Import demultiplexed paired-end fastq


I am trying to analysis gut microbes using demultiplexed paired-end fastq data.

This is what I did:

  1. I combined all data into one R1 fastq and one R2 fastq

  2. I used this command below to import data to qiime2 and produced demux-paired-end.qza
    qiime tools import
    –type ‘SampleData[PairedEndSequencesWithQuality]’
    –input-path casava-18-paired-end-demultiplexed
    –source-format CasavaOneEightSingleLanePerSampleDirFmt
    –output-path demux-paired-end.qza

  3. then I used this command below to produce the summary tables for forward and reverse reads.
    qiime demux summarize
    –i-data demux.qza
    –o-visualization demux.qzv

The next step should be trimming the data and this is where I got confused.

Since I have two summary tables forward and reverse, then how should I trim the data,
If the high quality for forward is between 1-300, and reverse is between 20-240. Is that means I need to trim the demux.qza from 20 to 240?

In qiime2 tutorial: Fecal microbiota transplant study. The data were imported, summarized, denoised separately. I was just wondering is this the correct way that I suppose to do for my analyse? Should I import the forward and reverse read separately as single read, then merge them later?

Thanks for your help

Yes, if you are analyzing multiple sequencing runs. Follow the steps in that tutorial.

No, import as you are doing above. Just import data from separate runs separately, do not merge into single files.

See the FMT study tutorial — dada2 has different trim settings for forward and reverse reads, so you can give different trim settings tailored to the needs of each read.

I hope that helps clarify!

Hello Nicholas

Thanks for your information. Another question is where does qiime2 store the error message. I was running the denoising analyse last night, and the run did was finished but nothing produced. I believe, if there is any problems occurred, there much be an error message generated right? So where am I able to find it?

Thanks for your help

If there is an error, QIIME 2 will report it in the standard output. In other words, it will appear right in the terminal. If you do not see any message, the command is probably still running (successful commands usually also produce a message).

Hello Nicholas

Thanks for your information.

Bowen :slight_smile:

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