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Microbiome Bioinformatics Analysis Workshop

General Information

Led by: ArrietaLab

Location: Oswaldo Cruz Foundation - Rio de Janeiro, Brazil

Dates: 3-5 April 2023

Objective: Analyzing microbial DNA sequences

Requirements:

  • Laptop with a Mac, Linux, or Windows operating system (not a tablet, Chromebook, etc.) with administrative privileges
  • Access to Wifi
  • R and Rstudio installed (instructions below)
  • Excel or any text editor installed (e.g. TextWrangler, Notepad, BBEdit, etc.)

Contact: mona.parizadeh@ucalgary.ca

Schedule

This workshop covers the following material:

Day 1

(9 a.m. to 12 p.m. & 1 p.m. to 4 p.m.)

  • Introduction to metabarcoding and amplicon sequencing
  • Introduction to R
  • DADA2 Tutorial

Day 2

(9 a.m. to 12 p.m. & 1 p.m. to 4 p.m.)

  • Introduction to phyloseq package in R
  • Data exploration
  • Statistical analyses of 16S rRNA gene sequences:
    • Taxonomic composition
    • Alpha diversity
    • Beta diversity (ordination)
    • PERmutational Multivariate ANalysis Of VAriance (PERMANOVA)
    • Differential analysis
  • Discussion
    • Wrap up of the results of 16S rRNA gene sequence analysis
    • Differences between DADA2 pipeline workflows in analyzing 16S vs ITS sequences

Setup

To participate in this Workshop, please install the following software and packages, complete the R courses for beginners, and let us know if you need any help before attending.

Install R and RStudio

R is a free and open-source programming language that is particularly powerful for data exploration, visualization, and statistical analysis. We use RStudio to interact with R.

Windows

Please download R for Windows from CRAN to install R, and also install the RStudio IDE. If you have separate user and admin accounts, please run the installers as an administrator by right-clicking on the .exe file and selecting "Run as administrator" instead of double-clicking. Otherwise, problems may arise later when installing R packages.

Video Tutorial

Mac OS X

Please download R for macOS from CRAN to install R, and also install the RStudio IDE.

Video Tutorial

Linux (Debian, Fedora/Redhat, Ubuntu)

Please download the binary files for your distribution from CRAN to install R, or use a package manager (e.g. run sudo apt-get install r-base for Debian/Ubuntu and run sudo yum install R for Fedora/Redhat). Additionally, please install the RStudio IDE.

R for beginners

To follow the workshop, you must have a basic understanding of R. Before attending the workshop, please go through the following courses:

Install R packages

Please install the following R packages in RStudio:
  • dada2
  • phyloseq
  • vegan
  • ggplot2
  • tidyverse (or tidyr): Tutorial by Data Carpentry
  • dplyr
  • BiocGenerics
  • SummarizedExperiment
  • rstatix
  • DESeq2
  • ggpubr
  • RColorBrewer
  • gridExtra

Download database for DADA2

Please download the silva_nr99_v138.1_wSpecies_train_set.fa.gz file for taxonomic assignment with DADA2.

BLAST

Sequence Example