Hello, I´m new in mothur.
I´m trying to follow the miseq sop with my own samples. I have got one sample with 120000 sequence.
I´m running the command:
mothur > make.contigs(file=stability.files, flip=T, processors=8)
and I get this:
Some of your sequences generated alignments that eliminated too many bases, a list is provided in stability.trim.contigs.good.unique.flip.accnos. if the reverse compliment proved to be better it was reported.
So I think I have to get rid of the primers, barcodes, linkers and spacers of my samples so I run,
mothur > make.contigs(ffastq=test_1.fastq, rfastq=test_2.fastq, oligos=test.oligos)
And my oligos file is:
forward AATGATACGGCGACCACCGAGATCTACACTATGGTAATTGTGTGCCAGCMGCCGCGGTAA
reverse GGACTACHVGGGTWTCTAAT
barcode GTCGTGTAGCCT
linker AGTCAGTCAGCC
Spacer CAAGCAGAAGACGGCATACGAGAT
and I get a error:
[ERROR]: cannot mix paired primers and barcodes with non paired or linkers and spacers, quitting.
I think my mistake is in the oligo file,
Could somebody help with this, please?