Ths folder contains additional files related to generic homology search and phylogenetic inference analyses, which are out of the scope of the book chapter. It is exepected that the user would apply it's own pipeline according to its needs. This folder also contains metadata files and templates for iTOL.
Contents:
Files description
Homology search and phylogenetic inference
| Files | Description |
|---|---|
| Bax1-I.hmm | hmm profile used to identify TMBIM proteins. The profile was obtained from Pfam. |
| Bax1-I_cutga_c0.8.tar.gz | Compressed output from executing the hmm_retrieve.sh script. |
| TMBIM_proteins.fasta | All 117 proteins in FASTA format used as example for the book chapter. |
| TMBIM_proteins.afa | Alignment of all 117 proteins in FASTA format used as example for the book chapter. |
| TMBIM_proteins.trimmed.afa | Trimmed alignment of all 117 proteins in FASTA format used as example for the book chapter. |
| TMBIM_proteins.trimmed.afa.* | Output files from iqtree2. The TMBIM_proteins.trimmed.treefile file was used to visualize the SynNet communities in iTOL. |
| community_colors.txt | Contains color hex codes for the communities. |
| create_table.R | Script used to create the nodes_metadata.csv file. |
| iTOL.xlsx | Excel file containing the information for the iTOL templates. |
| iTOL_template_connections.txt | iTOL template for communities. |
| iTOL_template_labels.txt | iTOL template for label colors. |
| iTOL_template_strips.txt | iTOL template for strop colors. |
| nodes_metadata.csv | Metadata of the 117 TMBIM proteins used as example for this study. |
| species_metadata.csv | Metadata of the 15 plant species. |
| species_tree.newick | Species tree used for phylogenetic profiling. The tree was generated with ete3 and furhter modified in iTOL. |
Homology searches were performed with HMMER3 implemented in the hmm_retrieve.sh script and using the Bax1-I.hmm (PF01027) obtained from Pfam. The command call was:
hmm_retrieve.sh -g -c 0.8 -p Bax1-I.hmm full/path/to/synnet_protocol_starter/db/*pep
The results were concatenated in the TMBIM_proteins.fasta file and the sequence identifiers in the TMBIM_id_list.txt file.
For protein alignment and trimming, the hmmalign command of hmmer3 and the trimAl programs were used, respectively:
hmmalign --trim --outformat afa Bax1-I.hmm TMBIM_proteins.fasta | sed -e '/>/!s/\./-/g' > TMBIM_proteins.afa
trimal -in TMBIM_proteins.afa -out TMBIM_proteins.trimmed.afa -gt 0.8
Phylogenetic inference was done with iqtree2:
iqtree2 --threads-max 3 -m TEST --mset WAG,LG,JTT -B 1000 -alrt 1000 -s TMBIM_proteins.trimmed.afa
Outputs from iqtree2 are TMBIM_proteins.trimmed.afa*. The TMBIM_proteins.trimmed.treefile was used to visualize the tree in iTOL in the protocol of the book chapter.