Overview of SEDNA users’ scripts and workflows

Who is doing what?

Phil Morin:

• reference-based genome assembly, genome-wide heterozygosity, historical demography (PSMC), genome characterization (BUSCO, QUAST), mitogenome assembly (reference based, de novo).

Eric Anderson

• lcWGS workflows for trimming, mapping, variant calling with GATK, and also using the indels discovered therein to do indel realignment on overlap-clipped BAM files to make BAMs ready for ANGSD processing.
• microhaplotype genotyping.
• Association with ANGSD.
• Population assignment/GSI with genotype likelihoods.

Bionformatic Topics

Whole Genome Sequencing:

• Eric Anderson’s Snakemake-based whole genome sequencing workflow for non-model organisms. (FastQC, Trimmomatic, bwa mem, MarkDuplicates, HaplotypeCaller, GenotypeGVCFs to get a BCF file, then bamutils clipOveraps and GATK 3.8 Indel Realignment to produce ANGSD-ready BAMs a al Nicolas Lou, Nina Therkildsen, et al. https://github.com/eriqande/mega-non-model-wgs-snakeflow

• Phil Morin’s genome-wide heterozygosity, 1MB non-overlapping windows (ANGSD, R): https://github.com/PAMorin/Genome_wide_heterozygosity_ANGSD

SNP discovery from single and multiplex genome assemblies

• Scripts from Morin et al. 2018, SNP discovery from single and multiplex genome assemblies of non-model organisms. In: Head, S.R., Ordoukhanian, P., Salomon, D. (Eds.), Next-Generation Sequencing. Methods in Molecular Biology. Humana Press, pp. 113-144.: https://github.com/PAMorin/SNPdiscovery

Microhaplotype genotyping

• The SWFSC MEGA Microhaplotype workflow: https://github.com/eriqande/mega-simple-microhap-snakeflow

Processing BCF files to ANGSD and other programs

• Eric’s workflows for post-BCF processing (PCA and association at the moment): https://github.com/eriqande/mega-post-bcf-exploratory-snakeflows

Computing Pairwise Fst from lcWGS data using ANGSD and winsfs

• Eric put this together and the folks in the MEGA have used it for a number of species/projects.: https://github.com/eriqande/mega-lcwgs-pw-fst-snakeflow

Phase chromosomes of WGS data

• Eric threw https://github.com/eriqande/phase-chromosomes-snakeflow together at one point when he was contemplating preparing some high coverage data for use with Leo Speidel’s RELATE program, that takes phased variant data to infer genealogies and then do inference from them.

Population assignment from lcWGS data

• Eric collaborated with Matt DeSaix at Colorado State University to put this together.
  • repo: https://github.com/mgdesaix/WGSassign
  • methods paper: https://besjournals.onlinelibrary.wiley.com/doi/full/10.1111/2041-210X.14286
  • application in redstarts: https://onlinelibrary.wiley.com/doi/full/10.1111/mec.17137

Annotating with SnpEff from a GFF file

• Eric put something together related to a guest lecture in a class at CSU. It shows a small Snakemake solution for it: https://github.com/eriqande/annotate-millet-variants-snakeflow

Download sequences from NCBI using fasterq-dump

• https://github.com/eriqande/fasterq-dump-snakemake-example

Making a FASTA file of ancestral bases

• Such a FASTA file is something that ANGSD requires when you want to polarize allele frequency spectra as derived and ancestral. One way to do it is to use the sequence of a closely related species (like using chimpanzee bases as the ancestral ones for human—as done in the ANGSD examples). The probably is that the ancestral sequence has to be perfectly concordant (in terms of number of bases, etc) with your focal species reference genome. This involves a fair bit of aligning and post-processing. There must be an easier way to do this, but here is one solution: https://github.com/eriqande/make-ancestral-fasta-snakeflow

Visualizing Missing data in RAD seq data, etc

• A simple R package that uses 012 files and some compiled code to quickly generate some summaries of missing data in individuals and at different loci. It also provides an interface to SNPRelate for PCAs and calculating Fst. https://github.com/eriqande/genoscapeRtools.

Historical demography from a single genome (PSMC)

• bash script to generate a consensus diploid genome from a bam file, and run PSMC (plus bootstrap) for historical demography. https://github.com/PAMorin/PSMC_boostrap_231122

Bioinformatically useful R packages

• ecaRbioinf: A few useful functions in a package:
  • Repo: https://github.com/eriqande/ecaRbioinf
  • Online function reference: https://eriqande.github.io/ecaRbioinf/reference/index.html

Using SEDNA topics

Work interactively from your local RStudio on SEDNA’s R

• A little R package Eric wrote to make this easier to do: https://github.com/eriqande/rView Don’t know if it still works, or if some of the packages it depends on have changed enough to break it!