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Next-Gen Sequencer Finds Genotyping Niche

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Keygene has used 454 Life Sciences’ Genome Sequencer 20 to help develop a technology platform that will enable large-scale SNP discovery and detection in higher eukaryotic organisms.

Specifically, the technology, called Complexity Reduction of Polymorphic Sequences, or CRoPS, will make large-scale polymorphism discovery more affordable in organisms with low levels of germplasm polymorphism and/or highly repeated genomes, which includes many crop species.

“The 454 technology, in combination with Keygene’s current technology platform, creates many new opportunities that further increase the efficiency of genotyping in plant species,” says Michiel van Eijk, manager of upstream research at Keygene.

For CRoPS, Keygene researchers use the company’s AFLP technology to prepare tagged complexity-reduced libraries of two or more genetically diverse samples, which are then sequenced at five- to 10-fold redundancy with 454’s instrument. A typical sequence run yields more than 200,000 sequence reads with a median length of 100 bases. The resulting sequences are clustered, and bioinformatics tools are used to inspect the sequence contigs for differences. Keygene then applies quality measures to separate sequence errors from true polymorphisms, based on redundant sequencing, sample origin information, and allele frequencies.

“On pepper, a typical crop for which only very limited sequence information is available, we identified ... roughly 6,000 high-quality polymorphic sites and hundreds of SSR sequences in only two CRoPS runs on the GS20, generating 450,000 reads,” comments Mark van Haaren, business development manager of Keygene. “Up until now, we have been looking at fingerprints in bands on a gel — no sequence information whatsoever. With this new technology ... you can actually take each of the bands and sequence them. The fact that you can do many thousands of reads at the same time opens up this possibility.”

— Kate O’Rourke

PATENT WATCH
US Patent No. 6, 982,165. Nucleic acid sequencing by Raman monitoring of molecular deconstruction. Inventors: Mineo Yamakawa, Andrew Berlin, Steve Kirch, Gabi Neubauer, Valluri Rao. Assignee: Intel. Issued Jan. 3, 2006

The methods and apparatus concern sequencing single molecules of single-stranded DNA or RNA by exposing the molecule to exonuclease activity, removing free nucleotides one at a time from one end of the nucleic acid, and identifying the released nucleotides by Raman spectroscopy or FRET.


US Patent No. 6,975,943. Clone-array pooled shotgun strategy for nucleic acid sequencing. Inventors: Richard Gibbs, Allan Bradley, Wei-Wen Cai. Assignee: SeqWright. Issued: December 13, 2005.

The patent covers a simplified strategy for sequencing large genomes. Clone-Array Pooled Shotgun Sequencing is based upon pooling rows and columns of arrayed genomic clones for shotgun library construction. Random sequences are accumulated and the data are assembled by sequential comparison of rows and columns, to resolve the sequence of clones at points of intersection. Compared to either a clone-by-clone approach or whole genome shotgun sequencing, CAPSS requires relatively few library constructions and only minimal computational power for a complete genome assembly.


The US Department of Agriculture has awarded $10 million to the University of Illinois to obtain a draft sequence of the swine genome.

Helicos BioSciences has partnered with a select group of experimental researchers, providing them early access to the company’s next-generation sequencer, True Single Molecule Sequencing technology.

Researchers at Korea Research Institute of Bioscience and Biotechnology have sequenced the genome of Hahella chejuensis, a marine bacterium capable of killing Cochlodinium polykrikoides, a major red-tide dinoflagellate problematic in the western coasts of the North Pacific. The work has led to the identification of a molecule that effectively kills red-tide phytoplankton.

The USDA and DOE will share resources and coordinate the study of plant and microbial genomics. The DOE will tackle the sequencing of the soybean genome as the first project resulting from the agreement.

Scientists at Indiana University and the Joint Genome Institute have completed a shotgun sequence for Daphnia pulex, commonly known as the water flea.

An international team of scientists has determined and compared the genome sequences of three aspergilliAspergillus fumigatus, a potentially deadly human pathogen, A. oryzae, used in the production of soy sauce and sake, and A. nidulans, a model genetic organism.

Datapoint
$130 million
The National Human Genome Research Institute has allocated $130 million for fiscal year 2006 to renew its large-scale sequencing program. The institute is encouraging the adoption of next-generation sequencing technologies that it expects to reduce the cost of large-scale sequencing projects by five-fold or more over the next four years.

 

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