Problem running Trinity
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Entering edit mode
8.3 years ago
biomagician ▴ 410

Dear Biostars, This is my first post on an online forum. I am seeking help with the installation and running of the Trinity software suite for the assembly of transcriptomes. I have spent two days trying different things to fix different errors but this is one sequence of commands I have tried to install and run the software. I am using a Mac OS X El Capitan version 10.11.5. I have highlighted the important lines in the sequences of commands, websites visited and outputs below. Thank you for your help.

brew install homebrew/science/trinity

==> Installing trinity from homebrew/science
==> Downloading https://homebrew.bintray.com/bottles-science/trinity-2.2.0_1.el_
Already downloaded: /Users/cr7/Library/Caches/Homebrew/trinity-2.2.0_1.el_capitan.bottle.tar.gz
==> Pouring trinity-2.2.0_1.el_capitan.bottle.tar.gz
**Warning: homebrew/science/trinity dependency bamtools was built with a different C++ standard
library (libc++ from clang). This may cause problems at runtime.**
==> Caveats
Trinity only officially supports Java 1.7. To skip this check pass the
option --bypass_java_version_check to Trinity. A specific Java version may
also be set via environment variable:
  JAVA_HOME=`/usr/libexec/java_home -v 1.7`
==> Summary
🍺  /Users/cr7/homebrew/Cellar/trinity/2.2.0_1: 1,222 files, 226.3M

http://stackoverflow.com/questions/18033255/install-libc-on-mac-10-6-8

https://github.com/Homebrew/homebrew-versions/blob/master/llvm33.rb

brew search llvm33

brew install homebrew/versions/llvm33

==> Installing llvm33 from homebrew/versions
==> Downloading http://llvm.org/releases/3.3/llvm-3.3.src.tar.gz
######################################################################## 100.0%
==> Patching
patching file Makefile.rules
patching file tools/llvm-shlib/Makefile
patching file tools/lto/Makefile
==> Downloading http://llvm.org/releases/3.3/cfe-3.3.src.tar.gz
######################################################################## 100.0%
==> Downloading http://llvm.org/releases/3.3/libcxx-3.3.src.tar.gz
######################################################################## 100.0%
==> Downloading http://llvm.org/releases/3.3/polly-3.3.src.tar.gz
######################################################################## 100.0%
==> Downloading http://llvm.org/releases/3.3/clang-tools-extra-3.3.src.tar.gz
######################################################################## 100.0%
==> ./configure --prefix=/Users/cr7/homebrew/Cellar/llvm33/3.3_1/lib/llvm-3.3 --enable-optimized --disable-b
==> make VERBOSE=1
==> make VERBOSE=1 install
==> make -C /private/tmp/llvm33-20160805-87927-1sdv239/llvm-3.3.src/projects/libcxx install CC=/Users/cr7/ho
==> Caveats
Extra tools are installed in /Users/cr7/homebrew/opt/llvm33/share/clang-3.3

To link to libc++, something like the following is required:
  CXX="clang++-3.3 -stdlib=libc++"
  CXXFLAGS="$CXXFLAGS -nostdinc++ -I/Users/cr7/homebrew/opt/llvm33/lib/llvm-3.3/lib/c++/v1"
  LDFLAGS="$LDFLAGS -L/Users/cr7/homebrew/opt/llvm33/lib/llvm-3.3/lib"

Python modules have been installed and Homebrew's site-packages is not
in your Python sys.path, so you will not be able to import the modules
this formula installed. If you plan to develop with these modules,
please run:
  mkdir -p /Users/cr7/.local/lib/python2.7/site-packages
  echo 'import site; site.addsitedir("/Users/cr7/homebrew/lib/python2.7/site-packages")' >> /Users/cr7/.local/lib/python2.7/site-packages/homebrew.pth
==> Summary
🍺  /Users/cr7/homebrew/Cellar/llvm33/3.3_1: 1,299 files, 140.8M, built in 8 minutes 45 seconds

brew reinstall homebrew/science/trinity

==> Reinstalling homebrew/science/trinity
==> Downloading https://homebrew.bintray.com/bottles-science/trinity-2.2.0_1.el_capitan.bottle.tar.gz
Already downloaded: /Users/cr7/Library/Caches/Homebrew/trinity-2.2.0_1.el_capitan.bottle.tar.gz
==> Pouring trinity-2.2.0_1.el_capitan.bottle.tar.gz
Warning: homebrew/science/trinity dependency bamtools was built with a different C++ standard
library (libc++ from clang). This may cause problems at runtime.
==> Caveats
Trinity only officially supports Java 1.7. To skip this check pass the
option --bypass_java_version_check to Trinity. A specific Java version may
also be set via environment variable:
  JAVA_HOME=`/usr/libexec/java_home -v 1.7`
==> Summary
🍺  /Users/cr7/homebrew/Cellar/trinity/2.2.0_1: 1,222 files, 226.3M

In the trinity installation folder

make -stdlib=libc++

Too much output to copy here

make

make: `all' is up to date.

cd sample_data/test_Trinity_Assembly

Running the sample data that comes with Trinity

**./runMe.sh**

#!/bin/bash -ve


if [ -e reads.right.fq.gz ] && [ ! -e reads.right.fq ]; then
    gunzip -c reads.right.fq.gz > reads.right.fq
fi

if [ -e reads.left.fq.gz ] && [ ! -e reads.left.fq ]; then
    gunzip -c reads.left.fq.gz > reads.left.fq
fi

if [ -e reads2.right.fq.gz ] && [ ! -e reads2.right.fq ]; then
    gunzip -c reads2.right.fq.gz > reads2.right.fq
fi

if [ -e reads2.left.fq.gz ] && [ ! -e reads2.left.fq ]; then
    gunzip -c reads2.left.fq.gz > reads2.left.fq
fi

Run Trinity to Generate Transcriptome Assemblies 

../../Trinity --seqType fq --max_memory 2G --left reads.left.fq.gz,reads2.left.fq.gz --right reads.right.fq.gz,reads2.right.fq.gz --SS_lib_type RF --CPU 4 --no_cleanup --normalize_reads
Trinity version: v2.2.0
-currently using the latest production release of Trinity.

Friday, August 5, 2016: 01:00:00    CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/support_scripts/ExitTester.jar 0
Friday, August 5, 2016: 01:00:00    CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/support_scripts/ExitTester.jar 1
Friday, August 5, 2016: 01:00:00    CMD: mkdir -p /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir
Friday, August 5, 2016: 01:00:00    CMD: mkdir -p /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir/chrysalis

Trinity Phase 1: Clustering of RNA-Seq Reads

    ---------------------------------------------------------------
    ------------ In silico Read Normalization ---------------------
    -- (Removing Excess Reads Beyond 50 Coverage --
    -- /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir/insilico_read_normalization --
    ---------------------------------------------------------------

        Friday, August 5, 2016: 01:00:00    

CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/insilico_read_normalization.pl --seqType fq --JM 2G  --max_cov 50 --CPU 4 --output /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir/insilico_read_normalization --SS_lib_type RF  --no_cleanup  --left /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.left.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.left.fq.gz --right /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.right.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.right.fq.gz --pairs_together --PARALLEL_STATS  
    Converting input files. (both directions in parallel)CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/fastool/fastool --rev  --illumina-trinity --to-fasta <(gunzip -c /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.left.fq.gz) >> left.fa
    CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/fastool/fastool --illumina-trinity --to-fasta <(gunzip -c /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.right.fq.gz) >> right.fa
    Sequences parsed: 30575
    CMD finished (0 seconds)
    CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/fastool/fastool --rev  --illumina-trinity --to-fasta <(gunzip -c /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.left.fq.gz) >> left.fa
    Sequences parsed: 30575
    CMD finished (0 seconds)
    Sequences parsed: 30575
    CMD finished (0 seconds)
    CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/fastool/fastool --illumina-trinity --to-fasta <(gunzip -c /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.right.fq.gz) >> right.fa
    Sequences parsed: 30575
    CMD finished (0 seconds)
    CMD: touch left.fa.ok
    CMD finished (0 seconds)
    CMD: touch right.fa.ok
    CMD finished (0 seconds)
    Done converting input files.CMD: cat left.fa right.fa > both.fa
    CMD finished (0 seconds)
    CMD: touch both.fa.ok
    CMD finished (0 seconds)
    -------------------------------------------
    ----------- Jellyfish  --------------------
    -- (building a k-mer catalog from reads) --
    -------------------------------------------

    CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 304805520  both.fa
    bash: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish: No such file or directory
    **Error, cmd: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 304805520  both.fa died with ret 32512 at /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/insilico_read_normalization.pl line 736.
    Error, cmd: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/insilico_read_normalization.pl --seqType fq --JM 2G**  --max_cov 50 --CPU 4 --output /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir/insilico_read_normalization --SS_lib_type RF  --no_cleanup  --left /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.left.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.left.fq.gz --right /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.right.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.right.fq.gz --pairs_together --PARALLEL_STATS   died with ret 512 at ../../Trinity line 2206.
RNA-Seq trinity installation • 6.7k views
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0
Entering edit mode

Welcome to biostars and congratulations on the comprehensive explanation. As suggested by Biogeek, the problem seems to be that trinity can't find jellyfish. Could you check if and where it is installed?

Alternatively, using git: git clone https://github.com/trinityrnaseq/trinityrnaseq.git

make

Might also do the trick :)

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0
Entering edit mode
8.3 years ago
Biogeek ▴ 470

Read the error:

trinity-plugins/jellyfish/bin/jellyfish: No such file or directory

Just get trinity off github, tar -xvzf it in your directory, and type make. I think your installation messed up via homebrew.

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0
Entering edit mode
8.3 years ago
biomagician ▴ 410

Thanks for your replies. I have tried the following but I do not know what to do with IRKE_run.cpp:9:10: fatal error: 'omp.h' file not found.

**git clone https://github.com/trinityrnaseq/trinityrnaseq.git**

**brew uninstall homebrew/science/trinity**

Uninstalling /Users/cr7/homebrew/Cellar/trinity/2.2.0_1... (1,242 files, 275.1M)

Cloning into 'trinityrnaseq'...
remote: Counting objects: 4090, done.
remote: Total 4090 (delta 0), reused 0 (delta 0), pack-reused 4090
Receiving objects: 100% (4090/4090), 207.36 MiB | 1.31 MiB/s, done.
Resolving deltas: 100% (2332/2332), done.
Checking connectivity... done.

**pwd
/PATHTO/trinity/installation/trinityrnaseq**

**make**

Using gnu compiler for Inchworm and Chrysalis
cd Inchworm && (test -e configure || autoreconf) \
                && sh ./configure --prefix=`pwd`  && /Applications/Xcode.app/Contents/Developer/usr/bin/make install
checking for a BSD-compatible install... /usr/bin/install -c
checking whether build environment is sane... yes
checking for a thread-safe mkdir -p... ./install-sh -c -d
checking for gawk... no
checking for mawk... no
checking for nawk... no
checking for awk... awk
checking whether make sets $(MAKE)... yes
checking for g++... g++
checking for C++ compiler default output file name... a.out
checking whether the C++ compiler works... yes
checking whether we are cross compiling... no
checking for suffix of executables... 
checking for suffix of object files... o
checking whether we are using the GNU C++ compiler... yes
checking whether g++ accepts -g... yes
checking for style of include used by make... GNU
checking dependency style of g++... gcc3
checking for library containing cos... none required
configure: creating ./config.status
config.status: creating Makefile
config.status: creating src/Makefile
config.status: creating config.h
config.status: config.h is unchanged
config.status: executing depfiles commands
Making install in src
g++ -DHAVE_CONFIG_H -I. -I..    -ansi -pedantic -fopenmp -Wall -Wextra -Wno-deprecated -m64 -g -O2 -MT Fasta_entry.o -MD -MP -MF .deps/Fasta_entry.Tpo -c -o Fasta_entry.o Fasta_entry.cpp
mv -f .deps/Fasta_entry.Tpo .deps/Fasta_entry.Po
g++ -DHAVE_CONFIG_H -I. -I..    -ansi -pedantic -fopenmp -Wall -Wextra -Wno-deprecated -m64 -g -O2 -MT IRKE_run.o -MD -MP -MF .deps/IRKE_run.Tpo -c -o IRKE_run.o IRKE_run.cpp
**IRKE_run.cpp:9:10: fatal error: 'omp.h' file not found**
#include <omp.h>
         ^
1 error generated.
make[2]: *** [IRKE_run.o] Error 1
make[1]: *** [install-recursive] Error 1
make: *** [inchworm_target] Error 2
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1
Entering edit mode

See if this fixes the problem.

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0
Entering edit mode

Please use ADD COMMENT to reply to earlier posts to keep this thread logical to follow.

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Entering edit mode
8.3 years ago
biomagician ▴ 410

Dear genomax2, I have tried your suggestion, without success:

**rm -r installation/
override r--r--r--  cr7/1535 for installation//trinityrnaseq/.git/objects/pack/pack-6900dc09485cffd5d0419a97902c3fab0d701d1c.idx? yes**

**brew install trinity --cc=gcc-4.9**

BLABLA
**Warning: homebrew/science/trinity dependency bamtools was built with a different C++ standard
library (libc++ from clang). This may cause problems at runtime.**
==> Caveats
Trinity only officially supports Java 1.7. To skip this check pass the
option --bypass_java_version_check to Trinity. A specific Java version may
also be set via environment variable:
  JAVA_HOME=`/usr/libexec/java_home -v 1.7`
==> Summary
🍺  /Users/cr7/homebrew/Cellar/trinity/2.2.0_1: 1,222 files, 225.9M, built in 54 seconds

pwd

**PATH/TO/…/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly**

**./runMe.sh**

#!/bin/bash -ve


if [ -e reads.right.fq.gz ] && [ ! -e reads.right.fq ]; then
    gunzip -c reads.right.fq.gz > reads.right.fq
fi

if [ -e reads.left.fq.gz ] && [ ! -e reads.left.fq ]; then
    gunzip -c reads.left.fq.gz > reads.left.fq
fi

if [ -e reads2.right.fq.gz ] && [ ! -e reads2.right.fq ]; then
    gunzip -c reads2.right.fq.gz > reads2.right.fq
fi

if [ -e reads2.left.fq.gz ] && [ ! -e reads2.left.fq ]; then
    gunzip -c reads2.left.fq.gz > reads2.left.fq
fi



BLABLA
----------- Jellyfish  --------------------
-- (building a k-mer catalog from reads) --
-------------------------------------------

CMD: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 304805520  both.fa
**bash: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish: No such file or directory**
Error, cmd: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 304805520  both.fa died with ret 32512 at /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/insilico_read_normalization.pl line 736.
Error, cmd: /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/insilico_read_normalization.pl --seqType fq --JM 2G  --max_cov 50 --CPU 4 --output /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/trinity_out_dir/insilico_read_normalization --SS_lib_type RF  --no_cleanup  --left /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.left.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.left.fq.gz --right /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads.right.fq.gz,/Users/cr7/homebrew/Cellar/trinity/2.2.0_1/sample_data/test_Trinity_Assembly/reads2.right.fq.gz --pairs_together --PARALLEL_STATS   died with ret 512 at ../../Trinity line 2206.
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0
Entering edit mode

As indicated by @Wouter earlier please use ADD REPLY/ADD COMMENT when responding to existing posts. e.g. this belongs against my comment up above. SUBMIT ANSWERS box is only meant for NEW answers for the original question.

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0
Entering edit mode

I don't want to discourage you but what kind of dataset/genome size are you working with? How much RAM do you have on this Mac?

Before you spend large amounts of time getting this going you should consider that Trinity requires large amounts of RAM to function (link1 and link2) . You may want to consider using a free web resource instead.

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0
Entering edit mode

The reads in fastq format are 3.46 GB. I have 16 GB on my laptop but I give it 15 GB to run. I will look into the free web resource.

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Entering edit mode

But I do have access to 64 GB on my institute cluster.

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Entering edit mode

Are you able to look in this location and see if the jellyfish executable is there?

ls -lh /Users/cr7/homebrew/Cellar/trinity/2.2.0_1/util/..//trinity-plugins/jellyfish/bin/jellyfish

At this point the install has gone through so many iterations that things could be subtly messed up. Are you feeling up to starting from scratch (remove the current install) and trying the install again?

ADD REPLY •
0
Entering edit mode

Thanks I will try. I have also decided to do it on Ubuntu in the meantime. It took me 5 minutes to install and make Trinity work on Ubunty.

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