic degradation, so the efficacy of Power Inhibitors is significantly better than the regular inhibitors (see figure Enhanced potency of miRCURY LNA miRNA Power Inhibitors.
HeLa cells were transfected with a plasmid with a Renilla luciferase (transfection efficiency control) and an hsa-miR-21-5p target sequence in the 3'UTR of a Firefly luciferase reporter gene. Firefly luciferase expression is therefore suppressed by the corresponding endogenous miR-21-5p level in the cell. The next day, the cell cultures were transfected with different concentrations of regular and Power miRCURY LNA hsa-miR-21-5p inhibitors and negative controls. Reporter gene expression was measured with a dual Luciferase assay 24 H after transfection. Ratios of firefly and Renilla luciferase activity were calculated and normalized to values obtained with a firefly luciferase reporter with no miR target sequence (pLuc). The results illustrate that our miRNA inhibitors display sub-nanomolar potency under optimal transfection conditions, and the Power inhibitors display superior activity compared with the regular inhibitors.
The high potency of miRCURY LNA miRNA Inhibitors and miRCURY LNA miRNA Power Inhibitors allows them to be used at low concentrations, minimizing the risk of undesired secondary effects unrelated to the antisense activity.
DNA/RNA duplexes are a substrate for RNase H, which degrades the RNA strand, and we use this catalytic activity in our GapmeRs for RNA silencing. To ensure that our miRNA inhibitors do not degrade mRNA and long non-coding RNAs that happen to contain a complementary sequence, we have positioned the LNA nucleotides such that they are never distanced by more than a few bases. LNA effectively inhibits RNase H activity, ensuring that the LNA inhibitor/RNA duplexes are not recognized as substrates by RNase H. As a result, off-target effects on mRNA and lncRNA stability are minimized. Finally, ribosomes have strong helicase activity that can effectively remove even LNA-enhanced, high-affinity, short oligonucleotides. Therefore, our inhibitors also have minimal effects on translation of mRNAs that have complementary sequences in the open reading frame.>
Predesigned miRCURY LNA miRNA Inhibitors and miRCURY LNA miRNA Power Inhibitors have been designed for all known human, mouse and rat miRNAs. Since many miRNAs are phylogenetically conserved, this set of inhibitors covers a large proportion of vertebrate and invertebrate miRNAs. These inhibitors are also available with fluorescein (6-FAM) labels. All miRNA inhibitors are desalted and delivered dried down in tubes in 1 nmol, 5 nmol and 15 nmol quantities. >
We currently offer four predesigned negative controls, two for use with miRCURY LNA miRNA Inhibitors and two for use with miRCURY LNA miRNA Power Inhibitors. The miRCURY LNA miRNA Power Inhibitor Controls have phosphorothioate-modified backbones to match the miRCURY LNA miRNA Power Inhibitors.
For each category of inhibitor, we offer the following two controls:
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