Trinity

Trinity is used for de novo reconstruction of transcriptomes from RNA-seq data. Trinity combines three independent software modules: Inchworm, Chrysalis, and Butterfly, applied sequentially to process large volumes of RNA-seq reads. Trinity partitions the sequence data into many individual de Bruijn graphs, each representing the transcriptional complexity at a given gene or locus, and then processes each graph independently to extract full-length splicing isoforms and to tease apart transcripts derived.

The Trinity module at CSC also includes TransDecoder and Trinotate tools to analyze the results of a Trinity run.

Trinity

License

Free to use and open source under Broad Institute License.

Available

Puhti: 2.8.5, 2.11.0, 2.13.2, 2.14.0, 2.15.1

Usage

Using Trinity

On Puhti, Trinity is set up with the command:

module load biokit

The biokit module sets up a set of commonly used bioinformatics tools including Trinity 2.8.5. If you want to use a newer version, e.g. 2.13.2, run the command:

module load trinty/2.13.2

Trinity should be used interactively in a compute node or preferably through the batch job system. Below is an example batch job file for Trinity.

#!/bin/bash 
#SBATCH --job-name=trinity
#SBATCH --output=output_%j.txt
#SBATCH --error=errors_%j.txt
#SBATCH --time=48:00:00
#SBATCH --ntasks=1
#SBATCH --nodes=1  
#SBATCH --cpus-per-task=6
#SBATCH --mem=24000
#SBATCH --account=project_1234567

module load trinity/2.13.2

Trinity --seqType fq --max_memory 22G --left reads.left.fq --right \
reads.right.fq --SS_lib_type RF --CPU $SLURM_CPUS_PER_TASK \
--output trinity_run_out --grid_exec sbatch_commandlist

The batch script above reserves 6 computing cores from one node for the job. The maximal run time of the sample job here is 48 hours. About 4 GB of memory is reserved for each core so the total memory reservation is 6 * 4 GB = 24 GB. On Puhti, you must use batch job option --account= to define the project to be used. You should replace project_1234567 used in the example with your own project. You can check your projects with command: csc-projects.

In the actual Trinity command the number of computing cores to be used (--CPU) is set using the environment variable $SLURM_CPUS_PER_TASK. This variable contains the value set by the --cpus-per-task Slurm option.

On Puhti, you can also use distributed computing to speed up the Trinity job. When the definition:

--grid_exec sbatch_commandlist

is added to the command, some phases of the analysis tasks are executed as a set of parallel sub-jobs. For large Trinity tasks the settings of the sbatch_commandlist tool are too limited. In these cases replace sbatch_commandlist with sbatch_commandlist_trinity.

--grid_exec sbatch_commandlist_trinity

When Trinity is executed with --grid_exec option, it generates a large amount of temporary files, and it is very likely that you will exceed the default limit of 100 000 files. Thus, it is advisable to apply for a larger file number quota for Puhti scratch before submitting large Trinity jobs. You can send the request to CSC Service Desk.

When the batch job file is ready, it can be submitted to the batch queue system with the command:

sbatch batch_job_file

Look here for more information about running batch jobs.

Please also check the Trinity website to get hints for estimating the required resources.

Using autoTrinotate

You can analyze the results of your Trinity job with autoTrinotate. You need two files resulting from a successful Trinity assembly.

Fasta-formatted nucleotide sequence file containing the final contigs created by Trinity (Trinity.fasta)
gene-to-trans map for the input fasta file (Trinity.fasta.gene_to_trans_map)

Note that depending on the Trinity version, these names may have a prefix as defined with the --output option (e.g. trinity_run_out.Trinity.fasta).

Copy a template sqlite database for your analysis:

cp $TRINOTATE_HOME/databases/Trinotate.sqlite mydb.sqlite

You can then launch autoTrinotate with the command:

$TRINOTATE_HOME/auto/autoTrinotate.pl --Trinotate_sqlite mydb.sqlite --transcripts Trinity.fasta --gene_to_trans_map  Trinity.fasta.gene_to_trans_map --conf $TRINOTATE_HOME/auto/conf.txt --CPU  $SLURM_CPUS_PER_TASK

Note

autoTrinotate analysis can require a lot of resources, so you should execute the command in an interactive session or as a batch job!

autoTrinotate produces an SQLite database file that can be further analyzed with the command:

$TRINOTATE_HOME/Trinotate

More information

Trinity home page