Pseudomonas population genomics
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Projects
- Build a local genome database
- Database schema:
- "genome": genome_id, strain_name, ncbi_taxid
- "orf": genome_id, locus_tag, start, stop, strand, genome_name, product_name
- "orth_orf": orth_orf_id, locus_name, genome_id, orth_class
- Parsing scripts
- Rayees Parsing code, requires that you remove columns 9-27 using bash command:
cut -c 1-8
(I will write a bash script that does this and runs the program) https://www.dropbox.com/s/lpxxbkxeyw7frrn/parser.pl
- Rayees Parsing code, requires that you remove columns 9-27 using bash command:
- Database loading scripts
- Database schema:
- Molecular Evolution of flagellum genes
- Download orthologs
- Reconstruct phylogenetic tree
- Run PAML tests
Update:August 5, 2013
Pseudomonas Pipeline Steps:
USAGE: ./pseudo_pipline.sh [options] [FASTQ file]
- Step 0: Creation of Simulated reads via GemSim: http://www.biomedcentral.com/1471-2164/13/74 (outputs FastQ file with quality scores).
- Step 1: Assembly of reads via ALL Paths
- Step 2: ORF finding via Glimmer
- Step 3: Gene finding via BLAST
- Step 4: Top Gene matches and Database orth_id matching via top_orth_match.pl
- Step 5: Predicted Ortholog Database import
- Step 6: Core genome extraction
- Step 7: Choose random number of orthologs via random_cds.sh
- Step 8: Translation of Orthologs via BioSeq
- Step 9: Alignment of peptide files via Muscle
- Step 10: Reverse translation of aligned sequences via align_coding_seq.pl
- Step 11: Creation of Aligned Fasta file via AlignConcat_Bioperl.final.pl
- Step 12: Creation of treefile via FASTTREE
Pseudomonas Pipeline options:
- -i: keep all intermediary files (default remove all intermediate files)
- -f: keep only intermediate fasta files
- -b: [integer] blast score threshold
- -n: [integer] number of genes to create treefile from
Update:July 22, 2013
Estimation of recombination hotspots via LD_hat Ld_hat results:
Data | Number of Genes | ||||
No PA7 | 10 | ||||
PA7 | 10 | ||||
No PA7 | 50 |
Update:July 17, 2013
Phylip dollop parsimony tree based on an matrix of ortholog changes
Update: July 16, 2013
Treefiles based on aligned orthologs
number of genes used | Method | treefile |
100 | FASTTREE | treefile |
100 | FASTTREE | Treefile 2 |
all orthologs | Phylip | treefile_phylip |
FleN, FleQ, Flhf unique strains
Gene | alignment | tree |
fleN | fleN unique align | |
fleQ | fleQ unique align | |
flhf | flhF unique align |
To do:
- Create pipeline
- LD_Hat w/ 10 genes excluding and including PA7
Update: June 28, 2013
Phylogenetic Analysis by Maximum Likelihood (PAML) Test performed on FleN, FleQ, FlhF orthologs and aeruginosa only orthologs
Gene | PAML outfile for orthologs | |
---|---|---|
fleN | fleN PAML | |
fleQ | fleQ PAML | |
flhf | flhF-corrected PAML |
To do:
- Estimate genome tree using MrBayes and BEST (http://www.stat.osu.edu/~dkp/BEST/introduction/).
- Analysis of positively selected genes using PAML and homozygosity analysis
Update: June 18, 2013
Ortholog aligning and phylogenetic tree material and methods
Protein ortholog data: fleN, fleQ, flhF
- Protocol:
- Fasta headers are too long for tree, run: rename.pl to shorten names. Usage:
>rename.pl <FASTA_file> > <OUTPUTfilename.fas>
(will rewrite script to create automatic output file) - To align use muscle Usage:
>muscle -in <FASTA_File> -out <OUTPUTfilename> -clwstrict
- To create tree file run clustalW2 Usage
>clustalw2 -infile= <Aligned_file> -output= Phylip
- To create tree run R using following commands:
setwd("<directory containing phylip files>")
library ("ape")
library ("phangorn")
<gene_name> = read.tree("<file_gene_name.phy>")
<gene_name> = midpoint(gene_name)
plot(<gene_name>)
- Fasta headers are too long for tree, run: rename.pl to shorten names. Usage:
Gene | Alignment | Tree | Notes |
---|---|---|---|
fleN | fleN pep alignment | Conserved Domain | |
fleQ | fleQ pep alignment | Conserved Domain | |
flhf | flhF pep alignment | Conserved Domain |
Benchmark: June 11, 2013
- Finish parsing the genome files to upload the "orf" table (Raymond & Rayees)
- Rayees Parsed genome files: https://www.dropbox.com/sh/k0zktvvmv39op9i/1zBercEky8
- Parsing the ortholog file to upload the "orth_orf" table (Raymond)
- Identify and download fleN, fleQ, and flhF orthologs & align them (Rayees)