Arrayed primer extension reaction for genotyping on oligonucleotide microarray.
Arrayed primer extension reaction (APEX) is a straightforward and robust enzymatic genotyping method in which hundreds to thousands of variations in the genome are simultaneously analyzed in a single multiplexed reaction. It differs from allele-specific hybridization in that the genotype information in APEX is obtained by single base extension, performed by a specific DNA polymerase, together with four different dye terminators. This approach ensures highly specific discrimination without allele-specific hybridization, because the primer to be extended anneals just adjacent to the DNA base that needs to be identified. Selection of primers for specific sites or their consecutive placement in tiled format, shifting them by one base, permits single-nucleotide polymorphism analysis, mutation detection, or resequencing of the DNA template. Careful primer design also permits the analysis of insertions, deletions, splice variants, gene copy numbers, or CpG islands within the genome for gene methylation studies, by performing additional bisulfite reactions.
Pullat
Janne
J
Department of Biotechnology, Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia.
Metspalu
Andres
A
D018899:CpG Islands; D004252:DNA Mutational Analysis / Q000379:methods*; D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase; D018628:Gene Dosage; D005838:Genotype; D006801:Humans; D025202:Molecular Diagnostic Techniques; D016254:Mutagenesis, Insertional; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D017384:Sequence Deletion
D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase