Import R1 and R2 .fastq files demultiplexed by qiime1

[daniel_rl (Daniel)]

Hi, congratulations for the work with qiime2. The facility use qiime1 to demultiplex the run according to each user and send the R1 and R2 .fastq files to us. I tried different combinations to import it ex.:

qiime tools import \
  --type 'SampleData[PairedEndSequencesWithQuality]' \
  --input-path seqs \
  --source-format QIIME1DemuxFormat \
  --output-path demux-paired-end.qza

but got a error:

"An unexpected error has occured:
  No transformation from <class
  'qiime2.plugin.model.directory_format.QIIME1DemuxDirFmt'> to <class 'q
  2_types.per_sample_sequences._format.SingleLanePerSampleSingleEndFastq
  DirFmt'> "

How is the best way to import R1 and R2 .fastq files demultiplexed by qiime1 to qiime2? thanks!

[stanislav.iablokov]

You should use manifest files:

qiime tools import \
--type 'SampleData[PairedEndSequencesWithQuality]' \
--input-path manifest.csv \
--output-path paired-end-demux.qza \
--source-format PairedEndFastqManifestPhred33

manifest.csv:
sample-id,absolute-filepath,direction
sample-1,$PWD/…/fastq/R1.fastq,forward
sample-1,$PWD/…/fastq/R2.fastq,reverse

[daniel_rl (Daniel)]

The first command you suggested worked, but the next one to demultiplex:

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

has an error:

"Plugin error from demux:

Argument to parameter ‘seqs’ is not a subtype of
EMPPairedEndSequences."

[thermokarst]

Hi @daniel_rl — looks like you were successfully able to import your already demultiplexed sequences — that means that they don’t need to be run through qiime demux (because that is for demultiplexing multiplexed samples). If you are following along with the Moving Pictures tutorial, you can skip the demux emp-* step in the “Demultiplexing Sequences” section, and proceed directly to the summarize step, to visualize your sequence quality scores. Keep us posted on your progress!

PS - thanks @stanislav.iablokov for helping @daniel_rl out!

[daniel_rl (Daniel)]

It makes sense, thank you for the answer. My only doubt is about how to combine the R1 and R2 files (did the previous command assembled the R1 and R2 contigs)? thanks

[thermokarst]

Hi @daniel_rl - qiime demux emp-* does not combine (or merge) the forward and reverse reads. If you decide to use q2-dada2, the DADA2 protocol will perform the merging for you. I believe q2-deblur only operates on the forward reads, even if you provide paired-end sequences - so merging isn’t necessary here. Either way you go, you should be all set!

Keep us posted on your progress!