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LaserGen Could Improve Sequencing Platforms

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Two years after publishing its first reversible terminator nucleotide, sequencing technology company LaserGen has developed a full set of reagents that it claims could improve a variety of existing and impending second-generation sequencing platforms.

The Houston-based startup plans to commercialize the reagents, called Lightning Terminators, for use with at least one existing sequencing platform. Long term, it wants to develop its own sequencing instrument.

The company's dye-labeled, photo-cleavable reversible terminators differ from many others in that they have an unblocked 3'-OH group, which enables researchers to use them in sequencing reactions with off-the-shelf polymerases.

A single light-induced cleavage step removes the terminating group as well as the dye and leaves a naturally occurring base. The company has also been working on chemically cleavable versions of the terminators.

Kinetic experiments in collaboration with researchers at New England Biolabs have shown that the Lightning Terminators have an "efficiency value" — a combination of how well they bind to the polymerase and how fast they get incorporated — that is better than that of natural nucleotides, according to LaserGen founder and CEO Michael Metzker, who is an associate professor at Baylor College of Medicine's Human Genome Sequencing Center.

Even though the researchers have to unblock the growing DNA chain after each cycle, using UV light, a polymerase incorporates the terminator nucleotides at a rate of 15 bases per second, faster than Pacific BioSciences' real-time sequencing technology at the moment, Metzker points out.

In addition, polymerase makes fewer mismatch errors using Lightning Terminators than using natural nucleotides, he says.

He and his colleagues have been using Vent, a wild type polymerase, and Therminator, a wild type enzyme with a single mutation, to test the reagents, and "both tend to show the same kind of performance with these Lightning Terminators," he says.

— Julia Karow

Sequencing Notes

Complete Genomics will sequence glioblastoma and melanoma samples for the Broad Institute as part of a pilot study. For the project, Complete Genomics will use its DNA sequencing technology to sequence five samples provided by the Broad: one sample well characterized by the community, and four genomes of glioblastoma and melanoma samples and their matched normal controls.

Febit joined the Revolutionary Approaches and Devices for Nucleic Acid Analysis consortium, a European group focused on developing DNA sequencing technologies, and plans to advance its gene capturing technology for use with the group's members. The group includes 16 industry and academic research entities and was funded by the European Union with €12 million.

Datapoint

10
Helicos BioSciences co-founder Steve Quake sequenced his own -genome to 10-fold -coverage using the company's platform.

Funded grants

$557,839/FY 2008
Genome-scale anaylsis of DNA methylation in CpG Islands with bisulfite sequencing
Grantees: Huidong Shi, University of Missouri, and Kun Zhang, University of California, San Diego
Began: Sep. 15, 2008; Ends: Jul. 31, 2012

Shi and Zhang plan to "[develop] two complementary methods that can specifically extract all non-repetitive CpG islands (or any subset) from the bisulfite converted genome, and coupling these target selection methods with next-generation DNA sequencing technologies," according to the abstract.

$392,500/FY 2008
Development of a universal tagging method for genome wide ChIP analyses
Grantees: Peter Scacheri and Zhenghe Wang,
Case Western Reserve University
Began: Sep. 1, 2008; Ends: Jun. 30, 2011

Scacheri and Wang will continue to develop their method to get around ChIP-grade antibodies by knocking in epitope tag-encoding DNA into endogenous loci. They will test the approach in various cell lines, "assess the fidelity of the 3xFLAG tagged transcription factors," and "develop methods for high-throughput tagging."

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