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Old 08-07-2013, 01:19 PM   #1
shbh
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Default Softclipping of primers for Fluidigm / Raindance PCR based methods

When performing target enrichment of large panels of genes using a PCR based method such as Fluidigm or Raindance, primers of varying lengths are used to amplify targets. I am looking for a tool to remove the known primer sequences. I know Illumina has incorporated such a tool in the miseq reporter for truseq custom amplicons, but I have not been able to find a similar tool to incorporate into the standard CASAVA (or other) pipelines. I contacted Illumina about this, but, of course, they were no help. I have also attempted to use cutadapt, but it is not working. And, since the primer sequences are of varying length, I cannot simply remove X number of bases at the beginning of the read.
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Old 08-07-2013, 06:13 PM   #2
frozenlyse
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In my pipeline for MiSeq amplicon analysis I align the reads keeping the primers on, and then just ignore the primer areas in downstream analysis.
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Old 03-03-2015, 01:23 PM   #3
bioinfosm
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Quote:
Originally Posted by frozenlyse View Post
In my pipeline for MiSeq amplicon analysis I align the reads keeping the primers on, and then just ignore the primer areas in downstream analysis.
In some cases, there might be overlapping primers and amplicons where simply ignoring the areas wouldnt be a good idea. In those cases, trimming is still important as the variant allele frequencies might be inaccurate, but seems there are no ready to use tools out there!
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