align.seqs_output file_ unique.flip.accnos

Hi,

After running align.seqs command I am getting 3 output files : …unique align , …unique.align.report and …unique flip accnos:

Here is the log:

mothur > align.seqs(fasta=Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.fasta, reference=silva.bacteria.fasta, processors=2)

Using 2 processors.

Reading in the silva.bacteria.fasta template sequences… DONE.
It took 33 to read 14956 sequences.
Aligning sequences from Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.fasta …

Reading in the silva.bacteria.fasta template sequences… DONE.
It took 41 to read 14956 sequences.
Some of you sequences generated alignments that eliminated too many bases, a list is provided in Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.flip.accnos. If you set the flip parameter to true mothur will try aligning the reverse compliment as well.
It took 106 secs to align 10030 sequences.


Output File Names: Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.align Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.align.report Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.flip.accnos
mothur > summary.seqs(fasta=Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.align, name=Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.names)

Using 2 processors.

Start End NBases Ambigs Polymer NumSeqs
Minimum: 0 0 0 0 1 1
2.5%-tile: 1044 5411 245 0 3 8617
25%-tile: 1044 5415 247 0 5 86170
Median: 1044 5429 256 0 5 172340
75%-tile: 1044 5716 265 0 5 258509
97.5%-tile: 1044 6212 269 0 5 336062
Maximum: 43116 43116 301 0 8 344678
Mean: 1252.59 5768.29 254.606 0 4.74675

of unique seqs: 10030

total # of seqs: 344678

Output File Names:
Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.summary

Well, previously in the trim seqs command I did not set the paparmeter Flip=T since I have only one direction of sequencing (5->3):

mothur > trim.seqs(fasta=Mus_36_LibL_unsplitted.IP2I14U02.shhh.fasta, name=Mus_36_LibL_unsplitted.IP2I14U02.shhh.names, oligos=Mus_36_LiBL.txt, pdiffs=2, bdiffs=1, maxhomop=8, minlength=200, processors=2)

Now my question is : can I go further With my analysis even when mothur giving me this warning:

“Some of you sequences generated alignments that eliminated too many bases, a list is provided in Mus_36_LibL_unsplitted.IP2I14U02.shhh.trim.unique.flip.accnos. If you set the flip parameter to true mothur will try aligning the reverse compliment as well.”

Thanks in advance.

This looks good. When you’re sequencing everything in the same direction the warning is really just a nuisance. This happens because you sometimes will get reads that are from non-specific amplification products. These will get chucked when you set your parameters in screen.seqs.