Biowulf High Performance Computing at the NIH
ANNOgesic: Accurate RNA-Seq-based annotation of bacterial and archaeal genomes

Processing and integrating RNA-Seq data in order to generate high-resolution annotations is challenging, time consuming and requires numerous different steps. ANNOgesic is a powerful and modular pipeline that provides the required analyses and simplifies RNA-Seq-based bacterial and archaeal genome annotation. It predicts and annotates numerous features, including small non-coding RNAs, with high precision.


Important Notes

Interactive job
Interactive jobs should be used for debugging, graphics, or applications that cannot be run as batch jobs.

Allocate an interactive session and run the program. Sample session:

[user@biowulf]$ sinteractive --mem=4g
[user@cn3316 ~]$ module load ANNOgesic
[+] Loading singularity  on cn3316 
[+] Loading ANNOgesic 1.0.2  ...
At this point, user has two options:
1) typing the command
[user@cn3316 user]$ ag 
(without arguments) will bring the user into the singularity container shell environment
Singularity ANNOgesic.sqsh:~>
from which one can run any script or command accessible within the container on any data accessible from inside the container. For example, the following commands will run built-in tests:
Singularity ANNOgesic.sqsh:~> python /ANNOgesic/tests/ 
Detecting operons of test
Warning: No proper file - test.gff
Ran 2 tests in 0.009s

Singularity ANNOgesic.sqsh:~> python3 /ANNOgesic/tests/ .......Plotting nusB .. ---------------------------------------------------------------------- Ran 9 tests in 1.567s OK
To exit from the container shell environment, type:
Singularity ANNOgesic.sqsh:~> exit

2) typing the same command followed by another supported command as an additional aggument will result in performiing the second command without explicitly entering the container shell. For example:
[user@cn3316 ~]$ ag python /ANNOgesic/tests/ 
Ran 1 test in 0.004s

[user@cn3316 ~]$ ag python3 /ANNOgesic/tests/ 
Ran 10 tests in 0.018s

In particular, the following command will display ANNOgesic help message:
[user@cn3316 ~]$ ag annogesic --help 

       ___    _   ___   ______                  _     
      /   |  / | / / | / / __ \____ ____  _____(_)____ \
  __ / /| | /  |/ /  |/ / / / / __ `/ _ \/ ___/ / ___/__\
 |  / ___ |/ /|  / /|  / /_/ / /_/ /  __(__  ) / /__    /
 | /_/  |_/_/ |_/_/ |_/\____/\__, /\___/____/_/\___/   /
 |                          /____/ 
 |                                                \

usage: annogesic [-h] [--version]

positional arguments:
    create              Create a project
    get_input_files     Get required files. (i.e. annotation files, fasta
                        Get fasta files of reference genomes if the reference
                        sequences do not exist.
                        Transfer the annotations from a closely related
                        species genome to a target genome.
    tss_ps              Detect TSSs or processing sites.
    optimize_tss_ps     Optimize TSSs or processing sites based on manual
                        detected ones.
    terminator          Detect rho-independent terminators.
    transcript          Detect transcripts based on coverage file.
    utr                 Detect 5'UTRs and 3'UTRs.
    srna                Detect intergenic, antisense and UTR-derived sRNAs.
    sorf                Detect expressed sORFs.
    promoter            Discover promoter motifs.
    operon              Detect operons and sub-operons.
    circrna             Detect circular RNAs.
    go_term             Extract GO terms from Uniprot.
    srna_target         Detect sRNA-mRNA interactions.
    snp                 Detect SNP/mutation and generate fasta file if
                        mutations were found.
    ppi_network         Detect protein-protein interactions suported by
    localization        Predict subcellular localization of proteins.
                        Predict riboswitches and RNA thermometers.
    crispr              Predict CRISPR related RNAs.
    merge_features      Merge all features to one gff file.
    screenshot          Generate screenshots for selected features using IGV.
                        Add color information to screenshots (e.g. useful for
                        dRNA-Seq based TSS and PS detection. It only works
                        after running "screenshot" (after running batch

optional arguments:
  -h, --help            show this help message and exit
  --version, -v         show version

Batch job
Most jobs should be run as batch jobs.

Create a batch input file (e.g. For example:

module load ANNOgesic
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python3 /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python3 /ANNOgesic/tests/
ag python  /ANNOgesic/tests/
ag python3 /ANNOgesic/tests/

Submit this job using the Slurm sbatch command.

sbatch [--cpus-per-task=#] [--mem=#]