DADA2 No reads pass filter

Hi everyone!
I am trying to use dada2 on forward and reverse reads which have been removed primers, barcodes. My primers are 515f-907R, for 16s V4-V5.
However, when i used denoise-paired, i got error mistakes. I also try other p-trunc-len-f/r 150 190 200 210 220 etc, but the machine still display “No read pass filter”.Here are my demux.qzv filegcb-demux.qzv (291.3 KB)
i am very dispirited, hope someone can help me!
qiime dada2 denoise-paired
--i-demultiplexed-seqs gcb-paired-end-demux.qza
--p-trim-left-f 0
--p-trim-left-r 0
--p-trunc-len-f 210
--p-trunc-len-r 210
--o-table gcb-table-1.0
--o-representative-sequences gcb-rep-seqs-1.0
--output-dir dada2-1.92
--p-n-threads 16
--verbose

Running external command line application(s). This may print messages to stdout and/or stderr.

The command(s) being run are below. These commands cannot be manually re-run as they will depend on temporary files that no longer exist.

Command: run_dada_paired.R /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/forward /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/reverse /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/output.tsv.biom /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/track.tsv /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/filt_f /mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/filt_r 210 210 0 0 2.0 2 consensus 1.0 16 1000000

Fatal error: cannot create 'R_TempDir'

Traceback (most recent call last):

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/q2_dada2/_denoise.py", line 231, in denoise_paired

run_commands([cmd])

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/q2_dada2/_denoise.py", line 36, in run_commands

subprocess.run(cmd, check=True)

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/subprocess.py", line 398, in run

output=stdout, stderr=stderr)

subprocess.CalledProcessError: Command '['run_dada_paired.R', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/forward', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/reverse', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/output.tsv.biom', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/track.tsv', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/filt_f', '/mnt/Amplicon/wqsu/xingancharcoal/tmp6r7j54g0/filt_r', '210', '210', '0', '0', '2.0', '2', 'consensus', '1.0', '16', '1000000']' returned non-zero exit status 2

During handling of the above exception, another exception occurred:

Traceback (most recent call last):

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/q2cli/commands.py", line 274, in call

results = action(**arguments)

File "<decorator-gen-436>", line 2, in denoise_paired

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/qiime2/sdk/action.py", line 231, in bound_callable

output_types, provenance)

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/qiime2/sdk/action.py", line 362, in callable_executor

output_views = self._callable(**view_args)

File "/home/wqsu/anaconda3/envs/qiime2-2018.6/lib/python3.5/site-packages/q2_dada2/_denoise.py", line 242, in denoise_paired

% (trunc_len_f, trunc_len_r))

ValueError: No reads passed the filter. trunc_len_f (210) or trunc_len_r (210) may be individually longer than read lengths, or trunc_len_f + trunc_len_r may be shorter than the length of the amplicon + 20 nucleotides (the length of the overlap). Alternatively, other arguments (such as max_ee or trunc_q) may be preventing reads from passing the filter.

Plugin error from dada2:

No reads passed the filter. trunc_len_f (210) or trunc_len_r (210) may be individually longer than read lengths, or trunc_len_f + trunc_len_r may be shorter than the length of the amplicon + 20 nucleotides (the length of the overlap). Alternatively, other arguments (such as max_ee or trunc_q) may be preventing reads from passing the filter.

See above for debug info.

Hi! Maybe you should try to truncate it at 180 nt instead.

ok Thank you i will try now!

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