using Mothur to demultiplex plate prepared using RLMIDs

I am trying to switch over from the Roche tools (sfffile, sffinfo…) to using Mothur to demultiplex
some metagenomics data. However one plate was done using RLMIDs
I am not sure as to how to set up the oligo file in such a case to process the data.
There seems to be two barcode sequences perline as opossed to a single barcode
in the MIDconfig.parse file:


I am wondering what the two barcodes represent and whether it is possible to set up
Mothur to process this plate.


Dunno, you might check with Roche or your sequence provider to figure out what they are. Are they perhaps for two different samples or were barcodes mistakenly put on both ends of the amplicons?

Hi, thanks for the reply it seems that there are two sequences are because the RLMIDs are
Y adaptors and correspond to the two arms of the adaptor. I tried just using just the first set
of the barcode sequences in each line and it seems to be producing similar results to what
the Roche tools are giving me… any insights would be appreciated!

Have you looked at the actual sequences? I’d look to see if I could find which barcode it is.