I tried to search for it, but am not sure whether MOTHUR allows variable length barcodes to be included in the same oligos file? Also, if my forward primer is denegerate and i include it as part of the barcode sequence, would it allow the degenracy in the barcode during trim.seqs? I know that i could use the degenrate portion of the sequence as a partial forward primer, and use only the non-degenerate potion of the primer as a part of barcode to avoid this.
Yup, you can have mixed length barcodes and degeneracies in your primers. But you don’t want to have degeneracies in your barcode. You should be able to have multiple barcodes for the same sample.