Patent ID: 11898201
Assignee: BIO-RAD LABORATORIES, INC.
Field: Biotechnology (Chemistry)
Classification: CPC C | IPC C

Claim 8:
9. A method for quantitating a frequency of wild-type and mutant target nucleic acid fragments in a nucleic acid sample, the method comprising:
(a) forming a plurality of mixture partitions of claim 1;
(b) incubating the mixture partitions under thermal cycling conditions suitable for amplification of the target DNA template molecules by a polymerase chain reaction, wherein the thermal cycling conditions comprise a first set of temperature cycles and a second set of temperature cycles, wherein the second set of temperature cycles comprises an annealing temperature that is at least 5° C. higher or lower than an annealing temperature of the first set of temperature cycles;
(c) detecting the presence or absence of amplified target DNA template in the mixture partitions, wherein the detecting comprises, determining:
i) a number of wild-type mixture partitions comprising amplified target DNA template consisting of wild-type target sequence;
ii) a number of mutant mixture partitions comprising amplified target DNA template consisting of mutant target sequence; and
iii) a number of double-positive mixture partitions comprising amplified target DNA comprising mutant and wild-type target sequence; and
(d) determining from the number of wild-type, mutant, and double-positive mixture partitions the frequency of wild-type and mutant target nucleic acid fragments in the nucleic acid sample.