For studying the effects of an individual miRNA on a single target site

Features

Custom-designed target site blockers for specific inhibition of miRNA targets

Sophisticated design and superior high affinity, regardless of target sequence

Unmatched high efficacy in vitro and in vivo

Unrivaled performance and high protein expression due to lack of RNase H-dependent mRNA degradation

Efficient at very low concentrations, outcompeting miRNAs for their target sites

Superior biological stability for long-lasting antisense activity

Product Details

miRCURY LNA miRNA Power Target Site Blockers are antisense oligonucleotides that bind to the miRNA target site of an mRNA, preventing miRNAs from gaining access to that site. This enables you to study the effects of an miRNA on a single target. miRCURY LNA miRNA Power Target Site Blockers are efficient at very low concentrations, and due to their high affinity from LNA enhancement, the target site blockers outcompete miRNAs for their target sites.

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Performance

Unravel miRNA function with target site blockers

miRNAs typically regulate gene expression of multiple targets, and inhibition of an miRNA will result in derepression of all of these targets. Therefore, a phenotype observed upon miRNA inhibition is a composite result of derepression of several targets. However, deregulation of a few of these targets will often contribute significantly to the phenotype. Identifying these targets is important to understanding the function of the miRNA.

miRCURY LNA miRNA Power Target Site Blockers can be used to:

Determine which pathway is involved in a phenotype observed upon miRNA inhibition

Determine which miRNA/mRNA interactions are most important in a pathway containing several predicted targets

Examples of target site blocker applications are described in figures  Examples of target site blocker applications – pathways X and Y and  Examples of target site blocker applications – pathway Z.

The figure  The CAV1 target site blocker phenocopies the miR-199a-5p inhibitor in many important aspects shows an example of using a target site blocker to demonstrate how miR-199a-5p functions as a key effector of TGFβ signaling in lung fibroblasts.

Principle

What are target site blockers?

miRCURY LNA miRNA Target Site Blockers are LNA-enhanced antisense oligonucleotides that bind to the miRNA target site of an mRNA, thereby preventing miRNAs from gaining access to that site. This allows researchers to study the effects of an miRNA on a single target. In contrast, the phenotype observed when inhibiting an miRNA reflects the combined effects of that miRNA on all targets.

LNA-enhanced target site blockers

The incorporation of LNA into the miRCURY LNA miRNA Power Target Site Blocker means that they will compete more effectively with the miRNA/RISC complex for the miRNA target site. In addition, LNA distribution throughout the LNA/DNA mixmer ensures that the antisense oligonucleotide does not catalyze RNase H-dependent degradation of the mRNA. As a result, the TSB will cause increased expression of the protein encoded by the targeted mRNA by preventing miRNA-mediated translational attenuation.

Procedure

Following resuspension, miCURY LNA miRNA Target Site Blockers are transfected into cells with a transfection reagent or via electroporation. Phenotypic effects of the target site blocker are normally assessed 24–72 hours after transfection. For some applications, such as cell differentiation assays, the phenotypic readout may take place 7–10 days after transfection.

Applications

miRCURY LNA miRNA Target Site Blockers can be used for:

Determining which pathway is involved in a phenotype observed upon miRNA inhibition

Determining which miRNA/mRNA interactions are most important in a pathway containing several predicted targets