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Custom ExiLERATE LNA™ qPCR primers

Design custom LNA™-enhanced primer sets for mRNA and ncRNA. Use our web tool to design high performance primers compatible with Exiqon’s ExiLERATE LNA™ qPCR cDNA Synthesis and SYBR® Green Master Mix kits.

  • Superior detection of any mRNA or lncRNA
  • Success guaranteed
  • Fast and easy – get results in under 3 hours
  • LNA™ qPCR primers – unmatched sensitivity and specificity
  • Greater flexibility in primer positioning – easily detect isoforms, SNPs or splice-variants
  • Optimized for use with ExiLERATE LNA™ qPCR cDNA Synthesis and SYBR® Green Master Mix kits

Features

Design ExiLERATE LNA™ qPCR primer sets for any RNA using Exiqon's Custom LNA™ qPCR Assay design tool. The tool is optimized for RNAs ranging between 50-10,000 bp in length, including mRNA, non-coding RNAs or your own custom sequences. For shorter RNA targets, < 50 bases, we recommend using our custom microRNA design tool.

LNA™-enhanced primers offer several advantages over conventional DNA primers. They increase the flexibility in primer positioning which is particularly important for AT-rich targets. In addition, LNA™-enhanced primers can be made shorter than conventional primers, which is advantageous when working with low abundant targets or targets from degraded/ highly complex samples. The use of LNA™ also allows precise adjustments of primer melting temperatures. Primers designed using our online tool have an optimal Tm for use with Exiqon's SYBR® Green Master Mix.

The easy-to-use design tool accepts sequence, accession or keyword as input. The resulting qPCR assays are displayed graphically for easy viewing and selection. In addition, the tool offers highly developed features, such as automatic handling of introns and variations as well as the ability to design common assays for transcript families. Advanced options let you select specific regions to target, anchor primers or make changes to introns and variations.

As a highly innovative feature, the design tool also links mRNA targets to predicted or validated microRNAs and allows easy ordering of associated microRNA qPCR assays.

Assays designed using the tool are optimized for use with ExiLERATE LNA™ qPCR cDNA Synthesis Kit and SYBR® Green Master Mix.

Coverage

PCR primer sets designed with Exiqon’s online software tool are provided in amounts sufficient for 200 or 500 PCR reactions when used with ExiLERATE LNA™ qPCR cDNA Synthesis Kit and SYBR® Green Master Mix. Please see the Instruction manual for ExiLERATE LNA™ qPCR for details.

Ordering

The primer sets can be ordered directly online or by contacting us with the web order reference number (obtained by downloading order as PDF from the shopping basket). Prices can be found by placing the designs in the shopping basket.

Design process

In a process which typically takes less than 30 seconds, an advanced algorithm evaluates approximately 2,500 primer pair designs based on more than 45 different criteria to find LNA™-enhanced optimal primer sets for your target sequence.

Note, the design software has been designed for mRNA but can also be used for long non-coding RNA and DNA templates.

The design criteria include:
  • Optimization of melting temperatures by varying the LNA™ distribution of the primers to ensure high amplification efficiency and specificity.
  • Calculations and adjustments of self-hybridization and cross-hybridization scores for efficient PCR reactions.
  • Adjustments to avoid potential primer-dimer formation in the PCR reaction.
  • Intelligent positioning of LNA™ bases in the primers based on Exiqon’s vast knowledge of LNA™ oligonucleotide design.
  • Sequence database searches to identify potential off-target hits in the transcriptome with high sequence similarity. This ensures that the designed primer sets are highly specific for the target for which they were designed.
The design of forward and reverse primers will be specifically adjusted toward each other and is optimized for use with ExiLERATE LNA™ qPCR SYBR® Green master mix.

Combined microRNA and mRNA analysis

The recommended PCR conditions are the same as for our pre-validated and custom miRCURY™ microRNA qPCR assays allowing mRNA and microRNA to be quantitated at the same time. For best co-reverse transcription of microRNA and mRNA/ncRNA we recommend using miRCURY Universal cDNA Synthesis Kit II in combination with a random-mer. A manual describing this protocol is available.

Figure 1 Schematic outline of the ExiLERATE LNA™ qPCR System

Schematic outline of the ExiLERATE LNA™ qPCR System. (Click to learn more)

Figure 2 Instant access to microRNAs that target your mRNA of interest

Instant access to microRNAs that target your mRNA of interest. (Click to learn more)

Unmatched sensitivity

The exceptional sensitivity of the ExiLERATE LNA™ qPCR primer sets is achieved by combining the following features:
  • Optimized design algorithm using LNA™ enhanced and Tm normalized primers
  • Universal RT with the ExiLERATE LNA™ qPCR cDNA synthesis kit
This combination enables accurate and reliable quantification of abundant mRNAs and lncRNAs from as little as 1 pg of total RNA input in the initial first-strand cDNA synthesis and efficient detection of most RNAs from 0.1 ng total RNA (Figure 1). High sensitivity is crucial for the detection of low abundance targets, such as lncRNA.

Excellent specificity

The incorporation of LNA™ in the PCR amplification primers (forward and reverse) facilitates the design of highly specific assays that can distinguish between RNA sequences that differ by only one nucleotide. In addition, the primers can be targeted freely over the target sequence despite AT content, due to the incorporation of LNA™ in the primers.

The use of ExiLERATE LNA™ qPCR SYBR® Green Master Mix, makes melt curve analysis possible, so that the specificity of an assay can be evaluated (Figure 2).

Superior performance – success guaranteed

The ExiLERATE LNA™ qPCR system offers superior performance compared to all major competitors, in terms of sensitivity (Figure 5). When testing the performance of assays designed to ten randomly selected mRNA transcripts, the ExiLERATE LNA™ qPCR assays were able to detect all targets at 1 ng total RNA input, a success rate far superior to all major competitors (Figure 6).
Enhanced by LNA™ – Success guaranteed

Note

The ExiLERATE LNA™ qPCR primers have been optimized for use with the ExiLERATE LNA™ qPCR, cDNA synthesis kit and SYBR® Green master mix kit. Use of other reagents will affect the quality of the results.

ExiLERATE guarantee

If you are unhappy with the performance of an ExiLERATE LNA™ qPCR assay, our technical support team will work with you to identify the issue and if required replace with a different assay for the same target at no extra cost. Data from specific customer experiments may be required to evaluate assay performance e.g. use of the GAPDH primer set and/or independent evidence for transcript expression. Any problem should be reported within 3 months of delivery. ExiLERATE LNA™ qPCR primers must be used with the ExiLERATE LNA™ qPCR, cDNA synthesis kit and SYBR® Green master mix kit. Exiqon reserves the right to modify the terms of this guarantee at any time.

Figure 1 ExiLERATE LNA™ qPCR system offers excellent sensitivity down to 0.1 ng total RNA
Excellent sensitivity. (Click to learn more)


Figure 2 Human GAPDH was amplified from a total RNA sample
Beautiful amplification and melting curves (human GAPDH). (Click to learn more)


Figure 3 Mouse GAPDH was amplified from a total RNA sample
Beautiful amplification and melting curves (mouse GAPDH). (Click to learn more)


Figure 4 Rat GAPDH was amplified from a total RNA sample
Beautiful amplification and melting curves (rat GAPDH). (Click to learn more)


Figure 5 ExiLERATE LNA™ qPCR sensitivity exceeds all major competitors
ExiLERATE LNA™ qPCR sensitivity exceeds all major competitors. (Click to learn more)


Figure 6 ExiLERATE LNA™ qPCR assays outperform all major competitors
ExiLERATE LNA™ qPCR assays outperform all major competitors. (Click to learn more)


NOTICE TO PURCHASER: DISCLAIMER OF LICENSE No license is conveyed with the purchase of this product under any of US Patents Nos. 5,210,015, 5,487,972, 5,804,375, 5,994,056, 6,171,785, 6,214,979, 5,538,848, 5,723,591, 5,876,930, 6,030,787, and 6,258,569, and corresponding patents outside the United States, or any other patents or patent applications, relating to the 5’ Nuclease and dsDNA-Binding Dye Processes. For further information contact the Director of Licensing, Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA.
 



Working with small RNAs?
Exiqon’s Custom LNA™ qPCR Assay design tool is optimized for RNAs ranging between 50-10,000 bp in length. For shorter RNA targets, we recommend using our custom microRNA design tool.
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