viral-ampliseq-assembly v1.0.0

Initial release (v1.0.0) of the viral-ampliseq-assembly Snakemake workflow for analysis and assembly of viral genomes from IonTorrent AmpliSeq data.

This workflow is especially suited for analysis of Classical Swine Fever Virus (CSFV) IonTorrent Ampliseq sequence data.

See the GitHub release page for more info

Workflow Overview

Preprocessing

• Duplicate reads were removed using Picard
• Reads were trimmed with Trimmomatic prior to SPAdes assembly
• BAM file stats computed using Samtools (coverage depth, extent, extent per genome, # of reads mapped)

Reference Genome Selection

• Downloading of all Classical swine fever virus (CSFV) (or FMDV, Ebola, Zika) virus genomes from NCBI Entrez API using BioPython
• Mash screen of deduplicated reads against all reference genomes with sketch size of 10000 and sketch k-mer size of 16, sorting by Mash screen identity to find top reference genome for read mapping and variant calling

Read Mapping & Variant Calling

• Read mapping with BWA MEM
• Removal of duplicate reads with Samtools
• Variant calling with FreeBayes
• SnpEff was used to predict and report variant effects using reference genome annotation

De Novo Assembly

• SPAdes de novo assembly of trimmed deduplicated reads.
• QUAST quality assessment of assemblies

Quality Control

• MultiQC interactive report of FastQC, Samtools, QUAST, SnpEff

Phylogenetic Tree

• Phylogenetic tree constructed with IQ-TREE (or Clearcut if a quick and dirty tree is okay)
• Interactive HTML phylogenetic tree visualization with PhyloCanvas using shiptv