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[Mutation sampling in such experiments is nearly unbiased because genetic drift overwhelms natural selection during the extreme bottlenecks; without selection, all non-lethal mutations can fix with roughly equal probability, regardless of their deleterious, advantageous or neutral effects [ 22- 24].
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We also verified the most important novel data, lower baseline Ki67 in mutation samples, in a large, publicly available data set.
The resistance mutation samples were collected in the US and Canada during 1998 2005, and included a portion of US samples from a previously reported surveillance study [19].
Compared with direct sequencing, in which mutant DNA needs to compose 25% or more of the total DNA to easily detect a mutation, the SNaPshot and sizing assays can detect mutations in samples in which mutant DNA composes 1.56% to 12.5% and 1.56% to 6.25% of the total DNA, respectively.
The SHH mutation in Sample 1, FLT3 mutation in Sample 3, and DMBT- 1 mutation in Sample 4 were validated using PCR and capillary sequencing.
We analyzed these two samples with a real-time PCR-PNA clamp assay that confirmed the mutation in sample 2768 but not in sample 3000.
Samples with a mutation in c108+c59 or c108+c51 of dhfr but no additional mutations were classified as double mutations; samples with mutations in c51+c59+c108 as triple mutations; samples with mutations in c51+c59+c108 of dhfr and c437 of dhps were considered quadruple mutations; and samples with mutations in c51+c59+c108 of dhfr and c437+c540 of dhps as quintuple mutations.
Point mutation in sample NE 16 was confirmed by four independent RT-PCR reactions.
This, in turn, allows for the molecular selection of mutated samples and makes mutation detection in premalignant samples feasible.
The two somatic mutations in sample 1 were homozygous reference in both parents.
In this study, we assessed the feasibility of detection of EGFR mutations in samples of pleural effusion fluid.
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