| name | bowtie2 |
| description | Gapped short-read alignment to contigs/genomes for coverage used in binning and abundance. Use when indexing assemblies, mapping cleaned PE reads, choosing sensitivity presets, QC of BAM rates, or preparing depths for MetaBAT2/CoverM/BASALT. Upstream: https://github.com/BenLangmead/bowtie2. Alternatives: strobealign, bwa-mem2. Always pair with samtools.
|
| license | MIT |
| category | analysis-tools |
| tags | ["mapping","alignment","short-read","coverage"] |
| upstream | https://github.com/BenLangmead/bowtie2 |
| stage | mapping |
Bowtie 2
Upstream: BenLangmead/bowtie2
Citation
Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2.
Nat. Methods 9, 357–359 (2012). https://doi.org/10.1038/nmeth.1923
Analytical thinking
Mapping is the bridge from assembly → binning. Coverage profiles (not raw
reads) drive MetaBAT2/MaxBin2/CONCOCT/SemiBin2 and inform BASALT. Index once
per assembly; map every sample that should contribute coverage.
Use the same cleaned PE reads that fed assembly (or the declared coverage
set from study-design). Mixing host-contaminated and dehosted libraries
silently biases bins.
Presets trade sensitivity vs speed: --sensitive / --very-sensitive for
catalogue work; --very-sensitive-local allows soft-clipping useful on
fragmented metagenome contigs. Do not treat higher sensitivity as “more true
species”.
Prerequisites
| Requirement | Minimum | Recommended |
|---|
| Contigs | FASTA from assembly | Filtered min-length if huge |
| Reads | Clean PE FASTQ | From kneaddata |
| Tools | bowtie2 + samtools | Same env |
| Threads | 8 | 16–32 |
| Disk | BAM ≈ 1–2× read set | Keep sorted BAM + .bai |
How to run
conda install -c bioconda bowtie2 samtools
mkdir -p temp/map
bowtie2-build --threads 8 contigs.fa temp/map/contigs
bowtie2 -x temp/map/contigs \
-1 temp/hr/${id}_1.fastq -2 temp/hr/${id}_2.fastq \
-p 16 --very-sensitive-local \
2> temp/map/${id}.bowtie2.log \
| samtools sort -@ 8 -o temp/map/${id}.bam
samtools index temp/map/${id}.bam
jgi_summarize_bam_contig_depths --outputDepth temp/map/depth.txt temp/map/*.bam
Key parameters
| Option | Guidance |
|---|
-x | Index prefix from bowtie2-build |
-1/-2 | PE mates; keep order from KneadData --reorder |
-U | SE only when truly single-end |
-p | Threads (aligner); samtools sort uses -@ |
--very-sensitive-local | Soft-clip friendly on short contigs |
--no-unal | Smaller SAM/BAM; lose unmapped diagnostics |
Output & QC
| Artifact | Use |
|---|
${id}.bam + .bai | CoverM, binners, inStrain |
depth.txt | MetaBAT2 -a |
| stderr overall alignment rate | Library/assembly sanity |
samtools flagstat temp/map/${id}.bam
Decision tree
Illumina → contig coverage?
├─ Large cohort / speed critical → strobealign (or bwa-mem2)
├─ Lab standard / KneadData ecosystem → bowtie2
└─ Long reads → minimap2 (not bowtie2)
Troubleshooting
| Symptom | Action |
|---|
| ~0% mapped | Contigs/reads mismatch; wrong sample IDs |
| PE sync errors | Re-KneadData with --reorder |
bowtie2-build RAM blowup | Split huge contig sets or raise node RAM |
| Unsorted BAM breaks MetaBAT | Always samtools sort + index |
Related skills
samtools · strobealign · bwa-mem2 · coverm · metabat2 · basalt · study-design
