allele-specific PCR

allele-specific PCR

The amplification of specific alleles, or DNA sequence variants, at the same locus by polymerase chain reaction. Specificity is achieved by designing one or both PCR primers so that they partially overlap the site of sequence difference between the amplified alleles.

AS-PCR is two-step nested PCR technique that allows detection of DNA with a point mutation in the presence of a 103–105-fold excess of normal DNA. AS-PCR is based on the observation that Taq polymerase will extend primers with a 3’ mismatch 103 to 106-fold less efficiently than perfectly matched primers; by choosing a primer that is complementary at its 3’ end to a point mutation, selective amplification of a mutant allele can be achieved.
References in periodicals archive ?
Allele-specific PCR differs from standard real-time PCR in that the primers themselves contain the specific complementary sequence to the relevant area of the EGFR gene, mutation or control.
Allele-specific PCR is a variant form of real-time PCR in which short oligonucleotide probes, specific for the wild-type and variant sequence, are mixed together in the amplification step.
HLA-B27 typing: evaluation of an allele-specific PCR melting assay and two flow cytometric antigen assays.
Such frequently used techniques as pyrosequencing and allele-specific PCR have analytical sensitivities ranging from 1% to 10% for the detection of mutant DNA diluted into a background of wild-type DNA (18-20).
An allele-specific PCR assay developed was used to detect sensitive/ resistance alleles among 55 isolates of W.
In comparison, allele-specific PCR offers a simple and easily accessible approach to SNP genotyping.
Material is in sections on basic principles and software, genome-scale PCR primer design, multiplex PCR primer design, allele-specific PCR, long PCR primer design, and DNA methylation mapping.
Standard genotyping * Sequencing the total virus population in plasma Allele-specific PCR * Use of specific primers to detect the presence of specific mutant sequences in the viral population * More sensitive than standard genotyping Single-genome sequencing * Sequencing individual viral genomes * Can detect the presence of minor variants but sensitivity depends on number of genomes sequenced Table 2: Advantages and disadvantages of allele-specific PCR.
By harnessing the sensitivity of DxS's ARMSTM allele-specific PCR and its ScorpionsTM quantitative real-time signalling system, the new assay can detect mutant copies even if they represent as little as 1% of the sample.
Compared to currently available methods, the BeadChip Test, comprising a set of 18 genetic markers, reliably produced results equivalent to those of either serological hemagglutination or manual restriction fragment length polymorphism (RFLP) and allele-specific PCR analysis of individual markers, but in a manner substantially accelerating throughput.