when i wanted to create a classifier, after running several hours!! it said No matches found, log message:
Traceback (most recent call last):
File "/home/qiime2/miniconda/envs/qiime2-2017.6/lib/python3.5/site-packages/q2cli/commands.py", line 222, in __call__
results = action(**arguments)
File "<decorator-gen-126>", line 2, in extract_reads
File "/home/qiime2/miniconda/envs/qiime2-2017.6/lib/python3.5/site-packages/qiime2/sdk/action.py", line 203, in callable_wrapper
File "/home/qiime2/miniconda/envs/qiime2-2017.6/lib/python3.5/site-packages/qiime2/sdk/action.py", line 305, in _callable_executor_
output_views = callable(**view_args)
File "/home/qiime2/miniconda/envs/qiime2-2017.6/lib/python3.5/site-packages/q2_feature_classifier/_cutter.py", line 157, in extract_reads
raise RuntimeError('No matches found') from None
RuntimeError: No matches found
To help you with this error, can you provide these details?
What version of QIIME 2 are you using? You can find that by running
What is the exact command you’re running?
What reference database (including version) are you using? e.g. Greengenes, SILVA, etc.
The error message indicates that none of the reference sequences are matching the primers you’re wanting to trim at. Can you double-check that the primers you’re specifying are covered by the reference sequences you’re using?
- QIIME 2017.6 i use
qiime tools import \
--type 'FeatureData[Sequence]' \
--input-path 97_otus.fasta \
- how to check the primers ? is it not the same?
thank you very much for your help!
It looks like you might’ve copied and pasted the wrong command in your response to @jairideout - that is an
import command, but the problem you originally reported implies that you were running a
feature-classifier command. Can you please provide the exact command that produced the error message above?
hi @thermokarst sorry, I didn’t record it. these days, i’ll run my own data, then i’ll see what happened, thanks for your reply
HI @thermokarst I have run agian, it said " plugin error from feature-classifier: no matched found"
i use : 1, QIIME2 2017.8;
2, exact command: qiime feature-classifier extract-reads
3, reference database: Greengenes
4, the primers was provided by the sequencing company
thank you in adance!
It looks like the primers you are inputting are the full tailed PCR primers provided in, e.g., Nextera kits. These primer sequences contain Illumina adapter sequences on the 5’ end that do not anneal to target DNA, and do not match any known sequences in nature (they are designed this way to avoid amplification bias and other issues). Hence, the 5’ portion of these sequences will not match any sequences in Greengenes, resulting in a lack of hits.
What you need to provide to
extract-reads is instead just the portion of these sequences that actually anneal to target DNA. This is probably the 20-30 nt on the 3’ end of each sequence, but I do not know for sure since I am not familiar with these primers. You should contact Illumina and/or search primary literature to determine which portion of the sequences are the actual PCR primer portion, and which are the Illumina adapter portion and other non-target-binding portions. Please post back here when you have an answer — others using these primers and running into the same issues would love to have your help!
@BenKaehler might have additional insight.
Hope that helps!
Hi Nicholas, thank you very muc for your reply. Until now, I have no idea what’s going on, I will keep ask, of course, i’ll post here as soon as i have an answer.
hi @Nicholas_Bokulich Good news, I solved the issue, you are right, there are the whole primer for the first PCR, so I should use the final parts (e.g. 10bp) of the primers. then i run again, it worked. thank you,
have a nice day!
Hi @Nicholas_Bokulich @thermokarst
another problem appear, after taining my own classifier, then i run with my data, but there are few taxononary , most of them are “bacteria;;;;”
do you know what happened?
It is difficult to tell what is occurring based on the information that you provided, but there are three most probable answers:
- The primers that you provided as input are not specific enough. 10 bp is probably not actually the full length of the primer and could incorrectly hit other sections of the input reference sequences outside of the region you intend to target. You should determine the full sequence of the target-DNA-binding section of your PCR primers and use those for input.
- You have used an incorrect parameter somewhere. What commands did you run at each step?
- Your sequence data are low quality. What is the read length?
If these do not offer any clues to what is going wrong (especially step 1), please provide more information, and we will try to find a more specific answer.
You could also try classifying your sequences will a full-length 16S rRNA gene classifier (i.e., don’t trim during extract-reads). If you get more specific taxonomy assignments, that’s probably a good indicator that the issue is related to one of the three possibilities I noted above.
Hi @Cheng50373640, @Nicholas_Bokulich,
It looks like the primers might be Bakt_341F (CCTACGGGNGGCWGCAG) and Bakt_805R (GACTACHVGGGTATCTAATCC).
I found them in Herlemann et al. (2011, The ISME Journal 5, 1571–1579).
Hope that helps!
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