ITSoneDB population pipeline

---

ITSoneDB is a database collecting Eukayotic ITS1 sequences and consistent taxonomic annotations. It is available at http://itsonedb.cloud.ba.infn.it/.

---

RATIONALE

The pipeline designed for ITSoneDB population integrates ad-hoc Python and BASH scripts and third-party tools (See the figure below).

1. In the initial step the ENA entries are locally downloaded and eukaryotic entries are extracted.

2. From each entry specific information (i.e. accession number, version, description line and annotation under specific keys) are pulled out and stored in a TSV file and consistent sequence data are annotated in FASTA files. TSV and FASTA files are analyzed by two parallel procedures to extract or to infer the ITS1 location. The TSV files are parsed out to extract the annotation relative to ITS1 boundaries by means of a commonly used ITS1 synonyms dictionary.

3. In parallel, HMM profiles for 18S and 5.8S rRNA genes are mapped on FASTA files by means of hmmsearch (HMMER 3.1) (right diagram part). The ITS1 boundaries information obtained by both procedures are merged in order to produce the files needed to populate the database.

---

---

REQUIREMENTS

1. The Biopython module is required (for installation info see http://biopython.org).
2. The Species Representative Entry Identification procedure require VSEARCH (for installation info see https://github.com/torognes/vsearch).

USAGE

Following the instruction to execute the scripts:

• ENA flat file parsing:
  1. flat file parsing (the script should be applied in a folder containing the gzipped ENA flat files):
     • python ENA_parser.py
  2. prepare FASTA folder
     • gzip *fasta
     • mkdir fasta_file_folder
     • mv *fasta.gz fasta_file_folder
  3. Prepare TSV folder:
     • mkdir tsv_folder
     • mv *tsv tsv_folder
• Extraction of ITS1 annotated in ENA entries:
  python extract_ITS1_loc.py tsv_folder
• HMM based ITS1 boundaries inference:
  1. mkdir script
  2. mv estrazione_localizzazione_from_hmm.py script
  3. mv hmmer_txt_parser.py script
  4. cd script && pwd && cd ..
  5. substitute the result of the previous step in the line 5 of the BASH script ITSoneDB_update_bf.sh
  6. cd fasta_file_folder
  7. ls *fasta.gz > sample_list
  8. ./ITSoneDB_update_bf.sh