reference alignment of ITS1 database

Theory behind mothur
1

Hi,

with special thanks to you for your great software, I have a problem in reference alignment of rumen fungi sequences.
I am using “ITSoneDB_ITS1_GB.fasta” as my reference and whatever I did, the mothur gives me the error:

[ERROR]: template is not aligned, aborting.

This is the first time I encounter this error.

This is the summary of my template:

Using 1 processors.

Start End NBases Ambigs Polymer NumSeqs
Minimum: 1 242 242 1 3 1
2.5%-tile: 1 242 242 2 7 6175
25%-tile: 1 242 242 3 8 61745
Median: 1 243 243 4 8 123490
75%-tile: 1 252 252 4 8 185235
97.5%-tile: 1 258 258 5 9 240805
Maximum: 1 258 258 5 9 246979
Mean: 1 246.49 246.49 3.91051 8.00635

of Seqs: 246979

Output File Names:
fungi.trim.contigs.good.unique.summary

It took 5 secs to summarize 246979 sequences.


would you please help me?

Thanks
Sam

2

I assume the error is coming up in align.seqs? If so, is the reference actually aligned? I’m pretty sure that a global alignment for ITS is impossible since there’s so little homology between sequences.

Pat