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very weird alignment and results #5
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Hi,
Thanks for using HmmUFOtu. Most of this type of problem is you are using an
older versio of HmmUFOtu, and failed to tell the correct strand-ness of the
reads. In other words, your fwd/rev reads were in wrong orientation (due to
primer mis-naming).
Try to upgrade to the latest HmmUFOtu, and either let it "guess" the
correct strand (by default), or manually tell it the strand orientation.
Check -h help message for details.
Best
Qi
…On Thu, Aug 8, 2019 at 12:41 PM marlenec ***@***.***> wrote:
Hi,
I am new using this software. I have an Illumina sequencing of V4 sequence
16S rDNA. I trimmed and assembled the sequences using DADA2.
I wanted to use hmmufotu on DADA2's ESVs to compare the taxonomic
classification of the two softwares, and get a phylogenetic tree for my
ESVs.
Here is the beginning of the input :
Otu1
AGCAGTGGGGAATAT[...]CAAACAGGATTAGATACCCTGGTA
Otu2
AGCAGTGGGGAATAT[...]GGATTAGATACCCTGGTA
Otu3
AGCAGTGGGGAATAT[...]AGGATTAGATACCCTGGTA
After running hmmufotu and hmmufotu-sum on the file using GreenGenes
(v13.8) species-level (97% OTU) reference + GTR DNA model that is
recommanded, I got a very weird alignment were almost all bases of most of
my ESVs (here they are called Otus, but it is only for compatibility with
other software) are replaced by gaps '-'
Example
5360
DBName=Archive/GTR/gg_97_otus_GTR;Taxonomy="k__Bacteria;p__Firmicutes;c__Clostridia;o__Clostridiales;f__Lachnospiraceae;g__[Ruminococcus];s__gnavus";AnnoDist=0.64307999999999976;ReadCount=71;SampleHits=1
--------------[...]----------------------------------------------
--------------------------TACCAGGGCTACACACGTGCT----
---[...]-----
[...] are were I reduced the sequence length for the purpose of this
message)
And the classification is also very weird, with only 7% of agreement at
Phylum level with classification on SILVA database using RDP classifier.
Perhaps I am using it wrong ? I know this software is supposed to be used
on raw reads, but I thought it would have been great to compare its
classification resolution with RDP classifier.
Thanks you in advance!
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Hi,
Thank you for your answer. I tried with -t option and it worked.
Thanks,
Marlène
…-----------------------------------------------
Dr Marlène Chiarello
Dpt of Biology
University of Mississippi
Oxford MS, 38655
USA
Cell (France): +33 (0)6-72-13-72-12
Cell (USA): +1 (662) 202-7433
Le 8 août 2019 à 13:12, Qi ***@***.***> a écrit :
Hi,
Thanks for using HmmUFOtu. Most of this type of problem is you are using an
older versio of HmmUFOtu, and failed to tell the correct strand-ness of the
reads. In other words, your fwd/rev reads were in wrong orientation (due to
primer mis-naming).
Try to upgrade to the latest HmmUFOtu, and either let it "guess" the
correct strand (by default), or manually tell it the strand orientation.
Check -h help message for details.
Best
Qi
On Thu, Aug 8, 2019 at 12:41 PM marlenec ***@***.***> wrote:
> Hi,
> I am new using this software. I have an Illumina sequencing of V4 sequence
> 16S rDNA. I trimmed and assembled the sequences using DADA2.
> I wanted to use hmmufotu on DADA2's ESVs to compare the taxonomic
> classification of the two softwares, and get a phylogenetic tree for my
> ESVs.
>
> Here is the beginning of the input :
>
> Otu1
> AGCAGTGGGGAATAT[...]CAAACAGGATTAGATACCCTGGTA
> Otu2
> AGCAGTGGGGAATAT[...]GGATTAGATACCCTGGTA
> Otu3
> AGCAGTGGGGAATAT[...]AGGATTAGATACCCTGGTA
>
> After running hmmufotu and hmmufotu-sum on the file using GreenGenes
> (v13.8) species-level (97% OTU) reference + GTR DNA model that is
> recommanded, I got a very weird alignment were almost all bases of most of
> my ESVs (here they are called Otus, but it is only for compatibility with
> other software) are replaced by gaps '-'
>
> Example
>
> 5360
> DBName=Archive/GTR/gg_97_otus_GTR;Taxonomy="k__Bacteria;p__Firmicutes;c__Clostridia;o__Clostridiales;f__Lachnospiraceae;g__[Ruminococcus];s__gnavus";AnnoDist=0.64307999999999976;ReadCount=71;SampleHits=1
> --------------[...]----------------------------------------------
> --------------------------TACCAGGGCTACACACGTGCT----
> ---[...]-----
>
> [...] are were I reduced the sequence length for the purpose of this
> message)
>
> And the classification is also very weird, with only 7% of agreement at
> Phylum level with classification on SILVA database using RDP classifier.
>
> Perhaps I am using it wrong ? I know this software is supposed to be used
> on raw reads, but I thought it would have been great to compare its
> classification resolution with RDP classifier.
>
> Thanks you in advance!
>
> —
> You are receiving this because you are subscribed to this thread.
> Reply to this email directly, view it on GitHub
> <#5?email_source=notifications&email_token=ABXZ44L7GEVEDY7PKMO4T3LQDREEDA5CNFSM4IKMPD5KYY3PNVWWK3TUL52HS4DFUVEXG43VMWVGG33NNVSW45C7NFSM4HEGRK3Q>,
> or mute the thread
> <https://github.com/notifications/unsubscribe-auth/ABXZ44JZ7DE6WSQCVDG5HE3QDREEDANCNFSM4IKMPD5A>
> .
>
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Hi,
I am new using this software. I have an Illumina sequencing of V4 sequence 16S rDNA. I trimmed and assembled the sequences using DADA2.
I wanted to use hmmufotu on DADA2's ESVs to compare the taxonomic classification of the two softwares, and get a phylogenetic tree for my ESVs.
Here is the beginning of the input :
After running hmmufotu and hmmufotu-sum on the file using GreenGenes (v13.8) species-level (97% OTU) reference + GTR DNA model that is recommanded, I got a very weird alignment were almost all bases of most of my ESVs (here they are called Otus, but it is only for compatibility with other software) are replaced by gaps '-'
Example
[...] are were I reduced the sequence length for the purpose of this message)
And the classification is also very weird, with only 7% of agreement at Phylum level with classification on SILVA database using RDP classifier.
Perhaps I am using it wrong ? I know this software is supposed to be used on raw reads, but I thought it would have been great to compare its classification resolution with RDP classifier.
Thanks you in advance!
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