Enzyme formulations and reaction mixtures for nucleic acid amplification
AssigneeGen-Probe Incorporated
InventorsVincent J. CHABOT, Mark E. FILIPOWSKY
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Abstract
Disclosed are formulations and reaction mixtures comprising at least one polymerase, α-cyclodextrin, and polysorbate 20. Also disclosed are related methods for preparing the disclosed formulations and reaction mixtures, as well as related kits and methods for nucleic acid amplification. The formulations and reaction mixtures are useful, for example, in mitigating the inhibitory effect of sample extraction detergents on nucleic acid amplification reactions.
Claims29
An aqueous polymerase formulation comprising:
- at least one polymerase;
- α-cyclodextrin at a concentration of from about 110 mg/mL to about 40 mg/mL;
- polysorbate 20 at a concentration of from about 0.002% to about 0.05% (v/v);
- and
- at least one buffering agent.
2-6. (canceled)
8-10. (canceled)
(canceled)
An amplification reaction mixture comprising:
- at least one polymerase;
- α-cyclodextrin at a concentration of from about 5 mg/mL to about 20 mg/mL;
- polysorbate 20 at a concentration of from about 0.001% to about 0.025% (v/v);
- and
- at least one buffering agent.
18-32. (canceled)
34-35. (canceled)
38-39. (canceled)
A method for performing an amplification reaction, the method comprising:
- (a) providing a sample containing a target nucleic acid, wherein the sample is an extracted sample containing at least one detergent;
- (b) contacting the sample with an aqueous mixture comprising at least one polymerase;
- α-cyclodextrin;
- polysorbate 20;
- at least one buffering agent;
- nucleotide triphosphates suitable for performing nucleic acid amplification;
- at least one cofactor;
- and
- at least one amplification oligomer configured to amplify a target region of the target nucleic acid;
- wherein contacting the sample with the aqueous mixture provides a reaction mixture in which the α-cyclodextrin is present at a concentration of from about 5 mg/mL to about 20 mg/ml and the polysorbate 20 is present at a concentration of from about 0.001% to about 0.025% (v/v);
- and
- (c) using the reaction mixture to perform an in vitro nucleic acid amplification reaction, wherein the target nucleic acid, if present in the sample, is used as a template for generating one or more amplicons corresponding to the target region.
(canceled)
Dependencies are inferred from the claim text.