Detection of specific alleles by using allele-specific primer extension followed by capture on solid support.
This article describes a method for determining whether a particular nucleic acid sequence is present in a sample and for discriminating between any two nucleic acid sequences if such sequences differ only by a single nucleotide. The method entails extension of a novel two-component primer on templates that may or may not include a target nucleic acid sequence. The 3' portion of the primer is complementary to a portion of the template adjacent to the target sequence (for example, the polymorphic nucleotide). The 5' portion of the primer is complementary to a different preselected nucleic acid sequence. Extension of the 3' portion of the primer with a labeled deoxynucleoside triphosphate yields a labeled extension product, but only if the template includes the target sequence. The presence of such a labeled primer-extension product is detected by hybridization of the 5' portion to the preselected sequence. The preselected sequence is immobilized on a solid support. The method has been applied to genotyping individuals for the two-allele polymorphism of the human tyrosinase gene.
Ugozzoli
L
L
Department of Molecular Biochemistry, Beckman Research Institute of the City of Hope, Duarte, California.
Wahlqvist
J M
JM
Ehsani
A
A
Kaplan
B E
BE
Wallace
R B
RB
D000483:Alleles; D001483:Base Sequence; D004247:DNA / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000235:genetics; D011110:Polymorphism, Genetic
D004247:DNA; D014442:Monophenol Monooxygenase