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Mar 1, 2011 (Vol. 31, No. 5)

Range of NGS Applications Rises Quickly

Advanced Technological Approach Generates Genomic Data Better, Faster, and Cheaper

  • Bearing Fruit

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    The GS Junior System offers a 10-hour run time for quick overnight sequencing and processing, according to Roche.

    Recent studies in the literature using Roche’s 454 sequencing technology include the discovery of a new human immunoglobulin (IGHV) gene and 16 new IGVH allelic variants published in the February 2011 issue of Immunogenetics. Additionally, a collaboration between 454, the University of Florida, the DOE Joint Genome Institute, and the Georgia Institute of Technology has borne fruit with completion of the first citrus genomes, including the sweet orange and the Clementine mandarin.

    Researchers from the Munich Leukemia Laboratory presented on the use of 454’s Genome Sequence (GS) FLX and GS Junior systems for targeted resequencing of genomic regions associated with blood cancers at the annual meeting of the American Society of Hematology. The ability of long-read sequencing technology to detect multiple types of variation including point mutations, insertions/deletions, and structural variation made it possible to identify novel mutations in patient samples and to stratify patients into disease risk subtypes, which aids in diagnosis, prognosis, and detection of reemergence of resistant disease following therapy.

    Soon to be introduced by 454 is new chemistry for the company’s GS FLX sequencing system that will enable longer reads of 700–800 base pairs and improve the quality of de novo genome assembly. The company also plans to launch a series of assays for application-specific workflows on both the GS FLX and GS Junior systems. The first assay will be primer sets for HLA genotyping, followed by assays targeting oncology, immunogenetics, and infectious disease applications.

    Roche is also partnering with DNA Electronics to develop a low-cost high-throughput DNA sequencing system that will combine 454’s current pyrosequencing-based platform with DNA Electronics’ semiconductor technology. The system would rely on electrochemical rather than optical detection technology to monitor nucleotide incorporation during sequencing.

  • Getting Personal

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    MiSeq™, Illumina’s new low-cost personal sequencing system, leverages the same TruSeq™ sequencing chemistry that drives the company’s flagship HiSeq™ platform. MiSeq can take purified DNA and generate analyzed sequence data in about eight hours and, in just over a day’s time produce more than one gigabase at a cost of $400–$750 per run. Illumina plans to ship the first commercial MiSeq units this summer.

    Key advantages of MiSeq are its fast turnaround time, ease of use, and simple sample prep, said David Bentley, Ph.D. Dr. Bentley, chief scientist at Illumina, envisions customers using the system for various types of applications: to check a small amount of sample before running it on HiSeq, to analyze large numbers of poor-quality DNA samples isolated from FFPE tissues, and to detect specific mutations in patient samples from clinical trial populations.

    Illumina has doubled the yield from the HiSeq system to 600 gigabases, increasing the instrument’s capacity to 4–5 sequenced genomes in a 10-day run. Sufficient scale and throughput are now available to allow users to analyze and compare hundreds of samples, and these improvements are requiring users “to set up more sophisticated automation platforms,” said Dr. Bentley.

    Illumina recently reached an internal milestone, completing a terabase run more than once. The company is nearing completion of work to validate HiSeq in its in-house CLIA sequencing laboratory in preparation for introducing the system into the clinical arena.

    Although the capability of next-gen sequencing technology continues to outstrip available computing power, “these problems are tractable,” said Dr. Bentley, and the need for more powerful informatics tools is systematically being addressed. “We are drastically reducing the amount of storage space needed to store the same amount of information,” he explained, pointing to positive signs for the future including Cloud-based initiatives for centralizing data analysis and the development of increasingly sophisticated and diverse informatics strategies for analyzing sequence data and extracting new and focused information.

    Dr. Bentley emphasized the need for software tools designed for “less expert users or experts in a different field.” As sequencing technology moves beyond the research laboratory, for example, it become increasingly important to design tools that will allow clinicians to extract and analyze the types of information most useful to them.

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    Ion Torrent’s Ion Personal Genome Machine is based on the company’s semiconductor sequencing chips, which translate chemical signals into digital information.

    Life Technologies’ Ion Personal Genome Machine (PGM™) is based on Ion Torrent’s semiconductor sequencing chips that translate chemical signals into digital information. The 314 sequencing chip contains an array of 1.3 million wells; each is the site of an individual sequencing reaction. A pH change is detected when incorporation of a new base onto a growing DNA strand produces hydrogen ions.

    The system is able to detect each base addition without the need for optics, a light source, or scanning detection technology. The 314 chip can yield at least 10 Mb of DNA sequence per run. From sample loading to raw data generation takes about two hours, with an additional hour for data analysis.

    Life Technologies is rapidly scaling throughput of the PGM, and when the new 316 chip becomes available in 2Q2011 it will generate 10-fold more sequence per run, or 100 Mb. The company expects to maintain a pace of increasing the throughput of its sequencer by a factor of 10 about every 6 months.

    The system’s housing and server will not change; users will only have to upgrade the chip and the software. The higher throughput chip will make it possible to sequence larger amplicon sets and whole microbial genomes on the PGM. Researchers have demonstrated the use of barcoding techniques to sequence multiple different samples on a single chip.

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