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ON-TARGETplus Set of 4 Upgrade

nr kat.: LU-MOUSE-XX-0010
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Producent: Horizon Discovery Group
nr kat.: LU-MOUSE-XX-0010

Opis

siRNA designed and modified for greater specificity

ON‑TARGETplus siRNA reagents reduce off-targets utilizing a patented dual-strand modification, while still providing guaranteed gene silencing of human, mouse and rat gene targets. This unmatched potency and specificity make ON‑TARGETplus siRNA the premium choice for optimal knockdown and reduced off-targets. Simply search for your gene of interest and choose from the available product formats and quantities.

Start your gene search:

 

Highlights

  • We now offer a 2 nmol size when purchasing pre-designed ON‑TARGETplus or siGENOME siRNAs for human, mouse, or rat.
  • Off-targets reduced by up to 90% compared to unmodified siRNA.
  • Guaranteed silencing by SMARTpool and 3 of 4 individual siRNAs (See Product Formats tab).
  • Sequence information provided with siRNA purchase.


A two-stranded mechanism requires a two-stranded solution

Our scientists and collaborators demonstrated in 2006† that siRNA off-targets are primarily mediated by antisense seed-region interactions, initiating the development of a dual-strand modification pattern to effectively reduce off-targets:

  • Sense strand is modified to prevent interaction with RISC and favor antisense strand uptake.
  • Antisense strand seed region is modified to destabilize off-target activity and enhance target specificity.


†Jackson et al., Position-specific chemical modification increases specificity of siRNA-mediated gene silencing. RNA12(7), 1197-1205 (2006).

Product Formats

  • SMARTpool: A mixture of 4 siRNA provided as a single reagent; providing advantages in both potency and specificity. siGENOME and ON‑TARGETplus are guaranteed† to silence by 75% or better. 
  • Set of 4: A convenient option for purchasing aliquots of all 4 individual siRNAs targeting a single gene. Three of four siGENOME and ON‑TARGETplus siRNAs are guaranteed† to silence by 75% or better.
  • Set of 4 Upgrade: Discounted price on the Set of 4 based on a concurrent or prior purchase of the SMARTpool reagent for the same gene.
  • Individual siRNAs: Select 1, 2, 3 or 4 individual siRNAs per gene.

    Our siRNA knockdown guarantee

    siGENOME and ON‑TARGETplus siRNA reagents (SMARTpool and three of four individual siRNAs) are guaranteed to silence target gene expression by at least 75% at the mRNA level when used under optimal delivery conditions (confirmed using validated positive control and measured at the mRNA level 24 to 48 hours after transfection using 100nM siRNA).

    Note: Most siGENOME and ON‑TARGETplus siRNA products are highly functional at 5 to 25nM working concentration.

 

Ordering Guidelines

MANUALS:

Dharmacon Product Line Selection

siGenome and on-Targetplus Product Insert 

siGenome on-Targetplus CONTROLS siRNA product insert 

Basic siRNA resuspension protocol

Off-target Technical Review


 

More information

ON‑TARGETplus modifications reduce the overall number of off-targets and pooling reduces them even further


Panels (A) and (B) are representative examples of off-target signatures with and without application of ON‑TARGETplus modifications to (A) a single siRNA and (B) a SMARTpool reagent. Green bars indicate genes with 2-fold or more reduction of expression when treated with the indicated siRNA reagent.The ON‑TARGETplus modifications reduced the off-targets when compared to unmodified siRNA. Pooling of siRNA and the ON‑TARGETplus modification pattern independently, and in combination, provide significant reduction in off-target gene silencing. Panel (C) represents quantitation of off-targets (down-regulated by 2-fold or more) induced by the indicated siRNA reagents targeting 10 different genes (4 siRNAs per gene or a single SMARTpool reagent). Off-targets were quantified using microarray analysis (Agilent) then compiled. Each shaded box represents the middle 50% of the data set. Horizontal line in box: Median value of the data set. Vertical bars: minimum and maximum data values.

 

siRNA designs with low-frequency seed regions ensure fewer off-targets


 

 

 

ON-TARGETplus siRNA designs leverage sophisticated bioinformatics to reduce the likelihood of miRNA-like off-targets from high-frequency or highly conserved miRNA seed regions. siRNAs with low seed frequency have a significantly lower number of off-targets than siRNAs with medium or high frequency seeds. Five siRNAs with low, medium, or high frequency seed regions were transfected into HeLa cells and their associated off-target signatures assessed via global expression profiling (Agilent 22K platform). siRNA sequences were constant at positions 1 and 8-19, only the seed regions (positions 2-7) were altered.


Low frequency seeds: < 350 occurrences in the HeLa transcriptome

Medium frequency: 2500-2800 occurrences

High frequency: >3800 occurrences

 

False phenotypes due to off-targets are alleviated by ON-TARGETplus SMARTpool reagents while target gene knockdown is maintained

 

The effect of silencing ARPC1B on cell migration was studied in a breast cancer cell line. A monolayer of cells was uniformly scraped and the rate of cell migration to close the scrape (wound healing) was evaluated. Both unmodified and ON-TARGETplus siRNA reagents induced potent target knockdown. Inconsistent phenotypes due to off-target effects (red outline), were observed for cells transfected with unmodified individual siRNAs. The unmodified SMARTpool improved the false phenotype considerably, while the ON-TARGETplus SMARTpool significantly reduced off-target effects to produce a consistent phenotype.

In collaboration with Kaylene Simpson, Laura Selfors, and Joan Brugge, Harvard Medical School.

 

ON-TARGETplus siRNA dual-strand modification pattern reduces off-targets 


A 2006 publication demonstrates that off-target effects are primarily driven by antisense strand seed activity†. Therefore, sense strand inactivation alone does not decrease the total number of off-target genes.

ON-TARGETplus modifications account for both strands:

Sense strand is modified to prevent interaction with RISC and favor antisense strand uptake
Antisense strand seed region is modified to minimize seed-related off-targeting

The ON-TARGETplus modification pattern dramatically reduces off-targets. Off-target effects induced by the indicated siRNAs were quantified using microarray analysis. For each target, three different siRNAs were used: unmodified, sense strand-inactivated, and ON-TARGETplus-modified. Data shown represents genes down-regulated by two-fold or more. HEK293 cells were transfected with 100 nM siRNA using 0.2 μL of DharmaFECT 1. Data was analyzed at 24 hours.

 

 

 


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