I'm using the QIIME1 scripts extract_barcodes.py to extract barcodes from a dual-indexed MiSeq library, and then demultiplex my reads using split_libraries_fastq.py. I have been having trouble correctly using a couple of parameters relating to reverse complementing barcodes.
When extracting the barcodes with extract_barcodes.py, I use
--rev_comp_bc1 (as directed by the individual in charge of sequencing). Then when I demultiplex the reads with split_libraries_fastq.py, sometimes I need to include the parameter
--rev_comp_mapping_barcodes to get the script to work, and sometimes I don't. I'm not sure why, unless the barcodes I'm receiving in my mapping files are sometimes written in different orientations (I don't write them).
I think the parameter
--rev_comp_bc1 is self-explanatory, but could someone explain what
--rev_comp_mapping_barcodes does and when to use it?
To make things simple we can assume that the two barcodes are written from 5' to 3' in the mapping file.