Bio Read Alignment Bwa Alignment

biotender-max/bio-read-alignment-bwa-alignment

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.

2k tokens

context cost

the whole folder, loaded on every use

files

ships runnable scripts

copies elsewhere

how many repositories repackaged it

132

stars on the repo

on the repository, not the skill itself

Install

one command, takes just this skill from the repository

npx skills add https://github.com/BioTender-max/awesome-bio-agent-skills --skill bio-read-alignment-bwa-alignment

What comes with it

3 495 bytes besides the instruction

examples/align_bwa.sh

usage-guide.md

The instruction itself

15 sections, as written by the author

Version Compatibility

Reference examples tested with: GATK 4.5+, samtools 1.19+

Before using code patterns, verify installed versions match. If versions differ:

CLI: <tool> --version then <tool> --help to confirm flags

If code throws ImportError, AttributeError, or TypeError, introspect the installed

package and adapt the example to match the actual API rather than retrying.

BWA-MEM2 Alignment

"Align reads with BWA" → Map DNA reads to a reference genome using BWA-MEM2, the standard aligner for whole-genome and exome sequencing.

CLI: bwa-mem2 mem -t 8 ref.fa R1.fq R2.fq | samtools sort -o aligned.bam

Build Index

# 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

# 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

# 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

# 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

Goal: Produce a duplicate-marked, sorted BAM file from raw reads in a single streaming pipeline.

Approach: Pipe BWA-MEM2 output through samtools fixmate (to add mate score tags), coordinate sort, and markdup in a single command chain to avoid intermediate files.

# 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

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

# 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

# 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)

# 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 |

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

How to use it

Copy the folder

Take biotender-max/bio-read-alignment-bwa-alignment from the repository into ~/.claude/skills for personal use, or into .claude/skills inside a project.

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