An optimal procedure for isolation of mutant mitochondrial...

C - Chemistry – Metallurgy – 12 – Q

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C12Q 1/68 (2006.01)

Patent

CA 2244101

A sample preparation procedure for mitochondrial (mt) DNA analysis is described. The present method for isolating mtDNA uses sedimentation techniques for separating erythrocytes from lymphocytes and platelets (crude buffy coat fraction), followed by DNA extraction from the crude buffy coat fraction by boiling in water. This approach improves the yield of mutant DNA, enhancing the sensitivity of subsequent mutation interrogation techniques and allowing for meaningful statistical treatment of the degree of heteroplasmy within the mitochondrial DNA.

L'invention concerne un procédé de préparation de spécimen dans le but d'effectuer l'analyse d'ADN mitochondrial (mt). Ce procédé d'isolation d'ADNmt met en application des techniques de sédimentation permettant de séparer les erythrocytes des lymphocytes et des plaquettes (fraction brute de couche leuco-plaquettaire), suivies par l'extraction de l'ADN de la fraction brute de couche leuco-plaquettaire par ébullition dans de l'eau. Ceci permet d'obtenir davantage d'ADN mutant, d'augmenter la sensibilité de techniques subséquentes d'interrogation sur la mutation et de réaliser un traitement statistique significatif du degré d'hétéroplasmie à l'intérieur de l'ADN mitochondrial.

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