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Using Dotplots for Comparative Genomic Analysis

By Andrew Kapinos1, Canela Torres1, Amanda Freise1, Sean McClory2, Kathleen Cornely3

1. UCLA 2. La Salle University 3. Providence College

In this activity, students are introduced to dotplots as a tool for analyzing phage genomic similarity.

Listed in Teaching Materials | resource by group HHMI Science Education Alliance (SEA) Faculty Group

Version 1.0 - published on 24 Aug 2020 doi:10.25334/RR9F-HD75 - cite this

Licensed under CC Attribution-ShareAlike 4.0 International according to these terms

Dot Plots.jpg


In this activity, students are introduced to dotplots as a tool for analyzing phage genomic similarity. An explanation of how dotplots are constructed is provided.  A pre-activity assignment reviews dotplot basics and requires students to practice analyzing pre-made dotplots. A final assessment requires students to develop their own research questions about phage similarity and to construct a dotplot to answer them.

Learning objectives

After completing this module, students will be able to…

  • Prepare concatenated FASTA files in the correct order.
  • Understand the purpose of different word lengths.
  • Use Gepard to generate high quality dotplot images that are labeled correctly and annotated in an informative and aesthetically pleasing manner.
  • Use dotplots to determine similarity or dissimilarity between two or more phages.
  • Interpret the dotplot results and draw conclusions about genomic relationships.
  • Identify repeat sequences and other features.
  • Suggest cluster relationships based on dotplot results.
  • Generate a hypothesis [make a prediction] to be tested with a dotplot.


How is the module structured to promote student development as a scientist? 

This activity supports elements important for student development as scientists in the following ways:

  • The activity introduces software (Gepard) used by practicing scientists who publish the results of their analyses using this tool in scientific publications.
  • The module provides students with the theoretical background necessary to construct and interpret dotplots.
  • Students are required to work in groups.
  • Students are asked to explain their answers, and to defend their answers to their peers.
  • One of the assessments involves an independent project in which students are required to use the dotplot tool to investigate their own research questions. Students are required to seek peer review before submitting their final project.


Intended Teaching Setting

Course level:  First-year and second-year science majors

Instructional Setting:  In-person classroom or online

Implementation Time Frame:

  1. Students complete the “Pre-Activity” assignment, either for homework or at the beginning of a laboratory period. Before proceeding to the activity, the facilitator assesses student comprehension and answers questions.
  2. The facilitator distributes the Learning Activity Outline and the Learning Activity Instructions. Students are instructed to install Gepard on their computers.
  3. The facilitator presents the “Using Gepard to Make Dotplots” PowerPoint presentation.
  4. Students construct their own dotplots using Gepard as described in the Learning Activity Instructions.
  5. The facilitator presents the “Annotating Dotplots Constructed Using Gepard” PowerPoint presentation.
  6. When the students have completed constructing and annotating their dotplots, the class concludes with the facilitator describing the assessment requirements to the students. If time permits at the end of the period, students can brainstorm project ideas with each other. The assessment may be completed outside of class for homework.
  7. During the next class period, students present their projects to the class.


Project Documents

Facilitator documents:

Using Gepard to make dotplots.pptx

Annotating dotplots constructed using Gepard.pptx

Dotplot_Part A_Pre-activity Answer Key

Smith KC, Castro-Nallar E, Fisher JN, Breakwell DP, Grose JH, Burnett SH. Phage cluster relationships identified through single gene analysis. BMC Genomics. 2013;14:410. Published 2013 Jun 19. doi:10.1186/1471-2164-14-410 (can be accessed at

Learning activity documents:

Dotplot_Part A_Pre-activity

Dotplot Learning Activity Outline

Dotplot_Learning Activity Instructions

Assessment documents:

Dotplot_Part B_Post activity assessment


Facilitator Instructions

  1. OPTIONAL: Read the Smith et al. (2013) paper for context.
    1. The Smith paper covers the use of different types of dotplots, and different input data, for the purposes of clustering mycobacteriophages. Dotplots are frequently used to analyze inter- and intra-cluster similarities, and this paper provides a useful overview of that purpose.
  2. Have students read “Dotplot Learning Activity Outline.
  3. Present “Using Gepard to make dotplots”: slides 1-6
    1. This is a very brief introduction to the concept of dotplots. Good time to allow questions for clarification.
  4. Have students complete “Dotplot_Part A_Pre-activity.
    1. Check for comprehension. Answer student questions.
  5. Ask students to install Gepard using “Dotplot_Learning Activity Instructions” file.
    1. This can be done outside of class time.
  6. Give the remainder of the “Using Gepard to make dotplots” presentation.
    1. Demo creating a dotplot in real time and/or use the presentation to walk students through the process.
    2. The presentation file can also be given to students as an instructional resource.
  7. Give the “Annotating dotplots constructed using Gepard” presentation.
    1. The presentation file can also be given to students as an instructional resource.
  8. Have students complete the “Dotplot_Part B_Post activity assessment” activity.
    1. Recommended: Include brainstorming session for students to come up with good questions. Review student questions to encourage development of testable hypotheses.
  9. Ask students to present questions and dotplots to each other.


Cite this work

Researchers should cite this work as follows:


HHMI Science Education Alliance (SEA) Faculty Group

HHMI Science Education Alliance (SEA) Faculty Group group image

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