Hi, Below are the files I received from a sequencing vendor.
- sample1-35_r1.fastq.gz file
- sample1-35_r2.fastq.gz file
- Map file with sample_id, barcodesequence and linkerprimer sequence.
R1 and R2 are both in the 5’-3’ orientation as raw files.
Forward primer format BARCODE-FORWARD PRIMER
Reverse primer format REVERSE PRIMER
After unzipping fastq.gz files, I am using make.contigs to get the demultiplexed contigs. (mothur > make.contigs(ffastq=sample1-35_r1.fastq, rfastq=sample1-35_r2.fastq, oligos=test.oligos))
But, I am getting the same error,
[ERROR]: cannot mix paired primers and barcodes with non paired or linkers and spacers, quitting.
It took 0 secs to process 0 sequences.
[WARNING]: your sequence names contained ‘:’. I changed them to ‘_’ to avoid problems in your downstream analysis.
I am not sure if the step is wrong or the oligos file. Any help in this regard is greatly appreciated.
Oligos file looks like this
barcode CATTCCGT sample1
barcode CATTCGTG sample2
barcode CATGGAAG sample3
barcode CATGTTGT sample4
barcode CATCACCG sample5
barcode CATGCGCA sample6
barcode ATGATTAG sample7