DNA Methylation Analysis Central Working Document

DNA Methylation Sequencing Analysis Central Document

This will be my working document to connect all the datasets and different analysis pipelines together. I’m working with 2 datasets for my own dissertation: Holobiont Integration and KBay Bleaching Pairs, and a separate project on oyster nutrition.


Diagram reference: Creative Biomart: Epigenetics


Diagram reference: Labster Theory

Projects

Holobiont Integration (WGBS)

Adult Pocillopora acuta fragments from Kaneohe Bay, Hawai’i.

Github repo link: here.

How do DNA methylation patterns in Pocillopora acuta:

  • Change with chronic stress treatments: temperature and ocean acidification?
  • Is there a core methylation pattern regardless of environmental change?

How do DNA methylation patterns in Symbiodinaeceae Cladocopium spp.:

  • Change with chronic stress treatments: temperature and ocean acidification?
  • Is there a core methylation pattern regardless of environmental change?

Holobiont Integration WGBS pipeline

Diploidy vs Triploidy: Holobiont Integration (WGBS)

Adult Pocillopora acuta fragments from Kaneohe Bay, Hawai’i.

No github repo link yet.

How are DNA methylation patterns in Pocillopora acuta impacted by ploidy status?

No pipeline link yet.

KBay Bleaching Pairs (WGBS)

Adult Montipora capitata fragments from Kaneohe Bay, Hawai’i.

Github repo link: here.

  • How do DNA methylation patterns change during seasonal changes (July vs. December)?
  • How do DNA methylation patterns change pre- and post- bleaching event (July vs. December)? This may be hard to distinguish from the above question of seasonality..
  • Are there particular DNA methylation patterns that are associated with ‘stronger’ phenotypes?

KBay WGBS pipeline

Point Judith, Rhode Island (MBD-BS)

Github repo link: here.

Adult C. virginica oysters from gut tissue. How nutrient enrichment change (or not change) DNA methylation patterns?

Cvir MBD-BS pipeline.

Resources on DNA methylation analysis

  • NF-Core methylseq pipeline: https://nf-co.re/methylseq/1.5/usage. This webpage from nf-core has been extremely helpful in learning all the flags and necessary parameters to use.
  • Bismark docs: http://felixkrueger.github.io/Bismark/Docs/.
  • Strand and Wong DNA methylation analysis guide: https://github.com/Putnam-Lab/Lab_Management/blob/master/Bioinformatics_%26_Coding/Workflows/Methylation_QC.md
  • Wong (Porites asteroides) pipeline: https://github.com/kevinhwong1/Thermal_Transplant_Molecular/blob/main/scripts/Past_WGBS_Workflow.md
  • Becker presentation from lab meeting 2021 on methylseq: https://docs.google.com/presentation/d/1qNMI2-LmyvqwNZ4J8FaHIg7d-niXm5jxEeeL1aobXeI/edit#slide=id.p4
  • Becker WGBS workflow (Pocillopora verrucosa): https://github.com/hputnam/Becker_E5/blob/master/Bioinformatics/Workflows/Becker_WGBS_Workflow.md
  • Methods comparison (Trigg et al 2021): https://onlinelibrary.wiley.com/doi/pdf/10.1111/1755-0998.13542.

Reference Genomes Used

  • Pocillopora acuta: http://cyanophora.rutgers.edu/Pocillopora_acuta/
  • Porites asteroides: https://www.biorxiv.org/content/10.1101/2022.07.01.498470v1.abstract
  • Pocillopora verrucosa: http://pver.reefgenomics.org/download/Pver_genome_assembly_v1.0.fasta.gz; Buitrago-Lopez et al 2020
  • Montipora capitata: http://cyanophora.rutgers.edu/montipora/
  • Crassostrea virginica: https://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/002/022/765/GCF_002022765.2_C_virginica-3.0/GCF_002022765.2_C_virginica-3.0_genomic.fna.gz

Before using a reference genome, be sure to double check if there is a newer version released and how that will affect your pipeline.

Lab work / Pre-bioinformatic processing

Whole Genome Bisulfite Sequencing

  • Pico Methyl Seqeuncing Kit testing: https://github.com/emmastrand/EmmaStrand_Notebook/blob/master/_posts/2020-10-21-WGBS-Pico-Methyl-Seq-Test-Run.md
  • Holobiont Integration lab work (WGBS): https://github.com/emmastrand/EmmaStrand_Notebook/blob/master/_posts/2021-04-08-HoloInt-WGBS-Sample-Processing.md
  • KBay Bleaching Pairs lab work (WGBS; done by M. Schedl from Putnam Lab): https://github.com/emmastrand/EmmaStrand_Notebook/blob/master/_posts/2021-05-18-KBay-Dec-July-2019-WGBS.md

Methyl Binding Domain Bisulfite Sequencing

  • Protocol from M. Schedl in the Putnam Lab (work done for Cvir project): https://github.com/hputnam/Cvir_Nut_Int#m-schedl-mbdbs-library-preps
Written on August 24, 2022