PO_2_MLSA.py v1.5.2

Automizes the creation of concatenated alignments of single copy genes with orthologues in comparison organisms for the construction of MLSA-based phylogenetic trees. Can optionally automize pylogenetic tree calculation using RaXML

Single-copy markers present in EVERY comparison organism EXACTLY ONCE are identified from a pre-calculated proteinortho5 resultfile, and extracted from corresponding supplied protein-fastas

Unalignable C- and N-terminal fregments will be clipped from each detected markergene and remaining poorly aligned positions will be automatically filtered from each single marker alignment using Gblocks

Optionally the resulting concatenated alignments can be specified to contain all gapped alignment positions or to be stripped either of all gapped positions or of only such gapped positions in the flanking regions of each composite ortholog

This script is supposed to be part of a pipeline consisting of:

1. Extraction of protein seqeunces (with unique identifiers!) fom Genbank/Embl-Files to one multi-fasta per comparison-organism using, e.g. CDS_extractor.pl by Andreas Leimbach

2. Calculation of orthologs and paralogs using proteinortho5 (with the '-single' and '-self' arguments!)

3. The creation of concatenated MLSA-sequences based on:

   • the fasta sequences of step 1.
   • the proteinortho5-results from step 2.

The output-file will be in fasta format (but including gapped positions, so remember to use 'fasta_wgap' if and when loading into Arb.

Dependancies:

• Python v2.7+
• Biopython
• proteinortho5
• CDS_extractor.pl
• muscle
• Gblocks
• RaXML (optional, only when creating trees with RaXML)

Usage:

usage: PO_2_MLSA.py [-h] -po PO_FILE [-f FASTA_PATH] [-t NTHREADS] [--filter]
                    [-op OUT_PATH] [-tp TEMP_PATH] [-kt] [-mb MUSCLE_BINARY]
                    [-gbb GBLOCKS_BINARY] [-rmlb RAXML_BINARY]
                    [-mt {raxml,raxml_rapidbs,nj,nj_bs,none}] [-sd SEED_NR]
                    [-bs NR_BOOTSTRAPS]
                    [--nj_substmodel]
                    [-v] [-s] [--debug]

Arguments:

Basic arguments:

  -h, --help            show a help message and exit
  -po PO_FILE, --proteinortho PO_FILE
                        (String) file with proteinortho5 results
  -f FASTA_PATH, --fastas FASTA_PATH
                        (String) path to fasta files (produced by CDS_extractor) 
                        Default = current working directory
  -t NTHREADS, --threads NTHREADS
                        Maximum number of threads to use for alignment steps
                        Default = 1
  --filter              OPTIONAL: Filter outliers based on single marker phylogenies in order to remove HGT events (Default: False)

Output-related arguments:

  -op OUT_PATH, --out_path OUT_PATH
                        Path to output (will be created if it does not exist)
                        Default = current working directory
  -tp TEMP_PATH, --temp_path TEMP_PATH
                        Path for temporary files (will be created if does not exist)
                        Default =  current working directory
  -kt, --keep_temp      Keep all temporary files and intermediate results
                        (Multifasta_files containing the singe genes involved in the MLSA-alignments are stored in any case)
                        Default: delete all temporary files

Dependancies/external binaries-related arguments:

  -mb MUSCLE_BINARY, --muscle_binary MUSCLE_BINARY
                        Muscle binary to use (including path if not in $PATH). default: assume "muscle" in $PATH
  -gbb GBLOCKS_BINARY, --gblocks GBLOCKS_BINARY
                        Gblocks binaries to use (include path to binaries if not listed in $PATH). Default: "Gblocks" (assumed to be in PATH)
  -rmlb RAXML_BINARY, --raxml RAXML_BINARY
                        RaxML excecutable biaries to use (Include path to binary if not listed in $PATH). Default: check for common naming of raxml binaries in $PATH, prioritizing binaries with "PTHREADS" in the name

Tree-calculation-related arguments:

  -mt {raxml,raxml_rapidbs,nj,nj_bs,none}, --make_tree {raxml,raxml_rapidbs,nj,nj_bs,none}
                        Generate ML phylogenetic trees using RAxML with the substitution model "PROTGAMMAAUTO"
                        choices: "raxml": single tree without bootstraps (using new rapid hill climbing)
                        	 "raxml_rapidbs": rapid bootstrap analyses and search for best ML tree in one run
                        	 "none"
                        Default = none
  -sd SEED_NR, --seed SEED_NR
                        Integer to provide as seed for RAxML
                        0 = seed generated randomly
                        Default = random seed
  -bs NR_BOOTSTRAPS, --bootstraps NR_BOOTSTRAPS
                        Number of bootstraps(if any)
                        default = 1000
  --nj_substmodel {identity,ident,blosum,pam,blosum30,blosum35,blosum40,blosum45,blosum50,blosum55,blosum60,blosum62,blosum65,blosum70,blosum75,blosum80,blosum85,blosum90,blosum95,blosum100,pam30,pam60,pam90,pam120,pam180,pam250,pam300}
                        Substitution model for distance matrix calculation. Only "identity" and variances of "blosum" and "pam" are offered. 
                        abbreviations:
                        	"identity"
                        	"blosum" = blosum62
                        	"pam" = pam120, default = "identity"

Other arguments:

  -v, --version         show version information and then quit (don't run complete script)
  -s, --silent          non-verbose mode
  --debug               Log extra info for debugging