Low-temperature cycle extension of dna with high priming...

Details Low-temperature cycle extension of dna with high priming... Low-temperature cycle extension of dna with high priming...

C - Chemistry, Metallurgy

12

P

C12P 19/34 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 2449560

The invention relates to methods for extending a primer or a pair of primers in low-temperature cycle DNA amplification for cycle sequencing and PCR. In particular, the methods contemplate the combined use of moderately thermostable DNA polymerases in the presence of a low concentration of glycerol or ethylene glycol, or the mixtures thereof, as an agent to reduce the melting temperature of DNA (that is, the temperature at which the double- strands of DNA are denatured). Predistributed reaction mixtures of a high- fidelity and high processivity DNA polymerase stable at room temperature for several weeks in ready-to-use kits are also contemplated by the invention.

L'invention concerne des méthodes d'extension d'une amorce ou d'une paire d'amorces dans l'amplification d'ADN cyclique à basse température de séquençage de cycle et PCR. Ces méthodes consistent en particulier à utiliser de façon combinée des ADN polymérases modérément thermostables en présence d'une faible concentration de glycérol ou d'éthylène glycol, ou de mélanges de ceux-ci, comme agents destinés à réduire la température de fusion de l'ADN (c'est-à-dire la température à laquelle les deux brins d'ADN sont dénaturés). L'invention concerne également des mélanges de réaction prédistribués d'ADN polymérase haute fidélité à grande capacité de traitement, stables à température ambiante pendant plusieurs semaines dans des kits prêts à l'emploi.

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