I am currently analysing an 18S amplicon dataset and am new to the world of mothur and microbiome analyses in general.
While working my way through the MiSeq SOP, I realised that the silva.nr_v123.align file does not fully cover my amplicons. My sequences are ca. 150bp long and extend about 20 bp over the end of the alignment. So using the align.seqs command with my dataset and the silva.nr_v123. align file removed 20bp at the end of my sequences. I solved this problem by making a custom alignment file with original files downloaded from SILVA.
However, I was not able to generate files that cover my entire amplicon and that can be used in the classify.seqs command. I followed the README on the mothur blog (http://blog.mothur.org/2015/12/03/SILVA-v123-reference-files/) to make these files, but did not succeed; permission issues on the computer I am working on prevented me unfortunately from saving the fasta_mothur.eft file in the right folder.
So, would it be possible that you provide the mothur compatible silva.full_v123.fasta file? Then I can make my own taxonomy and template files to use in the classify.seqs command.
Alternatively, do you have any idea if using the classify.seqs command with the silva.nr_v123.ng.fasta (i.e. 20 bp too short) and silva.nr_v123.tax files on my dataset would significantly impact the classifications?
Kind regards. Anke.