Filtering fastq file using prinseq, output problem

[zillur]

Sir,
I wanted to use prinseq to filter my fastq files. After running this command it seemed everything going perfect but when loading completed 100% there was output. What I am missing??
[zillur@workstation01 Strain_GI]$ perl /home/zillur/Desktop/zillur/Tools/prinseq-lite-0.20.4/prinseq-lite.pl -verbose -fastq SRR901607_1.fastq -fastq2 SRR901607_2.fastq -min_qual_score 36 -ns_max_n 0 -noniupac -derep 124 -out_good GI_qual_fltr -out_bad null


This gave me the following output but no filtered fastq files.

[zillur@workstation01 Strain_GI]$ perl /home/zillur/Desktop/zillur/Tools/prinseq-lite-0.20.4/prinseq-lite.pl -verbose -fastq SRR901607_1.fastq -fastq2 SRR901607_2.fastq -min_qual_score 36 -ns_max_n 0 -noniupac -derep 124 -out_good GI_qual_fltr -out_bad null
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
Estimate size of input data for status report (this might take a while for large files)
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
done
Parse and process input data
status: 0 %perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
done
Clean up empty files
Use of uninitialized value $fhmappings in unlink at /home/zillur/Desktop/zillur/Tools/prinseq-lite-0.20.4/prinseq-lite.pl line 1833.
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
perl: warning: Setting locale failed.
perl: warning: Please check that your locale settings:
LANGUAGE = (unset),
LC_ALL = (unset),
LC_CTYPE = "UTF-8",
LANG = "en_US.UTF-8"
are supported and installed on your system.
perl: warning: Falling back to the standard locale ("C").
done
Input and filter stats:
Input sequences (file 1): 26,419,950
Input bases (file 1): 2,668,414,950
Input mean length (file 1): 101.00
Input sequences (file 2): 26,419,950
Input bases (file 2): 2,668,414,950
Input mean length (file 2): 101.00
Good sequences (pairs): 0
Good sequences (singletons file 1): 0 (0.00%)
Good sequences (singletons file 2): 0 (0.00%)
Bad sequences (file 1): 26,419,950 (100.00%)
Bad bases (file 1): 2,668,414,950
Bad mean length (file 1): 101.00
Bad sequences (file 2): 0 (0.00%)
Sequences filtered by specified parameters:
min_qual_score: 52839900
[zillur@workstation01 Strain_GI]$ ls
SRR901607_1.fastq SRR901607_2.fastq

[dpryan]

I imagine the minimum quality score setting is causing at least part of the problem. Try a more reasonable setting, like 5.

[zillur]

Thank you very much for your kind suggestions. It seemed fine. After filtering I was trying to map the filtered fastq files using bowtie2. Mapping seems good, but when I was trying to generate variant calls in vcc format, smalls is giving me the following error message. What should I do now.

[zillur@workstation01 Strain_NaN_Hs_2011]$ nohup samtools mpileup -uf /home/zillur/Desktop/zillur/phd/Apecomplexan/B_microti/Reference/B_microti.fasta Nan_qual_trm.sorted.bam | bcftools view -O u- > Nan_qual_trm.bcf
nohup: ignoring input and redirecting stderr to stdout
Failed to open -: unknown file type
[zillur@workstation01 Strain_NaN_Hs_2011]$

** which file is unknown? The sorted bam?

[dpryan]

It's complaining about the pipe. Note that nohup mentions that it's redirecting stderr to stdout, which could seriously break things. Just don't use nohup and that alone may fix things.