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Qiagen Introduces Sequencing Workflow, Including NGS System and Library-Prep Instrument, at AGBT


At last week's Advances in Genome Biology and Technology conference in Marco Island, Fla., Qiagen discussed in more detail its next-generation sequencing workflow and said that early-access customers would have systems sometime this year, with data likely to be publicly available in 2014.

As part of its next-gen sequencing initiative, Qiagen has also developed two new instruments: QiaCube NGS for automated library prep and the GeneReader for sequencing. It has not settled on a price for either box but said the QiaCube NGS would be slightly more expensive than its QiaCube instrument for sample processing, which runs around $13,000.

The initial sequencing system will make use of the QiaCube for sample processing, but the company is also working on a second version that will run with its QiaSymphony instrument. The QiaCube is able to process 12 samples at once, while the QiaSymphony can process up to 96 samples at once, or batches of samples.

The company is also developing a sample-prep kit for processing DNA from formalin-fixed paraffin-embedded tissue, which it said is "artifact free" and automated on its QiaCube instrument. The kit is able to remove sequences with cytosine to thymine changes that have occurred as a result of a chemical reaction during the processing of FFPE tissue. It is planning to make the kit available mid-2013.

Qiagen aims to tap into the clinical sequencing market by focusing on targeted sequencing panels. It launched its first sequencing-based products last November, including the Repli-g Single Cell kit, GeneRead rRNA Depletion kit, the GeneRead Library Quant kit, and the GeneRead DNAseq Gene Panel kits, all of which are compatible with any next-generation sequencing system and will also be compatible with its own sequencing system (IS 11/6/2012).

The DNAseq Gene Panel kits include a comprehensive cancer panel encompassing 124 genes and eight subpanels targeting relevant genes for breast, liver, lung, prostate, colon, gastric, and ovarian tumors, as well as leukemia (CSN 11/7/2012).

At last week's AGBT meeting, CEO Peer Schatz told In Sequence that the company has an additional 200 panels in its pipeline that are being developed from its GeneGlobe portfolio of annotated genes. While the company is currently creating these as real-time PCR panels, Schatz said that they are "portable to next-generation sequencing."

Sequencing Workflow

Qiagen's next-gen sequencing workflow includes sample-prep and processing on its QiaCube instrument, followed by automated library prep on the QiaCube NGS, and then sequencing on the GeneReader.

The entire workflow will take just over one day, including 130 minutes on the QiaCube, two and a half hours on the QiaCube NGS, and between 22 and 23 hours of sequencing on the GeneReader.

In a presentation, Dirk Löffert, the company's vice president of research and development for sample and assay platform technologies, described the workflow. Rather than purified DNA, input can be an FFPE block or blood sample. DNA is extracted and processed within the QiaCube, with each enzymatic step occurring in a single tube and reactions separated with an incubation step.

Next, the sample moves on to the QiaCube NGS box for library preparation. Qiagen has developed a proprietary chemistry that Schatz said offers similar performance to emulsion PCR, but is fully automated. Löffert described the technology as bead-based single-molecule amplification with an automated method to enrich only the beads carrying amplification products on their surface.

QiaCube NGS also has a built-in quality control step for assigning a quality score to each sequencing library, so that the user can decide which are good enough for downstream processing.

The selected libraries then move onto the GeneReader for sequencing. The system is based on sequencing-by-synthesis technology, and uses technology developed by Jingyue Ju at Columbia University, in which fluorescently-labeled terminator nucleotides are used in conjunction with less expensive dark nucleotides for strand extension. In a first extend, a "sufficient number" of fluorescently-labeled nucleotides are used, with a second extend filling in the remaining gaps with non-labeled nucleotides.

Qiagen gained the rights to the technology when it acquired Intelligent Bio-Systems last year (IS 6/26/2012). IBS licensed the technology from Columbia University.

Qiagen's GeneReader sequencing system can run up to 20 flow cells, and the system includes random-access and continuous loading features. Each flow cell can generate up to 2 gigabases of data, and up to 240 samples can be assessed per sequencing run when 12 samples are multiplexed in each flow cell.

The company did not disclose what its initial read lengths would be, but said that the system should be able to scale with read lengths that are comparable to that of Illumina's MiSeq system.

The system also comes with analysis software that can be run on a laptop computer. Qiagen collaborated with SAP to develop the bioinformatics, which is based on SAP's Hana platform. After sequencing, the FastQ files can be uploaded to the user's computer. Each gene panel will include its own software program, which can be updated with plugins as new content is added to the panels.