tag barcode

Commands in mothur
1

Hi everyone, I am new in mothur so I have a question from my raw data:

I going to start to analyse my two fastq files (forward and reverse) with the reads of a miseq run of a mix of close to 300 experimental samples, with every sample having a different tag barcode in the forward and reverse primer to assign to their respective experiment. So my question is how I can classify my sequences to the proper experiment (group)? I was thinking in using in the make.contings command, the oligos options with the table of every tag to which experiment belongs, but I don’t know if in the new file will have attached for all the sequences its respective group, so I can use that for the next analyses steps.

Any suggestions?

Thank you for the help,
Sebastián

2

Welcome to the mothur community! The make.contigs command is a good place to start, http://www.mothur.org/wiki/Make.contigs. You want to set up your oligos file like:

primer yourForwardPrimer yourReversePrimer optionalPrimerName

Example:
primer ATTAGAWACCCBDGTAGTCC CCCGTCAATTCMTTTRAGT V5


BARCODE yourForwardBarcode yourReverseBarcode yourSampleName

Example:
BARCODE ccaac aacca F01R2B

Mothur will remove the primers and barcodes, assemble the reads and create a fasta and group file, http://www.mothur.org/wiki/Group_file, which assigns your sequences to a sample.