Argo Comparative View

  1. Overview
  2. Basic Concepts
  3. Installing and Starting Argo
  4. Downloading Sample Data
  5. Opening the Comparative View
  6. Bookmarks
  7. Perpendicular (Dot Plot) View
  8. Parallel View
  9. Biological Observations

Credits:
Tamaru Yu developed the initial version of the comparative view (ComBo).
Alan Derr developed the tutorial use case and sample data files.

Overview

This tutorial describes how to use the Argo Comparative View to view sequence alignments. Its purpose is to cover basic concepts and get you up and running with the viewer as quickly as possible. It is a how-to, not an exhaustive reference. Once you are comfortable with the basic operations, the best reference is the viewer itself; hyperlinks to specific and detailed help pages are provided from within the viewer's key menus and windows. All help pages, whether referenced within this document or from Argo itself, are also accessible from the main help page.

Basic Concepts

The Comparative View displays sequence alignments. You provide a reference sequence, one or more query sequences, and the alignments for each reference-query pair. The Comparative View displays the alignments in two ways:

Installing and Starting Argo

The Comparative View is part of Argo. This tutorial assumes that you have downloaded and started Argo.

If you haven't already, you can download argo here. No installation is required. If java 1.4 is properly set up on your system, you should be able to double click the jar file to start Argo. It may take a few seconds to start. For more detailed instructions, see the Argo User Guide.

Downloading Sample Data

For this tutorial, we'll use the Argo Comparative View to view three strains of Staphylococcus aureus:

Importance

S.aureus is the cause of many diseases, including food poisoning, various wound-related infections, and life-threatening conditions such as bacteremia, necrotizing pneumonia, and endocarditis. It is extremely successful at developing antibiotic resistance, and strains have evolved with resistance to one or more of penicillin, methicillin, and vancomycin.

Analysis

Genbank and fasta files for all three strains were downloaded from NCBI (http://www.ncbi.nlm.nih.gov/). Whole-genome comparison was performed using PatternHunter (http://www.bioinformaticssolutions.com/products/ph/). The output of PatternHunter was 'massaged' to create Argo-readable (Simple Alignment Format) alignment files. The same kind of analysis could be performed using MUMmer, BLASTZ, or other whole-genome comparison programs, with similar results.

Sample Files

Download the tutorial files from this directory, as a gzipped tar archive, or by using the links in the following table.

For your convenience, instead of downloading and opening these files individually, you may click on this link to open argo in webstart mode and automatically load these files. Note: it may take a few minutes for the data to download and display.

FileFile typeDescription
reference.gbGenbank Format Reference Sequence File (MSSA)
query_a.gbGenbank Format First Query Sequence File (MRSA)
reference_to_query_a.align Simple Alignment Format First Alignment File (MSSA to MRSA)
query_b.gbGenbank Format Second Query Sequence File (Mu50)
reference_to_query_b.align Simple Alignment Format Second Alignment File (MSSA to Mu50)

Descriptions of all supported argo file formats are provided on the main help page.

Opening the Comparative View

To open the Comparative View:

  1. Select Open Combo Files... from the combo menu. Argo displays the New Comparative View dialog box.
  2. In the reference sequence drop-down list, choose Sequence File, and then choose reference.gb (MSSA). Argo uses the file extension to determine how to interpret the file and then prompts you to confirm.
  3. When prompted to Interpret File as, accept the default (Genbank). There's a slight delay (2-3 second) while the file loads, then the filename replaces the "no data" label below the drop-down list.
  4. For Query Sequence 1, choose Sequence File and then query_a.gb (MRSA).
  5. For Alignment Data 1, choose File and then reference_to_query_a.align.
  6. To add a second query sequence, click Add another Alignment/Query Sequence.
  7. For Query Sequence 2, choose query_b.gb (Mu50).
  8. For Alignment Data 2, choose reference_to_query_b.align.

The New Comparative View dialog box should like this:

New Comparative View dialog

When you click OK, after a slight delay (10-15 second), your screen should look like this:

New Comparative Perpendicular View

Bookmarks

Specifying numerous sequence and alignment files can be tedious. To avoid doing it more than once, use the Bookmarks menu. After opening the Comparative View, select Add Bookmark from the Bookmarks menu. Argo adds a new entry for your comparative view to the Bookmarks menu for use in future sessions. Alternatively, you can use the Open Recent Sequence command on the File menu to display recent Comparative Views.

Perpendicular (Dot Plot) View

When you open the Comparative View, you first see the perpendicular view, as shown above. The reference sequence appears horizontally on the bottom. The query sequences are shown vertically on the left. The glyphs in the dot plot represent the alignments; they are color-coded to match your two query sequences.

The reference and query sequences are displayed in Argo feature maps. Click on a feature in a feature map or a glyph in the dot plot to highlight it in all panels and to view it in the inspector.

Hold the shift key down and left click to zoom in on a feature map or the dot plot, right click to zoom out. To scroll, use the scroll bars or drag the mouse while holding the alt key and the left mouse button (middle button for a three-button mouse). Zooming and scrolling are synchronized across panels.

Parallel View

The perpendicular view is useful for viewing large sequence regions. For a detailed comparison of smaller segments, open the parallel view:

  1. In the perpendicular view, zoom in to an area of interest.
  2. Right click in the dot plot and choose Parallel View from the menu.

Argo opens the parallel view. If you open a parallel view from a perpendicular view that contains multiple query sequences, Argo displays a separate parallel view for each query sequence (each displayed in its own tab).

In the parallel view, the reference sequence is shown on the bottom and the query sequence on the top. Alignments are drawn as a web connecting the two feature maps. Each feature map can be zoomed and scrolled independently. The alignment web will stretch as appropriate. If you select a feature or an alignment, the corresponding regions in the other panels are highlighted.

If the alignment panel is too busy, select alignment glyphs that are especially interesting (or uninteresting), right click and choose Hide Others (or Hide Feature). Another right click option, Show Hidden Features makes them visible again.

Biological Observations

  1. A phage is located in the vicinity of 995Kb (981,024-1,025,750bp) of the reference strain, MSSA. The comparative display clearly shows the translocation (and inversion) of this phage to around 1.6Mb (1591597-1637496bp) in the MRSA strain. In addition, it appears to be completely absent from the Mu50 strain, although some possible remnants can be seen in the region of 917-955kb of Mu50.
  2. In the region of 409-424Kb of MRSA, a "putative pathogenicity island" can be seen, which is absent in both MSSA (the reference) and Mu50.

Last Updated: Sept 18 2006
Contact: Reinhard Engels
argo-support@broad.mit.edu