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To study such extremely rare mutations requires "a very large denominator," or huge numbers of subjects, Roden says.
First, an excess of non-synonymous mutations requires a long time to accumulate [ 47].
In practice, any test that identifies driver genes by recurrent mutations requires a good estimate of the passenger mutation probability q.
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However, microarrays for diagnosis of disease-causing genetic mutations require a 100% assay success rate, suggesting that an alternative strategy for assay development is required.
Thus, to identify heterozygous mutations required a different approach to library construction and data analysis than has been used previously.
A structural understanding of the CFH mutations required a molecular structure for CFH, however intact CFH has not been crystallised to date.
Determination of the EGFR expression and the presence of mutations require a tumour biopsy, which is not possible to collect in all situations.
The fitness cost conferred by these last two mutations required a background of altered 23S rRNA however, as their transformation failed to affect the growth of S. pneumoniae 1974 WT (data not shown).
Accurate detection of LKB1 mutations require a combination of deep sequencing techniques including Sanger sequencing to detect point mutations and loss of heterozygosity as well as exon sequencing by Multiplex ligation-dependent probe amplification (Ji et al, 2007; Wingo et al, 2009).
Like the identification of NCO GCs, the identification of spontaneous mutations required a minimal coverage of 10, an allele frequency higher than the cutoff to distinguish between homozygous and heterozygous positions (as defined for the tetrad analysis), and equivalent evidence for the non-mutant allele in the other samples.
Moreover, the Ile219->Lys mutation requires a single nucleotide change, which also increases the probability of such a mutation to occur.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com