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Specifically, we identified the exon 19 mutation in sample 05 to be a 9-bp in-frame deletion encoding the amino acid deletion-substitution E746_R748del_A750, at a frequency of 11% of 1315 reads (Table 1).
The G12A substitution in sample 11 was present in 21% of 1358 reads and the T790M resistance mutation in sample 10 occurred at a frequency of 20% out of 909 reads (Table 1).
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.
Point mutation in sample NE 16 was confirmed by four independent RT-PCR reactions.
One PIK3CA mutation in sample P72_pos was called at 1% allele frequency by NGS, and this mutation was therefore unlikely to be detected by Sequenom analysis.
Nevertheless, the fact that PNA-clamp real-time PCR confirmed the L858R mutation in sample 2768 suggests that castPCR technology does not result in false positive findings.
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The FRET/MCA was used to detect the L1014S and L1014F kdr mutation in samples of the WHO bioassay (collected in 2004 2006).
Linkage analysis was used to determine the candidate region for the mutation in samples from 11 affected members and 14 unaffected members.
The two somatic mutations in sample 1 were homozygous reference in both parents.
Direct sequencing seems unable to provide satisfactory results for detection of EGFR mutations in samples containing a mixture of mutated and wild-type DNA.
Contrasting significance had already been reported for the KRAS-G13D mutation, found in sample ID_5443 [ 18].
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