QIAseq Library Quant System

For qPCR-enabled quantification of NGS libraries

Features

  • Ready-to-use, predispensed, sequentially diluted DNA standard (array format)
  • Automatic calculation of library concentration
  • Simple protocol, high sensitivity, and wide dynamic range
  • Compatibility with Illumina and Ion Torrent/Proton NGS platforms
  • Compatibility with most qPCR instruments
GeneRead Library Quant Array

Cat. No. / ID: 333304

PCR arrays with accessory components for sample library quantification prior to next-generation sequencing
Product TypeKit
GeneRead Library Quant Array
QIAseq Library Quant Assay Kit
 CONFIGURE AT GeneGlobe
The QIAseq Library Quant System is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

The QIAseq Library Quant System provides a simple, out-of-the-box solution to quantify NGS libraries, enabling consistent results with every NGS run. The QIAseq Library Quant System uses real-time PCR to specifically quantify DNA molecules with sequencing adaptors at both ends, ensuring optimal cluster density or template-to-bead ratio.

Please note: GeneRead Library Quant Array (product number 180611) and GeneRead Library Quant Kit (product number 180612) will be phased out by the end of 2016. Please use QIAseq Library Quant Assay Kit (product number 333314) instead.


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Performance

The sensitivity and dynamic range of the QIAseq Library Quant System were compared against Agilent's High Sensitivity DNA Kit, using two NGS libraries of differing concentrations. The first library was quantifiable by both systems, but the low concentration of the second library was below the detection limit of Agilent's 2100 BioAnalyzer. By contrast, the QIAseq Library Quant Array was able to quantify both libraries (see figure  QIAseq Library Quant System enables quantification of libraries with concentrations below the detection limit of conventional methods).

See figures

Principle

Accurate quantification of library molecules is essential for the efficient use of NGS platforms; however, current spectrophotometric methods only measure total nucleic acid concentrations. The QIAseq Library Quant System, by contrast, uses real-time PCR to quantify only DNA molecules with adaptors at both ends, which are the only amplifiable molecules during emulsion PCR (Ion Torrent platform) or bridge PCR (Illumina platform), and therefore provides highly accurate quantification of amplifiable library molecules. The high sensitivity of real-time PCR allows quantification of libraries at very low concentrations, even below the detection threshold of conventional spectrophotometric methods.

The QIAseq Library Quant System is available in both a tube and an array format, and is optimized with QIAseq qPCR SYBR® Green Mastermixes to provide superior sensitivity and a wide dynamic range. Both formats provide a DNA standard specific to the Illumina or Ion Torrent platform. The array format provides this standard in five predispensed, sequential 10-fold dilutions mixed with a PCR primer assay in triplicate, ensuring that your sample library will fall within the detection range of the array (see figure  Principle of QIAseq Library Quant System).
See figures

Procedure

The array format workflow (see figure  QIAseq Library Quant Array workflow) begins with two tenfold dilutions of the sample library, to ensure that its concentration falls within the range of the serially diluted standards. The samples are mixed with QIAseq qPCR SYBR Green Mastermix and aliquoted in technical triplicate across the array plate. PCR is performed, and raw CT values are exported to the provided data analysis Excel file for automatic calculation of library concentration (Illumina platform) or template dilution factor (Ion Torrent/Proton platform).

The tube format workflow is similar to the array format workflow, with a few small differences. The procedure begins with preparation of five sequential tenfold dilutions of DNA Standard and two tenfold dilutions of the sample library. This ensures that the concentration of the library will fall within the range of the standard dilutions. Next, the diluted DNA standards and sample libraries are mixed with the provided PCR assay and the appropriate QIAseq qPCR SYBR Green Mastermix. PCR is performed, and CT values are exported to the provided data analysis Excel file for automatic calculation of library concentration (Illumina platform) or template dilution factor (Ion Torrent/Proton platform).

See figures

Applications

The QIAseq Library Quant Array and QIAseq Library Quant Kit provide rapid, accurate measurement of sample library concentration prior to sequencing with the Illumina or Ion Torrent/Proton NGS platforms.

Supporting data and figures

Resources

Analysis Software (8)
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for 384-well plates for Illumina libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for 384-well plates for Ion Torrent libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for 96-well plates for Illumina libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for 96-well plates for Ion Torrent libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for Rotor-Disc 100 rings for Illumina libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) or GeneRead Library Quant Array for Rotor-Disc 100 rings for Ion Torrent libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) for 72-well Rotor rings for Illumina libraries
New version – Excel-based data analysis template for use with GeneRead Library Quant Kit (assay format) for 72-well Rotor rings for Ion Torrent libraries
Brochures & Guides (2)
Introducing QIAseq
PDF (450KB)
Accelerate your NGS performance through Sample to Insight solutions
Accelerate your NGS performance through Sample to Insight solutions

FAQ

Are there important considerations for plasma generation and urine handling?

It is strongly advised to follow the recommendations for preparing sample material provided in the corresponding Protocol Sheet to ensure reliable results.

Plasma: It is recommended to perform plasma separation immediately after blood collection when using EDTA or citrate as anticoagulant to prevent the release of genomic DNA into the plasma fraction.

Urine: Because circulating cell-free DNA in non-stabilized urine samples is rapidly degraded after sample collection due to high nuclease activity, eluates may contain no DNA or exhibit low DNA concentration. Therefore, it is recommended to stabilize urine samples. Even when using stabilized urine, it is recommended to perform a centrifugation step immediately after stabilization to prevent the release of genomic DNA from cells. Alternatively, non-stabilized urine samples can be processed immediately after collection and centrifugation using ATL-pretreatment and automated DNA extraction as described in the corresponding Protocol Sheet.

FAQ ID - 3699