Answers.Body: <p>The <a href="http://www.htslib.org/doc/samtools-0.1.19.html" rel="nofollow noreferrer">samtools v0.1.19 manual</a> confirms that there is no <code>--region</code> option.  In that version there is only the <code>-r</code> argument to pass in the targeted region.  If you update to a newer version of samtools this should execute correctly</p>
  
Answers.ContentLicense: CC BY-SA 3.0
Answers.CreationDate: 2017-12-27T17:50:03.753Z
Answers.Text: It worked after I reinstalled all the tools. The main problem was the mpileup error where --region was not an option. Cannot trust some of these yum packages as they are much older versions. Thank you so much for helping me with this issue.
Answers.UserDisplayName: Drops
Answers.ContentLicense: CC BY-SA 3.0
Answers.CreationDate: 2017-12-27T17:19:31.107Z
Answers.LastActivityDate: 2017-12-27T17:19:31.107Z
Answers.UserDisplayName: Bioathlete
Body: <p>This is the error that I am getting:</p>
  
  <pre><code>launch_varscan.pl: Creating a pileup /home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.mpileup
  removed ‘/home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.1239.mpileup’
  launch_varscan.pl: Running mpileup:
    echo "NC_001416.1
  " | xargs -P 1 -n 1 -I {} sh -c 'echo "MPileup on {}" &gt;&amp;2; samtools mpileup -f lambda/reference/reference.fasta 
  -Q 0 -B  --region "{}" /home/user/lyve-SET-1.1.4f/lambda/bam/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam &gt; 
  /home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.$$.mpileup'
  MPileup on NC_001416.1
  mpileup: unrecognized option '--region'
  open: No such file or directory
  [mpileup] failed to open NC_001416.1: No such file or directory
  launch_varscan.pl: main::mpileup: ERROR with xargs and samtools mpileup
  Stopped at /home/arahat/lyve-SET-1.1.4f/scripts/launch_varscan.pl line 106.
  QSUB ERROR
  31488
  launch_set.pl: Schedule::SGELK::command: ERROR with command: Inappropriate ioctl for device
    /usr/bin/perl /home/user/lyve-SET-1.1.4f/lambda/log/SGELK/qsub.2705.pl
  Stopped at /home/user/lyve-SET-1.1.4f/scripts/../lib/Schedule/SGELK.pm line 661.
  set_test.pl: ERROR: died while trying to run COMMAND
    launch_set.pl lambda --numcpus 1 --numnodes 20
  set_test.pl: See if you have the correct software installed for this command.
  6400 at /home/arahat/lyve-SET-1.1.4f/scripts/set_test.pl line 94.
  make: *** [test] Error 25
  </code></pre>
  
  <p>I think the main problem is the "mpileup: unrecognized option --region", which seems to be blank. In the beginning of the block, it says creating pileup and then it immediately removes the file, probably to create the pipeline file.</p>
  
  <p>I cannot change the code for Lyve-SET pipelining or create shell scripts for the pipeline because I am only configuring my server for use with the genome sequencing pipeline. Would this be a problem with the data set or some tool that I need to install? Any insight would be appreciated.</p>
  
Comments.ContentLicense: CC BY-SA 3.0
Comments.CreationDate: 2017-12-27T16:56:18.297Z
Comments.Text: What version of samtools mpileup to you have installed?  What version of varscan are you using?  Did you install the two tools separately or did varscan come with samtools?  I have version 1.3.1 and there is a `--region` option for mpileup.
Comments.UserDisplayName: Bioathlete
Comments.ContentLicense: CC BY-SA 3.0
Comments.CreationDate: 2017-12-27T16:59:25.427Z
Comments.Text: Have you tried running `samtools mpileup -f lambda/reference/reference.fasta 
  -Q 0 -B  --region NC_001416.1 /home/user/lyve-SET-1.1.4f/lambda/bam/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam`  directly to see if the command works outside the context of varscan?
Comments.UserDisplayName: Bioathlete
Comments.ContentLicense: CC BY-SA 3.0
Comments.CreationDate: 2017-12-27T17:06:34.500Z
Comments.Text: I installed samtools version 0.1.19 using yum install. Varscan was installed separately through make install from Lyve-SET. I did not try to run samtools mpileup separately because all the commands are implemented through Lyve-SET, which is how it will be run by the user. I will try reinstalling varscan and running the samtools mpileup command separately. Thank you for the help.
Comments.UserDisplayName: Drops
Comments.ContentLicense: CC BY-SA 3.0
Comments.CreationDate: 2017-12-27T17:15:30.150Z
Comments.Text: I would suggest installing a more recent version of samtools.  That version is from 2013 is likely is not compatible with the version of varscan you installed
Comments.UserDisplayName: Bioathlete
Comments.ContentLicense: CC BY-SA 3.0
Comments.CreationDate: 2017-12-27T17:21:21.080Z
Comments.Text: Yup. I noticed that yum patches but does not update the packages so I am manually installing the newest versions of all the tools specified in Lyve-SET. Thank you for bringing this to my attention. Hopefully it works after I reinstall.
Comments.UserDisplayName: Drops
Tags: <samtools><mpileup>
Title: I am getting this error when running a test data set for Lyve-SET on RHEL 7. Is this an MPileup error or something else?
ThreadText: # I am getting this error when running a test data set for Lyve-SET on RHEL 7. Is this an MPileup error or something else?
  
  (Asked by: username_0 on 2017-12-27)
  
  This is the error that I am getting:
  
  ```
  launch_varscan.pl: Creating a pileup /home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.mpileup
  removed ‘/home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.1239.mpileup’
  launch_varscan.pl: Running mpileup:
    echo "NC_001416.1
  " | xargs -P 1 -n 1 -I {} sh -c 'echo "MPileup on {}" >&2; samtools mpileup -f lambda/reference/reference.fasta 
  -Q 0 -B  --region "{}" /home/user/lyve-SET-1.1.4f/lambda/bam/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam > 
  /home/user/lyve-SET-1.1.4f/lambda/tmp/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam.$$.mpileup'
  MPileup on NC_001416.1
  mpileup: unrecognized option '--region'
  open: No such file or directory
  [mpileup] failed to open NC_001416.1: No such file or directory
  launch_varscan.pl: main::mpileup: ERROR with xargs and samtools mpileup
  Stopped at /home/arahat/lyve-SET-1.1.4f/scripts/launch_varscan.pl line 106.
  QSUB ERROR
  31488
  launch_set.pl: Schedule::SGELK::command: ERROR with command: Inappropriate ioctl for device
    /usr/bin/perl /home/user/lyve-SET-1.1.4f/lambda/log/SGELK/qsub.2705.pl
  Stopped at /home/user/lyve-SET-1.1.4f/scripts/../lib/Schedule/SGELK.pm line 661.
  set_test.pl: ERROR: died while trying to run COMMAND
    launch_set.pl lambda --numcpus 1 --numnodes 20
  set_test.pl: See if you have the correct software installed for this command.
  6400 at /home/arahat/lyve-SET-1.1.4f/scripts/set_test.pl line 94.
  make: *** [test] Error 25
  
  ```
  
  I think the main problem is the "mpileup: unrecognized option --region", which seems to be blank. In the beginning of the block, it says creating pileup and then it immediately removes the file, probably to create the pipeline file.
  
  I cannot change the code for Lyve-SET pipelining or create shell scripts for the pipeline because I am only configuring my server for use with the genome sequencing pipeline. Would this be a problem with the data set or some tool that I need to install? Any insight would be appreciated.
  
  **Tags:** samtools, mpileup
  
  > **Comments:**
  > **username_1**: What version of samtools mpileup to you have installed?  What version of varscan are you using?  Did you install the two tools separately or did varscan come with samtools?  I have version 1.3.1 and there is a `--region` option for mpileup.  
  > **username_1**: Have you tried running `samtools mpileup -f lambda/reference/reference.fasta  -Q 0 -B  --region NC_001416.1 /home/user/lyve-SET-1.1.4f/lambda/bam/NC001416.fasta.wgsim.fastq.gz-reference.sorted.bam`  directly to see if the command works outside the context of varscan?  
  > **username_0**: I installed samtools version 0.1.19 using yum install. Varscan was installed separately through make install from Lyve-SET. I did not try to run samtools mpileup separately because all the commands are implemented through Lyve-SET, which is how it will be run by the user. I will try reinstalling varscan and running the samtools mpileup command separately. Thank you for the help.  
  > **username_1**: I would suggest installing a more recent version of samtools.  That version is from 2013 is likely is not compatible with the version of varscan you installed  
  > **username_0**: Yup. I noticed that yum patches but does not update the packages so I am manually installing the newest versions of all the tools specified in Lyve-SET. Thank you for bringing this to my attention. Hopefully it works after I reinstall.  
  
  ---
  
  ## 1 answers
  
  ### Answer by username_1 on 2017-12-27. Score: 0 | Accepted answer
  
  The [samtools v0.1.19 manual](http://www.htslib.org/doc/samtools-0.1.19.html) confirms that there is no `--region` option. In that version there is only the `-r` argument to pass in the targeted region. If you update to a newer version of samtools this should execute correctly
  
  > **Comments:**
  > **username_0**: It worked after I reinstalled all the tools. The main problem was the mpileup error where --region was not an option. Cannot trust some of these yum packages as they are much older versions. Thank you so much for helping me with this issue.  
  
  ---
  
  