Method for the quantification of methylated dna

Details Method for the quantification of methylated dna Method for the quantification of methylated dna

C - Chemistry, Metallurgy

12

Q

C12Q 1/68 (2006.01) G06F 19/00 (2006.01)

Patent

CA 2559426

The method according to the invention concerns a method for the quantification of two different variations of a DNA sequence. Particularly, the invention relates to a quantification of methylated DNA. For this purpose, the DNA to be investigated is first converted so that cytosine is converted to uracil, while 5-methylcytosine remains unchanged. Then the converted DNA is amplified by means of a real-time PCR. However, probes are utilized, one of which is specific for the methylated status and one for the unmethylated status of the DNA. The degree of methylation of the investigated DNA can be calculated from the ratio of the signal intensities of the probes or from the Ct values. The method according to the invention is particularly suitable for the diagnosis and prognosis of cancer and other disorders associated with a change in the methylation status, as well as for the prediction of drug interactions.

L'invention concerne un procédé permettant de quantifier deux variations différentes d'une séquence d'ADN, et plus spécifiquement un procédé permettant la quantification de l'ADN méthylé. A cette fin l'ADN examiné est d'abord soumis à une conversion qui a pour effet de convertir la cytosine en uracile, tout en laissant la 5-méthylcytosine inchangée. Cet ADN converti est ensuite amplifié au moyen d'une PCR en temps réel. Ce procédé comprend toutefois l'utilisation de sondes, dont l'une est spécifique pour l'état méthylé, et l'autre, et une autre est spécifique pour l'état non méthylé de l'ADN. Le degré de méthylation de l'ADN examiné peut être calculé à partir du rapport entre les intensités des signaux des sondes, ou à partir des valeurs Ct. Ce procédé convient en particulier pour le diagnostic et le pronostic des cancers et d'autres troubles associés à une modification de l'état de méthylation, ainsi que pour le prédiction des interactions médicamenteuses.

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