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Mutect and ampliseq IonTorrent TES
I am trying to use Mutect (v1.1.4) with ultra-deep targeted ampliseq IonProton data.
The coverage is around ~7000x and here are the mutect parameters I use:
-rf BadCigar --analysis_type MuTect --reference_sequence $REF --cosmic $COSMIC --dbsnp $DBSNP --input_file:normal $1 --input_file:tumor $2 --out $2.mutect
now, I have the following problems:
- despite the very high coverage, only literally a handful of mutations are flagged as KEEP
- if the filtering is relaxed, and I select all the NOVEL+COVERED mutations that are not in dbsnp and with a coverage of at least 400x (columns 21 and 22) and at least 40 variants (column 22), the coverage gets "adjusted" to 1000 (columns 21 and 22 always sum up to at most 1000, despite having many >7000 covered spots).
- also, with the latest filtering step, I remain with a lot of low-frequency probably spurious variants that are not present in the original exome run (also analyzed with mutect)
- I looked at the overlap between mutect and varscan2 and it is very poor (<40%)
I was wondering if there is an optimal way to use mutect with ampliseq data avoiding all these problems.