organelle-assembly

A script to assemble a plastid or mitochondrial genome from long and short reads.

To run:

conda activate bio
bash assembler.sh -b baits.fasta -g 160000 -s 40000000 R1.fasta R2.fasta nano.fq.gz

Inputs:

• -b baits file, e.g. gene sequences of related species
• -g expected genome size
• -s target bases (for Filtlong - e.g. coverage (250) x genome size)
• R1 and R2 illumina reads, already trimmed and filtered (see fastp.sh script)
• Nanopore reads, raw

How it works (abridged):

• Uses a baits file to extract nanopore organelle reads (e.g. mitochondrial or chloroplast) from all the sequencing reads (e.g. nuclear, mitochondrial, chloroplast)
• Assembles these reads (Flye); polishes assembly (Racon)
• Uses this assembly as the new baits file to re-extract nanopore organelle reads
• Assembles (Flye); polishes assembly (Racon)
• Uses this assembly as baits to extract illumnina organelle reads
• Uses these reads to polish assembly (Pilon)
• Further assemblies for comparison: Raven, Miniasm, Unicycler
• Reports read and assembly stats

Where are the results?

• The results folder has assemblies, assembly graphs, extracted reads, read-mapping bam files, and read/assembly stats.
• The script run and screen output is saved as logfile.txt

Tools:

Installed with conda.

minimap2
samtools
filtlong
flye
racon
raven
fastp
rasusa
bwa
pilon
unicycler
miniasm
minipolish
mummer
seqkit

See packagae-list.txt for full details.

Test data

Here are some links to test data.

• Syme, Anna. (2019). Chloroplast genome sequencing reads from sweet potato [Data set]. Zenodo. https://doi.org/10.5281/zenodo.3567224
• Syme, Anna. (2020). Chloroplast genome sequencing reads from snow gum [Data set]. Zenodo. https://doi.org/10.5281/zenodo.3600662