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What are allele specific primers?

What are allele specific primers?

Allele Specific Primer Extension (ASPE) is a solution based, sequence specific enzymatic reaction technology that can be used to assay multiple SNPs in a single tube. This is done with the help of an appropriate capture sequence attached to the allele specific oligonucleotide.

What is allele specific PCR used for?

Allele-specific polymerase chain reaction (AS-PCR) is a technique based on allele-specific primers, which can be used to analyze single nucleotide polymorphism (SNP) effectively including the transition, transversion and insertion/deletion polymorphism and has been exploited in the study of diseases research, molecular …

What is the principle of allele specific PCR?

The basic principle is that one of your primers covers the variant you’re interested in, so that, for example, if your primer matches the mutant allele then it will bind to mutant DNA but not WT DNA, so that you only get amplification if the mutant allele is present.

What does allele specific mean?

An allele-specific oligonucleotide (ASO) is a short piece of synthetic DNA complementary to the sequence of a variable target DNA. It is designed (and used) in a way that makes it specific for only one version, or allele, of the DNA being tested.

What is SSP PCR?

Sequence-specific amplification (SSP) is simply a form of polymerase chain reaction (PCR) which involves designing one or both primers so that they will or will not allow amplification (the 3′-mismatch principle). Its origins are probably legion, i.e. many people probably thought of it at the same time.

How does methylation PCR work?

Methylation-specific PCR (MSP) is a method for analysis of DNA methylation patterns in CpG islands. For performing MSP, DNA is modified by and PCR performed with two primer pairs, which are detectable methylated and unmethylated DNA, respectively.

What is allele specific amplification?

Allele specific amplification differs from the normal PCR in that primers are carefully designed to target the region of difference in the different alleles so that a pair of primers only amplify a specific allele but not others.

What is inverse PCR used for?

Inverse polymerase chain reaction (Inverse PCR) is a variant of the polymerase chain reaction that is used to amplify DNA with only one known sequence.

What is the principle of allele specific oligonucleotide?

Allele-specific oligonucleotides allow amplification of a sequence of DNA to which they are perfectly homologous (1), but not one containing a mismatch (2).

What is SSP typing?

HLA typing by sequence-specific primers (PCR-SSP) is a commonly used technique in HLA typing in which multiple pairs of cis-located allele-specific primers are used to determine the alleles present in a given DNA sample.

How does PCR SSP work?

How do you design primers for Methylation Specific PCR?

Below are Herman’s tips for successful MSP primer design.

  1. Design primers to the bisulfite-converted template.
  2. Make sure there are non-CpG C’s in the target sequence.
  3. Aim for six to seven CpG’s.
  4. Keep the targets short.
  5. Obey the rules of PCR.
  6. Use the appropriate computer program.

How is allele specific primer extension ( Aspe ) used?

Allele Specific Primer Extension (ASPE) is a solution based, sequence specific enzymatic reaction technology that can be used to assay multiple SNPs in a single tube. The ASPE method involves two phases, an enzymatic reaction that determines the target genotype followed by a capture on solid microsphere surface for detection.

How does the allele-specific PCR principle work?

The Allele-specific PCR has the power to detect a single specific allele. Meaning, If you wish to amplify only a mutant allele, design a primer set accordingly and amplify it using this technique. Each set of specific primers is designed for each specific allele.

How can I make an allele specific oligonucleotide?

Allele specific oligonucleotides can be developed by following the assay design guidelines outlined here or by using PrimerPlex, a software tool for designing ASPE assays for multiplex analysis on xMAP based systems.

How is PCR specificity improved in SNP genotyping?

PCR specificity is improved by the introduction of a destabilizing mismatch within the 30 end of the allele-specific primers. This is a simple and inexpensive method for SNP detection that does not require PCR optimization. Alleles* Base Sequence