I was able to run standalone itsxpress successfully by changing the TMP directory. So thank you for that recommendation! I re-imported the sequences and am now running into issues when trying to run the demux command:
qiime demux emp-single --m-barcodes-column BarcodeSequence --i-seqs imported_data_forward.qza --o-per-sample-sequences demux_forward.qza --p-rev-comp-mapping-barcodes --o-error-correction-details demux_error.qza --p-no-golay-error-correction --m-barcodes-file Carey_ITS_Mappingfile.txt
This is the error:
Plugin error from demux: Mismatched sequence ids: M02149:257:000000000-CBYGW:1:1101:17084:1398 and M02149:257:000000000-CBYGW:1:1101:18572:1830
I’ve looked through other related posts and it seems like the barcode and sequence files don’t match up when using the file that was trimmed with ITSxpress. I checked how many lines are in each file per this post and it seems I’ve lost quite a bit from the sequences file after running through ITSxpress:
wc -l barcodes.fastq: 74,129,196
wc -l Undetermined_S0_L001_R1_001.fastq (raw sequences): 74,129,196
wc -l sequences.fastq (file I’m using that was run through ITSxpress): 48,958,160
Any thoughts on what might be going on and how to fix it?
Thanks!!
Chelsea