Demultiplexing paired end read Problem

User Support
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#import data
qiime tools import
–type ‘SampleData[PairedEndSequencesWithQuality]’
–input-path pe-64-manifest
–output-path paired-end-demux.qza
–source-format PairedEndFastqManifestPhred33

#demultiplex
qiime demux emp-paired
–m-barcodes-file metadata_4samples.tsv
–m-barcodes-column BarcodeSequence
–i-seqs paired-end-demux.qza
–o-per-sample-sequences demux
–p-rev-comp-mapping-barcodes

qiime demux summarize \
  --i-data demux_4samples.qza \
  --o-visualization demux_4samples.qzv

#error message
Plugin error from demux:

Argument to parameter 'seqs' is not a subtype of EMPPairedEndSequences.

Debug info has been saved to /tmp/qiime2-q2cli-err-ya8xuuh2.log

I could not see the logfile, actually, Could you help me figure out what’s wrong?
Thank you.

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Again, the error message
image

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How to identify the
DATA2 trimming and denoising size?
I could not view the demux.qzv file,
If I coud view it, how to identify the trim & tranc size?
Thank you so much.
qiime dada2 denoise-paired
–i-demultiplexed-seqs paired-end-demux.qza
–p-trim-left-f 0???
–p-trim-left-r 0???
–p-trunc-len-f 0???
–p-trunc-len-r 0???
–o-representative-sequences rep-seqs-dada2.qza
–o-table table-dada2.qza
–o-denoising-stats stats-dada2.qza

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image
Hi,
I put numbers randomly, however, I run successfully without any error, but without any outputfile.
Could you help me please?
THank you.

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Hi,
My story still going on.
I don't know how could I make this step happen....change sever to mac book maybe help me this moment.
However, the table.qzv SEEMs not qualified compare with the results discussed on qiime2 forum.

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image1198×1126 34 KB

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image1714×1148 68 KB

Could you please let me know, What happened?
Thank you.

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Hi,

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for the --p-sampling-depth
according to table.qzv ? frequency? or sequence account?
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image2436×1248 258 KB

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image1518×540 34.6 KB

Thank you so much,

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Hi,
The alpha compare figure looks weird.

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and I think BMI , BarcodeSequence, gender, sample-type, sequencing-run all listed in the metadata, why I got that information？
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metadata listed below.
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image1452×208 37.4 KB

Thank you so much.

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Hey @doudou2047 - looks like you made it through your initial problems if you made it this far, so I won’t address those posts.

This is because you only have 2 samples in each group.

As the warning states, those columns were ignored because they were either all unique (BarcodeSequence) or all the same value (sequencing-run, sample-type, gender).

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image1442×1142 181 KB

Hi，thank you so much for your reply.
Could you please let me know the classifier based on the Greengenes? Is there any other options? or update?
I just directly use this one? or ?
The bar-figure such like that?
level-5-bars
level-5-bars100×682 8.9 KB

Thank you,
Qiong

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Seems the file upload unsuccessfully.
This is the part of taxa-bar-plots.qzv
taxa-bar-plots.qzv (334.2 KB)

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Hi，@THERMOKARST
Even I went through this step, I still confusing about the trim size.
The numbers I just copy from one example from qiime2 forum.
Could you let me know your suggestion?
Thank you.
Qiong

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Have you read the DADA2 Documentation? This will be the best way for you to learn how this plugin works.

I am going to lock this thread now, we request that users keep it limited to one topic per thread - if you have any additional questions, please open them in new threads, individually. Thanks!

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