cutoff in classify.seqs

Commands in mothur
1

Hello Everyone,
I am kind of confused, what does cutoff in classify.seqs() mean? Is it taking the SearchScore percentage from the align.report file. I provided 80,95,99 cutoff and then I used classify.ots() command to generate OTUs and everytime it is giving me same no. of OTUs.
Also, I am using Silva db, I used silva bacteria, archaea and eukaryotes separately but to my surprise the archaea is generating more OTUs than bacteria and I am getting OTUs in eukaryotes also. How is that possible as I have only 16s sequences.

Thanks for the help!!!

2

http://www.mothur.org/wiki/Classify.seqs#cutoff

3

Thanks Dr. Schloss,
I read it before but didn’t get why I am getting same number of OTUs at different cutoffs. And also, why I am getting OTUS with eukaryotes when it is 16s. The align.report did show some matches for ex:
1-1023 151 U94371.1 1734 kmer 7.64 needleman 150 151 1 2 2 0 0 0 100.00
2-810 151 AJ243682.1 1758 kmer 6.25 needleman 1 14 1747 1758 14 0 2 1 57.14
3-774 151 AF352087.1 2059 kmer 9.03 needleman 147 151 1 5 5 0 0 0 100.00
4-772 151 L24747.1 1742 kmer 10.42 needleman 1 151 200 355 162 11 6 1 56.17
5-572 151 U51005.1 1733 kmer 6.25 needleman 150 151 1 2 2 0 0 0 100.00

for the first one the SimBtwnQuery&Template is 100 but the search score is just 7.64. I am wondering if the commands somewhere we are removing theses bad matched and then generating OTUs. I got that the cutoff is bootstraping value but where is the step in which we are removing the bad alignments.

Sorry for all the basic question but I am not getting it.

Thanks!!!

4

Well your questions are confusing too! :slight_smile: You are mixing classify.seqs and cluster or phylotype. I really don’t know what you’re trying to ask.

5

Hi Dr. Schloss,
I just want to know what does cut-off implies to in classify.seqs()

Thanks!!!

6

It corresponds to the confidence threshold. If an assignment is below the threshold, classify.seqs reports the next assignment above the threshold.