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Development and use of a microchip for molecular genotyping

Objective

- To develop a specific and cost-effective technology for detecting single nucleotide polymorphisms for genetic diagnostics and gene mapping by combining the AFLP and solid-phase minisequencing methods.
- To utilize the AFLP technology for the identification of novel single nucleotide polymorphisms and for the amplification of target material.
- To develop a micro-array format of the solid-phase minisequencing method.
- To show the feasibility of the technology by analysing population samples for Finnish disease causing point mutations and for Y-chromosomal and autosomal haplotypes.

For future molecular diagnostic purposes the aim will be to generate many data points reliably and at low cost. We will develop a technology for the simultaneous detection of large numbers of single nucleotide polymorphisms (SNPs) in a micro chip format. To achieve this goal we will utilize the advantages of the AFLP and solid-phase minisequencing methods. The AFLP technology allows co-amplification of thousands of target molecules simultaneously. The single nucleotide primer extension reaction used in the minisequencing assay alleviates the problem of low specificity of hybridization based methods. The project will consist of two phases. In its first phase the basic technology will be developed. The AFLP technology will be used to identify novel Y-chromosomal and autosomal SNPs and to enable multiplex amplification of genomic target sequences spanning the identified SNPs. A multiplex AFLP system will also be developed for amplification of DNA fragments spanning 50 disease causing mutations that are prevalent in the Finnish population. The minisequencing technique will be employed to acquire the selectivity and sensitivity to interrogate a complex mixture of multiplexed target material for identification of SNPs and disease mutations. Technology for semi-automatic construction solid-phase arrays of primers and for fluorescence detection of the SNP specific signals generated in the minisequencing reaction will be developed. In the second phase of the project the utility of microchip-based minisequencing SNP detection will be shown in a series of demonstration projects. A screening test for about 50 mutations that are common in the Finnish population will be set up and used for determination of the carrier frequency and regional distribution of the mutations in Finland. Y-chromosomal haplotype diversity will be studied to generate data on population diversity. In addition, in the same screening effort the diversity of autosomal haplotypes will be assessed.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

National Public Health Institute
Address
166,Mannerheimintie
00300 Helsinki
Finland

Participants (3)

KEYGENE NV
Netherlands
Address
90, 90 Agro Business Park
6700 AE Wageningen
Ludwig-Maximilians-Universitdt München
Germany
Address
Luisenstrasse
80333 München 2
University of Helsinki
Finland
Address
11,Kytosuontie
00014 Helsinki