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Metagenomics Summer School

Day
Lesson topic
Day 1: Assembly 1. (Pre-Summer School) Introduction I: Shell
2. Introduction II: HPC and job scheduler
3. Filter raw reads by quality
4. Assembly I: Assembling contigs
5. Assembly II: Variable parameters
6. Assembly evaluation
Day 2: Binning 1. Introduction to binning
2. Binning with multiple tools
3. Bin dereplication
4. Manual bin refinement
Day 3: Annotation 1. Assigning taxonomy to refined prokaryotic bins
2. Phylogenomics
3. Identifying viral contigs in metagenomic data
4. Gene prediction
5. Gene annotation I: BLAST-like and HMM
6. Gene annotation II: DRAM and coverage calculation
Day 4: Visualisation 1. Gene annotation III: DRAM distillation
2. Introduction to data presentation
3. Coverage heatmaps
4. Ordinations
5. KEGG pathway maps
6. Gene synteny
7. CAZy heatmaps


Timetable 2023
Time
Event
Session leader
09:00 – 09:30 Introductions
Welcome
Overview
Login to NeSI via Jupyter Hub		 Jian Sheng Boey
09:30 – 10:30 TASK Bash scripting and Slurm scheduler I Dinindu Senanayake
10:30 – 10:50 Morning tea
10:50 – 11:10 TASK Bash scripting and Slurm scheduler II Dinindu Senanayake
11:10 – 11:45 TALK Metagenomics decision tree Kim Handley
11:45 – 12:00 TALK Project introduction and description Kim Handley
12:00 – 12:45 Lunch
12:45 – 13:45 Read QC
TALK Quality filtering raw reads
TASK Visualisation with FastQC
TASK Read trimming & adapter removal
TASK Diagnosing poor libraries
TALK Common issues & best practices
TASK (Optional) Filter out host DNA		 Annie West
13:45 – 15:00 Assembly
TALK Choice of assemblers, parameter considerations, and when to stop!
TASK Prepare data for assembly
TASK Exploring assembler options
TASK Submit assembly jobs to NeSI via Slurm
TASK (Optional) Submit variant assembly jobs		 Kim Handley
15:00 – 15:20 Afternoon tea
15:20 – 17:00 Assembly
TALK Future considerations: co-assembly vs. single assemblies
TASK Assembly evaluation
TASK Short contig removal		 Mike Hoggard
Time
Event
Session leader
09:00 – 09:15 Introductions
Recap of day 1
Overview of the day		 Annie West
09:15 – 10:15 Binning I
TALK Overview of binning history, key parameters, and strategies I
TASK Read mapping		 Kim Handley
10:15 – 10:45 Binning II
TASK Multi-binning strategy with MetaBat and MaxBin		 Mike Hoggard
10:45 – 11:00 Morning tea
11:00 – 11:40 Binning II (continued)
TALK Overview of binning history, key parameters, and strategies II		 Kim Handley
11:40 – 12:00 DISCUSSION Binning strategies
TASK Compare and contrast outputs of binning tools		 Annie West
12:00 – 12:45 Lunch
12:45 – 14:00 Binning III
TASK Bin dereplication using DAS_Tool
TASK Evaluate bins using CheckM		 Kim Handley
14:00 – 14:30 Binning IV
TALK Additional dereplication strategies such as dRep
TALK Working with viral and eukaryotic bins
TALK Organisms possessing minimal genomes		 Mike Hoggard
Kim Handley
14:30 – 15:00 TALK Bin refinement: Strategies Mike Hoggard
Kim Handley
15:00 – 15:20 Afternoon tea
15:20 – 16:00 TASK Bin refinement: VizBin Jian Sheng Boey
Mike Hoggard
16:00 – 16:15 PRESENTATION Emilie Gios - Genetic exchange in ultra-small Patescibacteria Kim Handley
16:15 – 17:00 Wrap up
TASK Submit jobs for VirSorter2 and CheckV
Summary of the day		 Mike Hoggard
Kim Handley
Time
Event
Session leader
09:00 – 09:20 Introductions
Recap of day 2
Recap of binning outputs
Overview of the day		 Jian Sheng Boey
09:20 – 10:30 Taxonomic annotation of prokaryotic bins
TALK Taxonomic classification: Bin and species determination
TASK Taxonomic classification using GTDB-Tk
TASK View phylogenetic trait distribution using ANNOTREE		 David Waite
10:30 – 10:50 Morning tea
10:50 – 11:20 Phylogenomics
TALK Primer to phylogenetic analyses
TASK Build a phylogenetic tree using FASTTREE
TASK Visualise tree using iTOL		 Valter Almeida
11:20 – 12:00 Viruses
TALK Identifying viruses from metagenomic data
TASK Identify viral contigs using VirSorter2
TASK QC of viral contigs using CheckV
TALK Predicting viral taxonomy using vConTACT2
TASK (Optional) Visualise vConTACT2 viral gene sharing network in Cytoscape		 Mike Hoggard
12:00 – 12:45 Lunch
12:45 – 13:30 Gene prediction
TALK Tools for predicting genes: Prodigal, RNAmer, Aragorn, etc.
TASK Predict ORFs and protein sequences using Prodigal		 David Waite
13:30 – 14:00 Gene annotation I
TALK Primer on annotation method
TASK Annotate predicted genes using DIAMOND and HMMER3
DISCUSSION Evaluate the quality of gene assignments		 Jian Sheng Boey
14:00 – 15:00 Gene annotation II
TALK Using online resources (KEGG, BioCyc, MetaCyc, HydDB, PSORT)
TASK View KEGG annotations online		 Jian Sheng Boey
15:00 – 15:20 Afternoon tea
15:20 – 15:35 PRESENTATION Kitty Sriaporn - Genomic adaptations enabling Acidithiobacillus distribution across wide-ranging hot spring temperatures and pHs Kim Handley
15:35 – 16:15 Gene annotation III
TASK MAG annotation with DRAM
TASK Coverage calculation using Bowtie2		 Annie West
Mike Hoggard
16:15 – 16:30 Group project
TASK Group formation
TASK Topic selection		 Kim Handley
16:30 – 17:00 Wrap up
Summary of the day		 Jian Sheng Boey
Time
Event
Session leader
09:00 – 09:15 Introductions
Recap of day 3
Overview of the day		 Jian Sheng Boey
09:15 – 10:00 DRAM outputs
TALK Overview of DRAM results
TASK Explore DRAM results		 Annie West
10:00 – 10:30 Environmental distribution
TALK Visualising findings
TASK Coverage heatmap and nMDS ordination		 Mike Hoggard
10:30 – 10:50 Morning tea
10:50 – 11:00 TASK Workshop survey
11:00 – 12:00 Genomic features and metabolism
TALK Visualising findings
TASK KEGG metabolic maps
TASK Gene synteny		 Jian Sheng Boey
Annie West
12:00 – 12:45 Lunch
12:45 – 14:30 Group work
TASK Analyse data
TASK Prepare presentations		 Kim Handley
Mike Hoggard
David Waite
14:30 – 15:00 Group work
TASK Informal group presentations		 Kim Handley
Mike Hoggard
David Waite
15:00 – 15:20 Afternoon tea
15:20 – 15:40 Group work
TASK Informal group presentations		 Kim Handley
Mike Hoggard
David Waite
15:40 – 16:00 Wrap up
Final discussions


Appendices
Appendix ID
Appendix 1 Dereplicating data from multiple assemblies
Appendix 2 Generating input files for "VizBin" from "DAS_Tool" curated bins
Appendix 3 Normalise per-sample coverage values by average library size
Appendix 4 Viral taxonomy prediction via vContact2
Appendix 5 Preparing input files for visualising gene synteny


Post-workshop survey

Thank you for attending Metagenomics Summer School 2023! We would like your feedback on how we have done and what we can improve on. You can provide feedback here.


License

Genomics Aotearoa / New Zealand eScience Infrastructure / University of Auckland Metagenomics Summer School material is licensed under the GNU General Public License v3.0, 29 June 2007. (Follow this link for more information)


Slides for workshop

You can find a copy of the slides presented during the workshop, with published figures removed, in the slides/ folder.


Snapshots of results to download

If you are having trouble downloading files using scp, we are providing exemplar output files, which you can download through your browser, here, or via the following links: