MISMS MinION Workshop – Colombia

Training Workshop for the Implementation of MinION Sequencing for Influenza Viruses 

February 6-8, 2020

Irotama Resort

Santa Marta, Colombia

Workshop Objective

This three-day workshop, held February 6-8th, 2020 in Santa Marta, Colombia, introduced new technologies and advanced informatics approaches that can be used for infectious disease genomic characterization in resource-limited settings. Using the portable, low-cost Oxford Nanopore MinION sequencing platform, we will introduce the essential laboratory and computational skills required to perform extensive genetic characterization of viral pathogens, using influenza A virus as a primary demonstration. Topics will include sample preparation, bioinformatics, and phylogenetic approaches to characterize newly sequenced sampled against publicly available data.

By the completion of the workshop, attendees were provided access and training to all essential bioinformatics tools required to establish real-time genetic characterization capabilities in their primary laboratories or at field sites using the low-cost Oxford Nanopore MinION sequencing platform. Best practices for implementing these new technologies in research, clinical, and public health laboratories were emphasized.

 

Instructors/staff:

Johns Hopkins University: Tom Mehoke, Peter Thielen

Fogarty International Center: Martha Nelson, Nidia Trovao

 

Workshop Agenda

Day 1 (Thursday Feb 6) 

Introduction to Genomics Technologies and Oxford Nanopore Sequencing

 

Time Event Presenter
9:00am Welcome and Introductions

  • Pre-session surveys
Peter Thielen
9:30am Low-Resource Genome Sequencing with Oxford Nanopore: Outbreak response and Influenza Applications  Peter Thielen
10:00am Data Analysis Objectives: Rapid Characterization and Consensus Sequence Generation Peter Thielen
11:00am BREAK
11:15am Rapid Talks: Single Slide Presentations from Attendees Attendees
12:00pm Lunch / Surveillance Meeting Field Trips

 

Day 2 (Friday Feb 7):

Laboratory Considerations and Basic Bioinformatics

 

Time Event Presenter
9:00am Practical Considerations: Establishing a Genomics Wet Laboratory

  • Efficient isolation of nucleic acids (DNA/RNA)
  • Targeted and untargeted sequencing approaches
  • Accurate quantification of DNA
  • Minimizing contamination
Peter Thielen
9:30am Concepts: Optimized Influenza A Virus Sample Preparation with Ligation reagents

  • Time- and reagent-optimizing considerations
  • Sequencing of influenza multi-segment PCR amplicons
Peter Thielen
10:00am BREAK
10:15am Hands on: Sequencing with the MinION Peter Thielen
11:45pm Data Manipulation and Exploratory Data Analysis

  • Complete consensus generation on generated data
Tom Mehoke
12:15pm Lunch
1:15pm Introduction to Phylogenetics Concepts

  • 101 talks on principles of phylogenetics
  • Common mistakes
Martha Nelson
2:15pm Using data produced by the MinION

  • Outline of general analysis workflow
  • Initial processing vs. in depth analysis
Tom Mehoke
2:45pm BREAK
3:00pm Hands-on: Processing data produced so far

  • Visualization of processed data
  • Introduction to folder hierarchy
  • Consensus sequence generation
Tom Mehoke
5:00pm Day 2 Review

Revisit data from the earlier sequencing run

Recap of topics

Post-session evaluations

 

Day 3 (Saturday Feb 8):

Phylogenetics

 

Time Event Presenter
9:00am Identifying Genomes and Building a Genetic Sequence Dataset

  • Genbank
Nídia Trovão
10:00am Multiple Sequence Alignment and Building a Phylogenetic Tree

  • AliView
  • MEGA
Nídia Trovão
11:00am BREAK
11:15am Post-processing of Phylogenetic Trees

  • FigTree
Martha Nelson
12:15pm Lunch
1:15pm How To: Interpret a Phylogenetic Tree Martha Nelson
2:15pm Final Recap

  • Open Discussion: Moving forward with resource development and applied research
  • Post-session evaluations
3:00pm BREAK
3:15pm Time to Work With Your Own Data All instructors available for questions