OBJECTIVES: A reliable high-throughput assay system is necessary for the analysis of the ever-increasing numbers of single-nucleotide polymorphisms (SNP) relevant to genetic screening studies. We describe an assay suitable also for large-scale screening programs. DESIGN AND METHODS: The one-step assay is based on asymmetric PCR amplification of the target sequence and subsequent time-resolved fluorescence measurement. Asymmetric amplification results in a single-stranded PCR product that is detected in the amplification vessel with a highly sensitive, homogeneous hybridization method. RESULTS: A dual label, homogeneous high-throughput platform for nucleic acid sequence analysis was developed and validated using a C/T single-nucleotide polymorphism in the insulin gene as a model analyte and applied also to two other SNP-assays (poliovirus receptor A/G-polymorphism and CD86-gene exon 2 A/G-polymorphism). CONCLUSIONS: The described high-throughput genotyping technology is very competitive in price, simple in design and easily applied to any analyte sequence.
OBJECTIVES: A reliable high-throughput assay system is necessary for the analysis of the ever-increasing numbers of single-nucleotide polymorphisms (SNP) relevant to genetic screening studies. We describe an assay suitable also for large-scale screening programs. DESIGN AND METHODS: The one-step assay is based on asymmetric PCR amplification of the target sequence and subsequent time-resolved fluorescence measurement. Asymmetric amplification results in a single-stranded PCR product that is detected in the amplification vessel with a highly sensitive, homogeneous hybridization method. RESULTS: A dual label, homogeneous high-throughput platform for nucleic acid sequence analysis was developed and validated using a C/T single-nucleotide polymorphism in the insulin gene as a model analyte and applied also to two other SNP-assays (poliovirus receptor A/G-polymorphism and CD86-gene exon 2 A/G-polymorphism). CONCLUSIONS: The described high-throughput genotyping technology is very competitive in price, simple in design and easily applied to any analyte sequence.
Authors: Kristiina Luopajärvi; Janne K Nieminen; Jorma Ilonen; Hans K Akerblom; Mikael Knip; Outi Vaarala Journal: Pediatr Diabetes Date: 2012-02-15 Impact factor: 4.866
Authors: R Hermann; K Lipponen; M Kiviniemi; T Kakko; R Veijola; O Simell; M Knip; J Ilonen Journal: Diabetologia Date: 2006-04-14 Impact factor: 10.122