# Analysis of ubiome raw data
**URL:** https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797
**Category:** User Support
**Tags:** dada2
**Created:** [May 16, 2019, 1:41am UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797 "2019-05-16T01:41:03Z")
**Posts on this page:** 6
**Page:** 1
### Author:  [@kyeong\_yun](https://forum.qiime2.org/u/kyeong_yun)
#### Post date: [May 16, 2019, 1:41am UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/1 "2019-05-16T01:41:04Z")
Good morning,
I have some problem to running the DADA2 plugin.
Sample is pairedend FASTQ file from ubiome company.
Data information,
> sample\_A\_R1.fastq, sample\_A\_R2.fastq
> V4 region, 515F - 806R
I run plugins in qiime2-2018.11.
> #### [Importing]
>
> qiime tools import
> --type 'SampleData[PairedEndSequencesWithQuality]'
> --input-path $qiime2\_path/manifest.txt
> --output-path $import\_path/paired-end-demux.qza
> --input-format PairedEndFastqManifestPhred33
>
> #### [CutAdapt]
>
> qiime cutadapt trim-paired
> --i-demultiplexed-sequences $import\_path/paired-end-demux.qza
> --p-cores 4
> --p-adapter-f GTGCCAGCMGCCGCGGTAA
> --p-front-r GGACTACHVGGGTWTCTAAT
> --o-trimmed-sequences $cutadapt\_path/paired-end-demux-cutadapt.qza
>
> #### [DADA2]
>
> qiime dada2 denoise-paired
> --i-demultiplexed-seqs $import\_path/paired-end-demux-cutadapt.qza
> --p-trim-left-f $1
> --p-trunc-len-f $2
> --p-trim-left-r $3
> --p-trunc-len-r $4
> --p-n-threads 0
> --verbose
> --output-dir $qiime2\_path/DADA2
* * *
🙄 Error has occured
> 1. --p-trim-left-f 10, --p-trim-left-r 10
> --p-trunc-len-f 140, --p-trim-left-140
>
> 2. --p-trim-left-f 10, --p-trim-left-r 10
> --p-tunc-len-f 130, --p-trunc-len-r 130
>
> 3. --p-trim-left-f 10, --p-trim-left-r 10
> --p-trunc-len-f 145, --p-trunc-len-r 145
>
> ....
> miniconda2/envs/qiime2/lib/python3.5/site-packages/q2\_dada2/\_denoise.py", line 45, in \_check\_featureless\_table
> raise ValueError("No features remain after denoising. Try adjusting "
> ValueError: No features remain after denoising. Try adjusting your truncation and trim parameter settings.
>
> **Plugin error from dada2:**
> **No features remain after denoising. Try adjusting your truncation and trim parameter settings.**
So i change the option in dada2 plugin.
> --p-trim-left-f 10, --p-trim-left-r 10
> --p-trunc-len-f 150, --p-trunc-len-r 150
>
> ...
> miniconda2/envs/qiime2/lib/python3.5/site-packages/q2\_dada2/\_denoise.py", line 246, in denoise\_paired
> " and stderr to learn more." % e.returncode)
> Exception: An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more.
> **Plugin error from dada2:**
> **An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more.**
Thank you for your time.
---
### Author:  [@ChrisKeefe](https://forum.qiime2.org/u/ChrisKeefe)
#### Post date: [May 16, 2019, 5:44pm UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/2 "2019-05-16T17:44:48Z")
Hi @kyeong_yun!
Thanks for writing up a detailed post - unfortunately, your process is not entirely clear to me. A few syntax issues jump out (in 1, multiple `trim-left`s, in 1, a `--p-*tunc*-left`), but I suspect this is not cut/pasted from your actual code.
> [@kyeong\_yun](#):
>
> - –p-trim-left-f 10, --p-trim-left-r 10
> –p-trunc-len-f 140, --p-trim-left-140
> - –p-trim-left-f 10, --p-trim-left-r 10
> –p-tunc-len-f 130, --p-trunc-len-r 130
> - –p-trim-left-f 10, --p-trim-left-r 10
> –p-trunc-len-f 145, --p-trunc-len-r 145
How did you select your trim and trunc parameters? Can you share the DADA2 stats from the first run (in which the error message suggested you modify the parameters?
```auto
qiime metadata tabulate \
--m-input-file $qiime2_path/DADA2/stats-dada2.qza \
--o-visualization $qiime2_path/DADA2/stats-dada2.qzv
```
Best,
Chris
---
### Author:  [@kyeong\_yun](https://forum.qiime2.org/u/kyeong_yun)
#### Post date: [May 17, 2019, 1:31am UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/3 "2019-05-17T01:31:25Z")
Thank you for the prompt reply .
I turned dada2 plugin four times with the above options.
When i turned it like this, the file was not created.
> qiime dada2 denoise-paired
> --demultiplexed-seqs CutAdapt/paired-end-demux-cutadapt.qza
> --p-trimed-left-f 10 --p-trimed-left-r 10
> --p-trunc-len-f 140 --p-trunc-len-r 140
> --verbose
> --o--denoising-stats dada2-stats
> --o-table table
> --o-representative-sequences rep-seq
> --p-n-threads 0
The demultiplexed sequence length has a value between 147 and 150 in paired-end-demux-cutadapt.qzv, the above option are selected.
And quality declined before 7 base.
Thank you so much~ 🙂
---
### Author:  [@ChrisKeefe](https://forum.qiime2.org/u/ChrisKeefe)
#### Post date: [May 17, 2019, 8:02pm UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/4 "2019-05-17T20:02:15Z")
@kyeong_yun, sorry if my post was a bit long - if you can share the requested file it will be easier for me to help troubleshoot.
> [@ChrisKeefe](#):
>
> How did you select your trim and trunc parameters? Can you share the DADA2 stats from the first run (in which the error message suggested you modify the parameters?
>
> ```auto
> qiime metadata tabulate \
> --m-input-file $qiime2_path/DADA2/stats-dada2.qza \
> --o-visualization $qiime2_path/DADA2/stats-dada2.qzv
>
> ```
---
### Author:  [@Nicholas\_Bokulich](https://forum.qiime2.org/u/Nicholas_Bokulich)
#### Post date: [May 30, 2019, 1:48pm UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/5 "2019-05-30T13:48:19Z")
2 off-topic replies have been split into a new topic: [ubiome paired-end reads will not join!](https://forum.qiime2.org/t/ubiome-paired-end-reads-will-not-join/10025)
Please keep replies on-topic in the future.
---
### Author:  [@system](https://forum.qiime2.org/u/system)
#### Post date: [June 30, 2019, 7:48pm UTC](https://forum.qiime2.org/t/analysis-of-ubiome-raw-data/9797/6 "2019-06-30T19:48:22Z")
This topic was automatically closed 31 days after the last reply. New replies are no longer allowed.