A reproducible workflow for bacterial whole genome sequencing (WGS) upstream analysis. This repository is aligned with the abstract workflow for Pseudomonas aeruginosa strain 2025SY-00129 (SRR33893847), using the Pseudomonas aeruginosa strain-2507 reference for quality control, trimming, mapping, variant calling, de novo assembly, assembly assessment, and annotation.
Project Structure
All scripts should be run from the base directory:
bacterial-upstream-analysis
Tools & Conda Environments
Each step requires a separate conda environment with the listed tool(s) installed using the Bioconda channel:
| Step | Tool(s) | Conda Env Name |
|---|---|---|
| Download raw reads | sra-tools |
ncbi-sra |
| Quality control | fastqc |
fastqc |
| QC summary | multiqc |
multiqc |
| Trimming | trimmomatic |
trimmomatic |
| Mapping and variant calling | bwa, samtools, freebayes, tabix, bcftools |
bwa-sam |
| Assembly | spades |
spades |
| Assembly evaluation | quast |
quast |
| Annotation | prokka |
prokka |
Setup Conda Environments
Create separate environments for each step/tool by running:
conda env create -f envs/ncbi-sra.yml conda env create -f envs/fastqc.yml conda env create -f envs/multiqc.yml conda env create -f envs/trimmomatic.yml conda env create -f envs/spades.yml conda env create -f envs/quast.yml conda env create -f envs/prokka.yml conda env create -f envs/mapping.yml
Run a Script
./scripts/script_name.sh
Data Input
- SRA accession list:
data/accession.txt - Reference genome path:
data/reference/reference.fasta - Active reference:
NZ_CP141269.1 Pseudomonas aeruginosa strain 2507 chromosome, GI2647842840
Script Execution
Run the scripts in this order:
00_download_sra.sh- Download raw reads from Sequence Read Archive01_fastqc.sh- Run quality control on raw reads02_multiqc.sh- Summarize QC reports03_trimmomatic.sh- Trim low-quality reads04_mapping.sh- Map trimmed paired reads to the Pseudomonas reference genome05_variant_calling.sh- Call variants against the reference genome06_assembly.sh- Perform de novo genome assembly07_quast.sh- Evaluate the assembly08_prokka.sh- Annotate the genome
Included Outputs
- Dataset accession:
SRR33893847 - QUAST assembly report:
results/quast/report.txt - Annotation summary:
results/annotation/SRR33893847.txt - Variant calls:
results/vcf/SRR33893847.vcf.gz - Assembly outputs:
results/assembly/contigs.fastaandresults/assembly/scaffolds.fasta - QC reports:
results/qc/andresults/multiqc/multiqc_report.html
Access to Large Data Files
Due to GitHub's file size limits, raw FASTQ files, trimmed reads, and mapping output files such as .fastq, .bam, and .sam are hosted on Google Drive.
Access the large data files on Google Drive