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The authenticity of all constructs was verified by dideoxy sequencing using ALF sequencer (Amersham Pharmacia Biotech, Piscataway, NJ).
Dideoxy sequencing was performed on an ABI 3730 Sequencer.
We could PCR-amplify 304 sequence variants; dideoxy sequencing revealed that all carried the predicted mutation (Supplementary Table 2).
The results obtained for tumor samples were 95% comparable to those obtained by dideoxy sequencing.
Dideoxy sequencing of PCR products, however, confirmed 29 out of 36 such SNPs genes with known single mutant phenotypes (Supplementary Table 2).
To evaluate false positive and negative rates, dideoxy sequencing was done for 335,420 bases interrogated by the arrays.
The results show that MUCH-AMASE can readily detect multiple infections, whereas conventional dideoxy sequencing resulted in ambiguous sequence.
Sanger dideoxy sequencing was performed by using BigDye Terminator v.3.1 Cycle Sequencing kits (Applied Biosystems, USA).
The segments amplified by PCR are transcribed to further increase the signal and to provide an abundance of single-stranded template for reverse transcriptase-mediated dideoxy sequencing.
Dideoxy sequencing of at least two independent productions of each amplified DNA template was done to confirm the validity of the mutations found.
Pyrosequencing confirmed all mutations detected by dideoxy sequencing (Table 1).
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