Rhodococcus opacus DSM43205 genome assembly, annotation, and metabolic model simulations

This repository contains the Rhodococcus opacus DSM43205 genome scale metabolic model in SBML format associated with the genome assembly available at the European Nucleotide Archive (ENA) project PRJEB45460. The documentation for how the model was produced is given below.

Genome Assembly

Unicycler (https://github.com/rrwick/Unicycler) was used for the hybrid (long and short read) de novo assembly of the Rhodococcus opacus DSM43205 genome. The software versions used during the Unicycler assembly are listed in the table below.

Software	version
Unicycler	0.4.6
spades	3.12.0
racon	1.3.1
NCBI-BLAST+	2.5.0
bowtie2	2.3.4.2
samtools	1.9
java	1.8.0_121
pilon	1.22

Note that, the PacBio reads and Illumina paired-end reads should be downloaded from ENA study PRJEB45460 (PacBio reads, ERR6004137; Illumina reads, ERR6004016) to be able to perform the assembly with the commands shown below.

RAW_PACBIO_READS="data/rawdata/3-E143388-gDNA_33337_subreads.fastq.gz"
RAW_ILLUMINA_R1_READS="data/rawdata/e143388_R1_deduped.fq.gz"
RAW_ILLUMINA_R2_READS="data/rawdata/e143388_R2_deduped.fq.gz"

FASTP_PACBIO_READS="results/fastp/e143388_pb_fastp.fq.gz"
FASTP_ILLUMINA_R1_READS="results/fastp/e143388_R1_deduped_fastp.fq.gz"
FASTP_ILLUMINA_R2_READS="results/fastp/e143388_R2_deduped_fastp.fq.gz"

fastp --qualified_quality_phred=20 \ 
  --dont_overwrite \
  --cut_by_quality3 \
  --cut_by_quality5 \
  --low_complexity_filter \
  --trim_poly_g \
  --trim_poly_x \
  --overrepresentation_analysis --thread 6 \
  --json=illumina_fastp.json \
  --html=illumina_fastp.html \
  -i "${RAW_ILLUMINA_R1_READS}" -I "${RAW_ILLUMINA_R2_READS}" \
  -o "${FASTP_ILLUMINA_R1_READS}" -O "${FASTP_ILLUMINA_R2_READS}"
  
fastp --dont_overwrite \
  --low_complexity_filter \
  --disable_quality_filtering \
  --trim_poly_g --trim_poly_x \
  --overrepresentation_analysis --thread 6 \
  --json=pacbio_fastp.json \
  --html=pacbio_fastp.html \
  -i "${RAW_PACBIO_READS}" \
  -o "${FASTP_PACBIO_READS}"
  
unicycler --keep 3 \
  -1 "${FASTP_ILLUMINA_R1_READS}" -2 "${FASTP_ILLUMINA_R2_READS}" \
  -l "${FASTP_PACBIO_READS}" --threads 24 -o assembly

Gene prediction and annotation

The genome annotation was performed using prokka (https://github.com/tseemann/prokka). The software versions used during the prokka annotations are listed in the table below.

Software	version
prokka	1.13.3
aragorn	1.2
barrnap	0.9
NCBI-BLAST+	2.7
cmpress, cmscan	1.1
hmmscan, hmmpress	3.2
minced	3.2
prodigal	2.6
tbl2asn	25.6
BioPerl	1.007002

prokka --outdir annotation --prefix prokka assembly.fasta

Metabolic model reconstruction

The metabolic model was reconstructed using CarveMe (https://github.com/cdanielmachado/carveme). The software versions used during the reconstruction of the model provided in this repository. Note that GLPK (GNU Linear Programming kit, https://www.gnu.org/software/glpk/) can be used as an open source alternative to the CPLEX (https://www.ibm.com/analytics/cplex-optimizer).

Software	version
carveme	1.2.2
cplex (CPLEX studio and python interface)	12.8.0.1
framed	0.5.2

Note that, the set of all protein sequences from the Rhodococcus opacus DSM43205 data set should be obtained by either performing the genome assembly and annotation as described above or by downloading the protein fasta file from ENA project PRJEB45460 (genome assembly, ERZ2486663), to be able to perform the metabolic model reconstruction.

PROTEIN_FASTA_FILE=prokka.faa
OUTPUT_SBML_FILE=e143388.xml

carve "${PROTEIN_FASTA_FILE}" -o "${OUTPUT_SBML_FILE}"

Metabolic model simulations

The code for metabolic model simulations is presented in the python notebook ROP_simulations.ipynb.