Allélotypage

The allélotypage is a technique of Molecular biology allowing the determination of the relative frequencies of the Allèle S.

Principle

The principle consists in amplifying by PCR sequences Microsatellites, dispersed on the Génome and located between the coding sequences of genes or in large Introns. They are highly polymorphic in a population, but very stable at a given individual.

Application

This technique allows phenotypical classification some solid tumors. After the amplification of the microsatellites selected by PCR, the amplifiats are separate on freezing denaturing using an automated sequencer. This apparatus allows, by a technique of marking not Radioactive of the products of PCR, to determine their size, therefore the number of repetitions of the sequence Microsatellite, and to quantify them. A Microsatellite is known as informative so after migration, it appairait two peaks of which each one represents the amplification of the sequence Microsatellite carried by each Allèle: that means that the number of repetitions of the Microsatellite is different between the 2 alleles. The Microsatellite is known as not informative so only one peak is observed: that means that the number of repetitions of the Microsatellite is identical between the 2 alleles. The ratio height of the two peaks of each microsatellite amplified starting from DNA of the tumoral cells is compared with the report/ratio of the peaks obtained starting from nontumoral cells of the same individual. This makes it possible to calculate the percentage of the rehandling allelic according to the formula:

Percentage of the rehandling allelic = ((b/a) Fabric tumoral - (b/a) Fabric nontumoral)/(b/a) Fabric nontumoral
has and B representing the height of the two peaks of each amplified sequence Microsatellite.

This percentage of the rehandling allelic translates the tumoral proportion of fabric having a deterioration with the Locus considered: one speaks about rehandling allelic. A variation of the relationship between tumoral fabric and nontumoral fabric higher than a value threshold calculated starting from a pilot population is regarded as a deterioration. However, this method does not make it possible to distinguish a Délétion from a allelic Amplification.

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