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STAR-Fusion / STAR-Fusion-Tutorial

Tutorial for STAR-Fusion, FusionInspector, and de novo reconstruction of fusion transcripts using Trinity
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No such file or directory at /usr/local/src/STAR-Fusion/PerlLib/Pipeliner.pm line 181 #3

Open mapostolides opened 5 years ago

mapostolides commented 5 years ago

The Docker container tutorial isn't working for me. I successfully ran:

docker run --rm -it -vpwd :/data trinityctat/ctatfusion:latest bash

Once the container was pulled, I successfully ran...

${STAR_FUSION_HOME}/FusionFilter/prep_genome_lib.pl \ --genome_fa minigenome.fa \ --gtf minigenome.gtf \ --fusion_annot_lib CTAT_HumanFusionLib.mini.dat.gz

...with no errors. However, when running the following command...

${STAR_FUSION_HOME}/STAR-Fusion \ --left_fq rnaseq_1.fastq.gz \ --right_fq rnaseq_2.fastq.gz \ --genome_lib_dir ctat_genome_lib_build_dir

...I ran into an error which I'm not sure how to resolve.

root@50aaf3eb3bd1:/data# ${STAR_FUSION_HOME}/STAR-Fusion --left_fq rnaseq_1.fastq.gz --right_fq rnaseq_2.fastq.gz --genome_lib_dir ctat_genome_lib_build_dir 
${STAR_FUSION_HOME}/STAR-Fusion --left_fq rnaseq_1.fastq.gz --right_fq rnaseq_2.fastq.gz --genome_lib_dir ctat_genome_lib_build_dir 
* Running CMD: /usr/local/bin/STAR --genomeDir /data/ctat_genome_lib_build_dir/ref_genome.fa.star.idx  --outReadsUnmapped None  --chimSegmentMin 12  --chimJunctionOverhangMin 12  --chimOutJunctionFormat 1  --alignSJDBoverhangMin 10  --alignMatesGapMax 100000  --alignIntronMax 100000  --alignSJstitchMismatchNmax 5 -1 5 5  --runThreadN 4 --outSAMstrandField intronMotif  --outSAMunmapped Within  --outSAMtype BAM Unsorted  --readFilesIn /data/rnaseq_1.fastq.gz /data/rnaseq_2.fastq.gz  --outSAMattrRGline ID:GRPundef  --chimMultimapScoreRange 10 --chimMultimapNmax 10 --chimNonchimScoreDropMin 10  --peOverlapNbasesMin 12 --peOverlapMMp 0.1  --genomeLoad NoSharedMemory  --twopassMode Basic  --readFilesCommand 'gunzip -c' 
Nov 23 20:37:50 ..... started STAR run
Nov 23 20:37:50 ..... loading genome
Nov 23 20:37:59 ..... started 1st pass mapping
Nov 23 20:40:06 ..... finished 1st pass mapping
Nov 23 20:40:06 ..... inserting junctions into the genome indices
Killed
Error, cmd: /usr/local/bin/STAR --genomeDir /data/ctat_genome_lib_build_dir/ref_genome.fa.star.idx  --outReadsUnmapped None  --chimSegmentMin 12  --chimJunctionOverhangMin 12  --chimOutJunctionFormat 1  --alignSJDBoverhangMin 10  --alignMatesGapMax 100000  --alignIntronMax 100000  --alignSJstitchMismatchNmax 5 -1 5 5  --runThreadN 4 --outSAMstrandField intronMotif  --outSAMunmapped Within  --outSAMtype BAM Unsorted  --readFilesIn /data/rnaseq_1.fastq.gz /data/rnaseq_2.fastq.gz  --outSAMattrRGline ID:GRPundef  --chimMultimapScoreRange 10 --chimMultimapNmax 10 --chimNonchimScoreDropMin 10  --peOverlapNbasesMin 12 --peOverlapMMp 0.1  --genomeLoad NoSharedMemory  --twopassMode Basic  --readFilesCommand 'gunzip -c'  died with ret 35072 No such file or directory at /usr/local/src/STAR-Fusion/PerlLib/Pipeliner.pm line 181.
    Pipeliner::run(Pipeliner=HASH(0x1572cd0)) called at /usr/local/src/STAR-Fusion/STAR-Fusion line 862
    main::run_STAR(Pipeliner=HASH(0x1572cd0), "/data/rnaseq_1.fastq.gz", "/data/rnaseq_2.fastq.gz", "") called at /usr/local/src/STAR-Fusion/STAR-Fusion line 509

I went to the file /usr/local/src/STAR-Fusion/PerlLib/Pipeliner.pm inside the Docker container, and that exact file was, in fact, there. I'm not sure exactly what the reason for the above error might be, since I followed the instructions exactly.

Any help would be much appreciated!

Thanks, Michael

brianjohnhaas commented 5 years ago

Hi,

It probably ran out of memory. STAR can require a lot of RAM (40G) for running on the human genome.

On Fri, Nov 23, 2018 at 3:45 PM mapostolides notifications@github.com wrote:

The Docker container tutorial isn't working for me. I successfully ran:

docker run --rm -it -v pwd:/data trinityctat/ctatfusion:latest bash

Once the container was pulled, I successfully ran...

${STAR_FUSION_HOME}/FusionFilter/prep_genome_lib.pl \ --genome_fa minigenome.fa \ --gtf minigenome.gtf \ --fusion_annot_lib CTAT_HumanFusionLib.mini.dat.gz

...with no errors. However, when running the following command...

${STAR_FUSION_HOME}/STAR-Fusion \ --left_fq rnaseq_1.fastq.gz \ --right_fq rnaseq_2.fastq.gz \ --genome_lib_dir ctat_genome_lib_build_dir

...I ran into an error which I'm not sure how to resolve.

`root@50aaf3eb3bd1:/data# ${STAR_FUSION_HOME}/STAR-Fusion --left_fq rnaseq_1.fastq.gz --right_fq rnaseq_2.fastq.gz --genome_lib_dir ctat_genome_lib_build_dir ${STAR_FUSION_HOME}/STAR-Fusion --left_fq rnaseq_1.fastq.gz --right_fq rnaseq_2.fastq.gz --genome_lib_dir ctat_genome_lib_build_dir

  • Running CMD: /usr/local/bin/STAR --genomeDir /data/ctat_genome_lib_build_dir/ref_genome.fa.star.idx --outReadsUnmapped None --chimSegmentMin 12 --chimJunctionOverhangMin 12 --chimOutJunctionFormat 1 --alignSJDBoverhangMin 10 --alignMatesGapMax 100000 --alignIntronMax 100000 --alignSJstitchMismatchNmax 5 -1 5 5 --runThreadN 4 --outSAMstrandField intronMotif --outSAMunmapped Within --outSAMtype BAM Unsorted --readFilesIn /data/rnaseq_1.fastq.gz /data/rnaseq_2.fastq.gz --outSAMattrRGline ID:GRPundef --chimMultimapScoreRange 10 --chimMultimapNmax 10 --chimNonchimScoreDropMin 10 --peOverlapNbasesMin 12 --peOverlapMMp 0.1 --genomeLoad NoSharedMemory --twopassMode Basic --readFilesCommand 'gunzip -c' Nov 23 20:37:50 ..... started STAR run Nov 23 20:37:50 ..... loading genome Nov 23 20:37:59 ..... started 1st pass mapping Nov 23 20:40:06 ..... finished 1st pass mapping Nov 23 20:40:06 ..... inserting junctions into the genome indices Killed Error, cmd: /usr/local/bin/STAR --genomeDir /data/ctat_genome_lib_build_dir/ref_genome.fa.star.idx --outReadsUnmapped None --chimSegmentMin 12 --chimJunctionOverhangMin 12 --chimOutJunctionFormat 1 --alignSJDBoverhangMin 10 --alignMatesGapMax 100000 --alignIntronMax 100000 --alignSJstitchMismatchNmax 5 -1 5 5 --runThreadN 4 --outSAMstrandField intronMotif --outSAMunmapped Within --outSAMtype BAM Unsorted --readFilesIn /data/rnaseq_1.fastq.gz /data/rnaseq_2.fastq.gz --outSAMattrRGline ID:GRPundef --chimMultimapScoreRange 10 --chimMultimapNmax 10 --chimNonchimScoreDropMin 10 --peOverlapNbasesMin 12 --peOverlapMMp 0.1 --genomeLoad NoSharedMemory --twopassMode Basic --readFilesCommand 'gunzip -c' died with ret 35072 No such file or directory at /usr/local/src/STAR-Fusion/PerlLib/Pipeliner.pm line 181. Pipeliner::run(Pipeliner=HASH(0x1572cd0)) called at /usr/local/src/STAR-Fusion/STAR-Fusion line 862 main::run_STAR(Pipeliner=HASH(0x1572cd0), "/data/rnaseq_1.fastq.gz", "/data/rnaseq_2.fastq.gz", "") called at /usr/local/src/STAR-Fusion/STAR-Fusion line 509`

I went to the directory /usr/local/src/STAR-Fusion/PerlLib/Pipeliner.pm inside the Docker container, and that exact file was, in fact, there. I'm not sure exactly what the reason for the above error might be, since I followed the instructions exactly.

Any help would be much appreciated!

Thanks, Michael

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mapostolides commented 5 years ago

Hi Brian, thanks for your reply!

Are you sure that's the reason? I am using the exact files provided in the tutorial, and it states in the tutorial that 4G of RAM should be sufficient for the specific files provided inside the Docker container. The computer I am using has 8G of RAM, which is more than enough according to the tutorial. Is it possible that there is some other issue with the Docker container? This tutorial seems really useful, and it would be great if I could use your tools!

Thanks, Michael

brianjohnhaas commented 5 years ago

right, for the tutorial it shouldn't require much RAM. Are you using the ctat genome lib that goes with the tutorial, or are you using the larger one that's used in regular star-fusion runs? The tutorial has a tiny genome lib and shouldn't require much ram.

On Sat, Nov 24, 2018 at 11:45 AM mapostolides notifications@github.com wrote:

Hi Brian, thanks for your reply!

Are you sure that's the reason? I am using the exact files provided in the tutorial, and it states in the tutorial that 4G of RAM should be sufficient for the specific files provided inside the Docker container. The computer I am using has 8G of RAM, which is more than enough according to the tutorial. Is it possible that there is some other issue with the Docker container? This tutorial seems really useful, and it would be great if I could use your tools!

Thanks, Michael

— You are receiving this because you commented. Reply to this email directly, view it on GitHub https://github.com/STAR-Fusion/STAR-Fusion-Tutorial/issues/3#issuecomment-441380246, or mute the thread https://github.com/notifications/unsubscribe-auth/AHMVX-cS8dg854j-ENf20wbyLuNbC0n1ks5uyXelgaJpZM4YxG5m .

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

brianjohnhaas commented 5 years ago

Also, the limitation is that the ctat genome lib that goes with the tutorial is only useful for that tutorial, and not for general use.

On Sat, Nov 24, 2018 at 1:13 PM Brian Haas bhaas@broadinstitute.org wrote:

right, for the tutorial it shouldn't require much RAM. Are you using the ctat genome lib that goes with the tutorial, or are you using the larger one that's used in regular star-fusion runs? The tutorial has a tiny genome lib and shouldn't require much ram.

On Sat, Nov 24, 2018 at 11:45 AM mapostolides notifications@github.com wrote:

Hi Brian, thanks for your reply!

Are you sure that's the reason? I am using the exact files provided in the tutorial, and it states in the tutorial that 4G of RAM should be sufficient for the specific files provided inside the Docker container. The computer I am using has 8G of RAM, which is more than enough according to the tutorial. Is it possible that there is some other issue with the Docker container? This tutorial seems really useful, and it would be great if I could use your tools!

Thanks, Michael

— You are receiving this because you commented. Reply to this email directly, view it on GitHub https://github.com/STAR-Fusion/STAR-Fusion-Tutorial/issues/3#issuecomment-441380246, or mute the thread https://github.com/notifications/unsubscribe-auth/AHMVX-cS8dg854j-ENf20wbyLuNbC0n1ks5uyXelgaJpZM4YxG5m .

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

mapostolides commented 5 years ago

I'm using the one that comes inside the Docker container. I'm pretty sure that's the ctat genome library that goes with the tutorial.

Thanks!

On Sat, Nov 24, 2018 at 1:14 PM Brian Haas notifications@github.com wrote:

Also, the limitation is that the ctat genome lib that goes with the tutorial is only useful for that tutorial, and not for general use.

On Sat, Nov 24, 2018 at 1:13 PM Brian Haas bhaas@broadinstitute.org wrote:

right, for the tutorial it shouldn't require much RAM. Are you using the ctat genome lib that goes with the tutorial, or are you using the larger one that's used in regular star-fusion runs? The tutorial has a tiny genome lib and shouldn't require much ram.

On Sat, Nov 24, 2018 at 11:45 AM mapostolides notifications@github.com wrote:

Hi Brian, thanks for your reply!

Are you sure that's the reason? I am using the exact files provided in the tutorial, and it states in the tutorial that 4G of RAM should be sufficient for the specific files provided inside the Docker container. The computer I am using has 8G of RAM, which is more than enough according to the tutorial. Is it possible that there is some other issue with the Docker container? This tutorial seems really useful, and it would be great if I could use your tools!

Thanks, Michael

— You are receiving this because you commented. Reply to this email directly, view it on GitHub < https://github.com/STAR-Fusion/STAR-Fusion-Tutorial/issues/3#issuecomment-441380246 , or mute the thread < https://github.com/notifications/unsubscribe-auth/AHMVX-cS8dg854j-ENf20wbyLuNbC0n1ks5uyXelgaJpZM4YxG5m

.

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

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brianjohnhaas commented 5 years ago

sorry about that. We've used it before for workshops and it hasn't required much ram. Not sure why it's causing trouble here. If you can try it on a larger machine, let's see if it works there.

On Mon, Nov 26, 2018 at 11:02 AM mapostolides notifications@github.com wrote:

I'm using the one that comes inside the Docker container. I'm pretty sure that's the ctat genome library that goes with the tutorial.

Thanks!

On Sat, Nov 24, 2018 at 1:14 PM Brian Haas notifications@github.com wrote:

Also, the limitation is that the ctat genome lib that goes with the tutorial is only useful for that tutorial, and not for general use.

On Sat, Nov 24, 2018 at 1:13 PM Brian Haas bhaas@broadinstitute.org wrote:

right, for the tutorial it shouldn't require much RAM. Are you using the ctat genome lib that goes with the tutorial, or are you using the larger one that's used in regular star-fusion runs? The tutorial has a tiny genome lib and shouldn't require much ram.

On Sat, Nov 24, 2018 at 11:45 AM mapostolides < notifications@github.com> wrote:

Hi Brian, thanks for your reply!

Are you sure that's the reason? I am using the exact files provided in the tutorial, and it states in the tutorial that 4G of RAM should be sufficient for the specific files provided inside the Docker container. The computer I am using has 8G of RAM, which is more than enough according to the tutorial. Is it possible that there is some other issue with the Docker container? This tutorial seems really useful, and it would be great if I could use your tools!

Thanks, Michael

— You are receiving this because you commented. Reply to this email directly, view it on GitHub <

https://github.com/STAR-Fusion/STAR-Fusion-Tutorial/issues/3#issuecomment-441380246

,

or mute the thread <

https://github.com/notifications/unsubscribe-auth/AHMVX-cS8dg854j-ENf20wbyLuNbC0n1ks5uyXelgaJpZM4YxG5m

.

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Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

--

Brian J. Haas The Broad Institute http://broadinstitute.org/~bhaas http://broad.mit.edu/~bhaas

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ruizgo commented 4 years ago

Great, it solved my problem.

Daemon-ser commented 3 years ago

Great, it solved my problem.

Same problem here, how do you solve it ?

mapostolides commented 3 years ago

It was a while ago that I did this, but I know that Docker containers are launched by default with 2G of RAM. Could it be because Docker needs to give more memory to containers? image

Daemon-ser commented 3 years ago

It was a while ago that I did this, but I know that Docker containers are launched by default with 2G of RAM. Could it be because Docker needs to give more memory to containers? image

Thanks a lot.