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Pseudomonas Genome Database - Overview

How to identify orthologs in different strains/species

The Pseudomonas Genome Database provides pre-computed ortholog predictions based on the Ortholuge method for validation of reciprocal best-BLAST predictions (Fulton et al., 2006) and Pseudomonas Ortholog Groups (Winsor et al., 2009).

Ortholuge

One major advantage of Ortholuge is that its algorithm compares the ratio of phylogenetic distance between two putative orthologs and that of a third gene from a more distantly-related outgroup species. If these ratios match what you would expect for the phylogenetic distance of the strains as a whole, they are more likely to be true orthologs. This technique can be very useful for distinguishing paralogs from true orthologs, which may be of concern when dealing with genome annotations that may be missing one or more annotated genes.

Some points to note with respect to different confidence classes in Ortholuge:

  1. Supporting Species Divergence (SSD) Orthologs: Ortholuge analysis determined that phylogenetic distance between the two putative orthologs supports species divergence. These proteins have a higher probability of being orthologs.
  2. The phylogenetic distance between the two putative orthologs sometimes falls within a region where Ortholuge is unable to predict if they are paralogs or orthologs. These are marked as Uncertain.
  3. Non-SSD ortholog: Unusual rate of divergence in relation to species divergence. Important Warning: These proteins have a higher chance of being paralogs.
  4. Reciprocal best-BLAST hit: No Ortholuge analysis is available to validate this RBBH hit. NOTE: (1) Not all orthologs may be identified using the Ortholuge method. An example of when this may occur is when the outgroup used in the Ortholuge analysis does not have an ortholog corresponding to an ortholog pair occurring in the two ingroups. (2) Not all putative ortholog pairs in this database have been evaluated by Ortholuge analysis (they may just simply be reciprocal-best-blast hits).

Pseudomonas Ortholog Groups (POGs)

COG classification has been used for sorting genes into orthologous groups, however many Pseudomonas genes cannot be categorized into a COG group. In order to generate a more inclusive dataset of Pseudomonas genes mapped to their putative orthologs in other Pseudomonas species/strains, we developed a Pseudomonas Orthologous Groups (POG) classification system.

This method is more useful for identifying orthologs between closely-related strains. In addition, intra-genome BLASTp analysis is utilized to acquire in-paralog information (i.e. gene duplications occurring after species divergence).

Locating orthologs from within a gene card

When starting from a specific gene card (e.g. orpF/PA1777 ), links to putative orthologs in other strains may be found either directly under the top image (Figure 4a: blue arrow) or in the Orthologs/Comparative Genomics section of the Overview tab (Figure 4b).

Figure 4a. Using the gene card as a starting point for finding putative orthologs. The blue arrow marks a link to Ortholuge-validated orthologs.

Figure 4b. On the same gene card, a link to Pseudomonas Ortholog Groups and Otholuge-validated orthologs can be found in the Orthologs/Comparative Genomics section of the Overview tab.

Navigating between species' orthologs from within GBrowse

You can navigate between species via computationally predicted, putative orthologs based on reciprocal best BLAST hits (RBBHs). When viewing the annotation for a specific gene (Figure 5a), follow the drop-down menu entry, "Putative Orthologs" to its associated GBrowse entry. Once there, scroll down until you find the Predicted Orthologs track whereby orthologs are coloured based on their Ortholuge classification. Clicking on any of the glyphs in this track will bring you to its respective gene card.

Figure 5a: Navigation between closely related Pseudomonas genomes via putative ortholog tracks in GBrowse.

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Figure 5b: Navigation between closely related Pseudomonas genomes via putative orthologs in GBrowse.

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High-throughput Identification and Comparison of Orthologs

Downloading Ortholog Sets

In addition to viewing orthologs of a specific Pseudomonas gene, it is possible to download the entire set of orthologs for a particular strain. They can be retrieved under the Orthologs tab on the Download page.

Comparing Orthologs Between Strains

By going to the Comparative Genome Search page at Database Search » Comparative Search, you can also perform a search to return orthologs (as determined by reciprocal-best BLAST) that are present in one set of genomes and absent in another set.

This can be useful for situations when you want to:

1) Find orthologs present in one set of genomes (e.g. two or more strains selected from left box)

Example:You would like to find all orthologous genes that are present in BOTH P. aeruginosa PAO1 and P. aeruginosa PA14.

Steps:

a) Select P. aeruginosa PAO1 and P. aeruginosa PA14 from the left box

b) Click on submit.

2) Find orthologs present in one set of genomes (e.g. two or more strains selected from left box) while absent from another set (e.g. one or more genomes selected from right box).

Example:You would like to find all orthologous genes that are present in BOTH P. aeruginosa PAO1 and P. aeruginosa LES but NOT present in P. aeruginosa PA7.

Steps:

a) Select P. aeruginosa PAO1 and P.aeruginosa LES from the left box

b) Select P. aeruginosa PA7 from the right box

c) Click on submit.

3) Find a list of genes in one strain/species that do not have orthologs in another strain/species

Example: You would like to find all genes in P. aeruginosa PAO1 that do not have orthologs in P. aeruginosa PA7.

Steps:

a) Select P. aeruginosa PAO1 from the left box

b) Select P. aeruginosa PA7 from the right box

c) Click on submit.

These results can be further limited by up to four boolean search fields and filtered by existence of transposon mutants and human homologs.