Abstract
Allele-specific polymerase chain reaction (ASPCR) is an application of the polymerase chain reaction (PCR) that permits the direct detection of any point mutation in human DNA by analyzing the PCR products in an ethidium bromide-stained agarose or polyacrylamide gel. ASPCR works because an oligonucleotide primer that forms a 3′ mismatch with the DNA template will be refractory to primer extension by Thermus aquaticus DNA polymerase. Therefore, oligonucleotide primers specific for all known alleles can be synthesized and used to detect the alleles in DNAs of unknown genotype. ASPCR has already been used in DNA-based diagnostic techniques involving the diagnosis of genetic and infectious diseases. In this paper, we describe the use of ASPCR for the direct diagnosis of sickle cell anemia and discuss different methological aspects of ASPCR.
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