Agent skill

bio-read-alignment-bwa-alignment

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Install this agent skill to your Project

npx add-skill https://github.com/FreedomIntelligence/OpenClaw-Medical-Skills/tree/main/skills/bio-read-alignment-bwa-alignment

SKILL.md

name: bio-read-alignment-bwa-alignment description: Align DNA short reads to reference genomes using bwa-mem2, the faster successor to BWA-MEM. Use when aligning DNA short reads to a reference genome. tool_type: cli primary_tool: bwa-mem2 measurable_outcome: Execute skill workflow successfully with valid output within 15 minutes. allowed-tools:

  • read_file
  • run_shell_command

BWA-MEM2 Alignment

Build Index

bash
# Index reference genome (required once)
bwa-mem2 index reference.fa

# Creates: reference.fa.0123, reference.fa.amb, reference.fa.ann, reference.fa.bwt.2bit.64, reference.fa.pac

Basic Alignment

bash
# Paired-end reads
bwa-mem2 mem -t 8 reference.fa reads_1.fq.gz reads_2.fq.gz > aligned.sam

# Single-end reads
bwa-mem2 mem -t 8 reference.fa reads.fq.gz > aligned.sam

Alignment with Read Groups

bash
# Add read group information (required for GATK)
bwa-mem2 mem -t 8 \
    -R '@RG\tID:sample1\tSM:sample1\tPL:ILLUMINA\tLB:lib1' \
    reference.fa reads_1.fq.gz reads_2.fq.gz > aligned.sam

Direct to Sorted BAM

bash
# Pipe to samtools for sorted BAM output
bwa-mem2 mem -t 8 \
    -R '@RG\tID:sample1\tSM:sample1\tPL:ILLUMINA' \
    reference.fa reads_1.fq.gz reads_2.fq.gz | \
    samtools sort -@ 4 -o aligned.sorted.bam -

# Index the BAM
samtools index aligned.sorted.bam

Mark Duplicates Pipeline

bash
# Full pipeline: align, fixmate, sort, markdup
bwa-mem2 mem -t 8 -R '@RG\tID:sample1\tSM:sample1\tPL:ILLUMINA' \
    reference.fa reads_1.fq.gz reads_2.fq.gz | \
    samtools fixmate -m -@ 4 - - | \
    samtools sort -@ 4 - | \
    samtools markdup -@ 4 - aligned.markdup.bam

samtools index aligned.markdup.bam

Common Options

bash
bwa-mem2 mem -t 8 \         # Threads
    -M \                     # Mark shorter split hits as secondary (Picard compatible)
    -Y \                     # Use soft clipping for supplementary alignments
    -K 100000000 \           # Process INT input bases in each batch
    -R '@RG\tID:s1\tSM:s1' \ # Read group
    reference.fa r1.fq r2.fq

Key Parameters

Parameter Default Description
-t 1 Number of threads
-k 19 Minimum seed length
-w 100 Band width for extension
-r 1.5 Re-seeding trigger ratio
-c 500 Skip seeds with more than INT hits
-A 1 Match score
-B 4 Mismatch penalty
-O 6 Gap open penalty
-E 1 Gap extension penalty
-M off Mark secondary alignments

Output Filters

bash
# Filter unmapped and low quality
bwa-mem2 mem -t 8 reference.fa r1.fq r2.fq | \
    samtools view -@ 4 -bS -q 20 -F 4 - | \
    samtools sort -@ 4 -o aligned.filtered.bam -

Split Read Alignment

bash
# For SV detection, use -Y for soft clipping
bwa-mem2 mem -t 8 -Y reference.fa r1.fq r2.fq > aligned.sam

Memory Requirements

  • Index loading: ~10GB for human genome
  • Per thread: ~1-2GB
  • Typical human WGS: 30-50GB RAM with 8 threads

BWA-MEM (Alternative)

bash
# Build index
bwa index reference.fa

# Paired-end alignment
bwa mem -t 8 reference.fa reads_1.fq.gz reads_2.fq.gz > aligned.sam

# With read groups
bwa mem -t 8 -R '@RG\tID:sample1\tSM:sample1\tPL:ILLUMINA' \
    reference.fa reads_1.fq.gz reads_2.fq.gz > aligned.sam

# Direct to sorted BAM
bwa mem -t 8 -R '@RG\tID:sample1\tSM:sample1\tPL:ILLUMINA' \
    reference.fa reads_1.fq.gz reads_2.fq.gz | \
    samtools sort -@ 4 -o aligned.sorted.bam -

BWA-MEM vs BWA-MEM2

Feature BWA-MEM BWA-MEM2
Status Active Archived
Speed 1x 2-3x faster
Index format .bwt .bwt.2bit.64
Results Baseline Nearly identical
Memory ~5GB ~10GB

Related Skills

  • read-qc/fastp-workflow - Preprocess reads before alignment
  • alignment-files/alignment-sorting - Post-alignment processing
  • alignment-files/duplicate-handling - Mark duplicates
  • variant-calling/variant-calling - Call variants from BAM

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