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Amplicon Sequencing Data Analysis

I am looking for a tutorial and journal of amplicon sequencing data analysis. Is there any specific free website to teach beginner the pipeline of data analysis and what should we report in our result. Any journal recommended?

sequencing sequence amplicon data analysis
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updated by WouterDeCoster WouterDeCoster • written by t_stranger • t_stranger

1 answer

I would suggest you to follow the GATK best practices. Amplicon sequencing mainly follows the guidelines for exome sequencing/panel sequencing, with two important exceptions:

  • You shouldn't mark duplicates because all your reads are duplicates
  • You should softmask your primer sequences: you can't get valid variant calls in those
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written by WouterDeCoster WouterDeCoster

Thanks for your suggestion. May I know why we should not mark the duplicates before variant call? Because I read some resource they included it in the pipeline.

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written by t_stranger • t_stranger

Since you are doing amplicon sequencing all reads mapping to the same location will mimic duplicates so you will only keep one read per amplicon per direction.

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written by WouterDeCoster WouterDeCoster

Can one use BAMClipper instead of softmasking primers?

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written by lamteva.vera lamteva.vera

Assuming you mean bamclipper which removes primers using soft-clipping, the answer would be yes, although I haven't tested the software I assume it works as described in their README.

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written by WouterDeCoster WouterDeCoster

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