Project: High-quality, flexible, and low cost gene capture for High-Throughput DNA Sequencing in research and clinical diagnostics.

Background_x000D_Considerable efforts have been invested in developing ultra high-throughput DNA sequencing technologies. These next generation sequencing (NGS) technologies enable rapid and cost-effective DNA analysis. However, sequencing a whole human genome in one single run is still a complicated and expensive endeavor. Albeit very powerful and promising, resequencing whole human genomes using NGS is not expected to be carried out in clinical diagnostics due to various reasons related to ethics, liability, complexity, and costs. _x000D__x000D_In fact, clinicians are rarely interested in semi-reliable complete genome sequences from individual patients. Instead, in-depth interrogations of subgenomes with a high level of confidence is often preferred. These small subgenomic regions with proven or suggested clinical relevance (also known as candidate genes) are different for each disease. _x000D__x000D_Existing technologies suffer from a range of drawbacks; high costs, limited scalability, incomplete and uneven capture of regions of interest, and incompatibility with sample multiplexing approaches. _x000D__x000D_Project Goal_x000D_The goal is to establish a high quality, highly flexible, low-cost, genomic selection reagent kit based on the integration of the Olink Genomics and FlexGen technologies. Olink and FlexGen aim to use the project's result to achieve commercial success in both academic and industrial genetic research and diagnostic markets._x000D__x000D_Project Outline_x000D_Olink Genomics (Uppsala, Sweden) has developed a proprietary DNA sample preparation method, called "Selector Technology", which enables sequencing of selected genomic regions in a single reaction volume. This technology is based on targeted capture of regions of interest using a pool of amplification guiding oligonucleotides. This technology currently requires the separate synthesis of all individual oligonucleotide sequences, a very expensive and time consuming process. _x000D__x000D_Flexgen (Leiden, Holland) has developed an instrument called the FlexArrayer that enables fast, affordable, and high-throughput manufacturing and copying of oligonucleotides using a laser-derived DNA synthesis procedure. _x000D__x000D_These two technologies combined will make it possible to manufacture, optimize and assemble reagent kits in a much faster and cost-effective way than is possible to date. Protocols will also be developed to process a variety of different sample types, such as paraffin embedded samples, blood samples, and fine needle biopsies. Finally, reagents kits will be developed to process multiple samples simultaneously using incorporation of sample specific bar-codes. Protocol optimization will be performed in collaboration with Uppsala University. _x000D__x000D_The fourth project P, the Vascular Medicine department of the Amsterdam Medical Centre (AMC) will implement the new genomic selection protocols in screening for novel and known mutations linked to familial hypercholesterolemia (FH), and validate and implement the new approach in clinical diagnostics and in molecular research of dyslipidemia and vascular disease. Cardiovascular disease is a major human disease-group; genetic research and screening is therefore of large clinical relevance.

Acronym	GeneSelect (Reference Number: 5480)
Duration	01/03/2010 - 31/08/2011
Project Topic	Integrating Olink Genomic's powerful gene-capture method with Flexgen's parallel DNA synthesis will generate genomic selection kits of superior quality at ultra low costs. The first kits will be used in vascular medicine and then be commercialized in genetic research and diagnostics worldwide.
Project Results (after finalisation)	WP 3: The department of vascular medicine at the AMC was closely involved in the design of the probes and oligonucleotide _x000D_pools for capture in collaboration with the Ps at Flexgen and HaloGenomics. _x000D_Several capture experiments have been performed to enrich for all exons and regulatory sequence of interest (LDLR, PCSK9 and apoB) in 10 subjects with different mutations. All exons were captured by the novel approach._x000D_The goal was to replace the expensive and time consuming Sanger based analysis with the novel technology. _x000D_The molecular diagnostics approach has not been changed yet, but given the promising results (point mutations were identified by the novel strategy) we will continue optimizing the strategy investigated with the collaborators from this grant._x000D_
Network	Eurostars
Call	Eurostars Cut-Off 4

Project partner

Number	Name	Role	Country
4	Amsterdam Medical Centre	Partner	Netherlands
4	FlexGen B.V.	Partner	Netherlands
4	Halo Genomics	Coordinator	Sweden
4	Uppsala University, Department of Genetics and pathology	Partner	Sweden