Range Downloads

Extract specific genomic regions from BAM and VCF files using coordinates or BED files with automatic tool integration.

Overview

Range downloads allow you to extract specific genomic regions instead of downloading entire files, providing:

Focused data retrieval - Select regions of interest
Reduced data transfer - Download only what you need
Storage efficiency - Smaller files for specific analyses
Faster processing - Work with relevant data quickly

Prerequisites

Range downloads require external bioinformatics tools:

Tool	Version	Purpose
samtools	v1.17+	BAM file processing and region extraction
tabix	v1.20+	VCF file indexing and querying
bgzip	v1.20+	VCF file compression

Tool Installation

Ubuntu/Debian:

bash

sudo apt update
sudo apt install samtools tabix

macOS (Homebrew):

bash

brew install samtools htslib

Conda/Mamba:

bash

conda install -c bioconda samtools tabix

Verification

Check tool availability and versions:

bash

samtools --version | head -1
tabix --version 2>&1 | head -1
bgzip --version 2>&1 | head -1

The tool automatically verifies these dependencies when using range features.

Basic Range Syntax

Genomic Coordinates

Use standard genomic coordinate format: chromosome:start-end

bash

# Single genomic region
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000"

# Multiple regions (space-separated)
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000 chr2:500000-1500000"

# Another region
./varvis-download.cjs -t mytarget -a 12345 -g "chr7:5500000-5600000"

BED File Regions

Use BED files for complex region definitions:

Create regions.bed:

bed

chr1    1000000      2000000      region_a
chr2    500000       1500000      region_b
chr7    5500000      5600000      region_c
chrX    1000000      1100000      region_x

Use BED file:

bash

./varvis-download.cjs -t mytarget -a 12345 -b regions.bed

File Type Behavior

BAM Files

Range extraction for BAM:

Uses samtools view with region specification
Requires BAM index (.bai) file
Automatically downloads index if not present
Creates new index for extracted BAM

bash

# BAM range download
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -f "bam,bam.bai"

Output files:

sample_001.chr1_1000000_2000000.bam      # Extracted region
sample_001.chr1_1000000_2000000.bam.bai  # New index

VCF Files

Range extraction for VCF:

Uses tabix for region querying
Requires VCF index (.tbi) file
Automatically compresses output with bgzip
Creates new index for extracted VCF

bash

# VCF range download
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -f "vcf.gz,vcf.gz.tbi"

Output files:

sample_001.chr1_1000000_2000000.vcf.gz     # Extracted and compressed
sample_001.chr1_1000000_2000000.vcf.gz.tbi # New index

Advanced Range Examples

Region-Specific Downloads

Multiple defined regions:

bash

# Region A
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000"

# Region B
./varvis-download.cjs -t mytarget -a 12345 -g "chr2:500000-1500000"

# Both regions
./varvis-download.cjs -t mytarget -a 12345 \
  -g "chr1:1000000-2000000 chr2:500000-1500000"

Research region set:

bash

# Create research_regions.bed
cat > research_regions.bed << EOF
chr1    1000000     2000000     region_a
chr2    500000      1500000     region_b
chr7    5500000     5600000     region_c
chr12   25000000    25100000    region_d
chr20   1000000     1100000     region_e
EOF

# Download research regions
./varvis-download.cjs -t mytarget -a 12345 -b research_regions.bed

Unmapped Read Extraction

The --unmapped flag extracts reads with no reference assignment from BAM files using the samtools wildcard chromosome *. This is useful for identifying contamination, adapter sequences, or novel sequences (e.g., in Illumina NovaSeq data).

Unmapped Only

bash

# Extract only unmapped reads (creates sample.unmapped.bam)
./varvis-download.cjs -t mytarget -a 12345 --unmapped

Only samtools is required (no tabix/bgzip). VCF files are automatically skipped.

Combined with Range Downloads

When used with --range, both ranged and unmapped reads are included in a single BAM file:

bash

# Single BAM with a genomic region + unmapped reads
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" --unmapped

This uses command-line regions instead of a BED file to allow the * wildcard alongside genomic coordinates.

Exome Regions

Target-enriched data retrieval:

bash

# Download exome target regions
./varvis-download.cjs -t mytarget -a 12345 -b exome_targets.bed -f "bam,bam.bai"

Custom region panel:

bash

# Create custom region BED file
cat > custom_regions.bed << EOF
chr1    1000000     2000000     region_a
chr2    500000      1500000     region_b
chr3    2500000     2600000     region_c
chr7    5500000     5600000     region_d
chr11   1000000     1100000     region_e
EOF

./varvis-download.cjs -t mytarget -a 12345 -b custom_regions.bed

Whole Chromosome Downloads

Single chromosome:

bash

# Chromosome 21 (smallest autosome)
./varvis-download.cjs -t mytarget -a 12345 -g "chr21:1-48129895"

# X chromosome
./varvis-download.cjs -t mytarget -a 12345 -g "chrX:1-156040895"

Multiple chromosomes:

bash

# Chromosomes 21 and 22
./varvis-download.cjs -t mytarget -a 12345 \
  -g "chr21:1-48129895 chr22:1-50818468"

Range Download Workflow

Complete Workflow Example

bash

#!/bin/bash
# range_download_workflow.sh

ANALYSIS_ID="12345"
REGION_LABEL="region_a"
REGION="chr1:1000000-2000000"
OUTPUT_DIR="./range_${ANALYSIS_ID}_${REGION_LABEL}"

echo "Starting range download workflow for $REGION_LABEL..."

# Create output directory
mkdir -p "$OUTPUT_DIR"

# Download BAM region
echo "Downloading BAM region: $REGION"
./varvis-download.cjs -t mytarget -a "$ANALYSIS_ID" \
  -g "$REGION" \
  -f "bam,bam.bai" \
  -d "$OUTPUT_DIR"

# Download VCF region
echo "Downloading VCF region: $REGION"
./varvis-download.cjs -t mytarget -a "$ANALYSIS_ID" \
  -g "$REGION" \
  -f "vcf.gz,vcf.gz.tbi" \
  -d "$OUTPUT_DIR"

# Verify downloads
echo "Verifying downloaded files..."
for FILE in "$OUTPUT_DIR"/*.bam; do
    if samtools view -H "$FILE" >/dev/null 2>&1; then
        echo "✓ Valid BAM: $(basename "$FILE")"
    else
        echo "✗ Invalid BAM: $(basename "$FILE")"
    fi
done

for FILE in "$OUTPUT_DIR"/*.vcf.gz; do
    if tabix -l "$FILE" >/dev/null 2>&1; then
        echo "✓ Valid VCF: $(basename "$FILE")"
    else
        echo "✗ Invalid VCF: $(basename "$FILE")"
    fi
done

echo "Range download workflow complete: $OUTPUT_DIR"

Quality Control Integration

Post-download QC:

bash

#!/bin/bash
# range_qc.sh

REGION_BAM="sample_001.chr1_1000000_2000000.bam"
REGION_VCF="sample_001.chr1_1000000_2000000.vcf.gz"

# BAM QC
echo "BAM Quality Control:"
samtools flagstat "$REGION_BAM"
samtools view "$REGION_BAM" | wc -l | xargs echo "Total reads:"

# VCF QC
echo "VCF Quality Control:"
zcat "$REGION_VCF" | grep -v "^#" | wc -l | xargs echo "Total variants:"
zcat "$REGION_VCF" | grep -v "^#" | cut -f7 | sort | uniq -c | sort -nr

Performance Considerations

File Size Impact

Typical size reductions:

Single small region: 99% size reduction
Exome regions: 98% size reduction
Chromosome: 96% size reduction
Multiple regions: 95-99% reduction

Size estimation:

bash

# Check original file sizes
./varvis-download.cjs -t mytarget -a 12345 --list | grep -E "bam|vcf"

# Estimate region size (rough calculation)
# Region size / Genome size * Original file size
# Example: 10kb region / 3.2Gb genome * 2GB file = ~6KB

Network Optimization

Range downloads are faster because:

Smaller data transfer
Less network time
Reduced bandwidth usage
Parallel processing possible

Optimize for multiple regions:

bash

# Download regions in parallel (separate processes)
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" &
./varvis-download.cjs -t mytarget -a 12345 -g "chr2:500000-1500000" &
./varvis-download.cjs -t mytarget -a 12345 -g "chr7:5500000-5600000" &
wait  # Wait for all downloads to complete

Storage Optimization

Efficient storage patterns:

bash

# Organize by region
mkdir -p ./regions/{region_a,region_b,region_c}

# Download to specific directories
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -d "./regions/region_a/"
./varvis-download.cjs -t mytarget -a 12345 -g "chr2:500000-1500000" -d "./regions/region_b/"
./varvis-download.cjs -t mytarget -a 12345 -g "chr7:5500000-5600000" -d "./regions/region_c/"

Integration with Downstream Research Tools

Optional Downstream Processing

Region-specific processing:

bash

# Download region
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -f "bam,bam.bai"

# Run a downstream research workflow on the region
REGION_BAM="sample_001.chr1_1000000_2000000.bam"
REFERENCE="/data/reference/hg38.fa"
OUTPUT_VCF="research_output.chr1_1000000_2000000.vcf"

# Example with a user-managed external tool
gatk HaplotypeCaller \
  -R "$REFERENCE" \
  -I "$REGION_BAM" \
  -O "$OUTPUT_VCF" \
  -L "chr1:1000000-2000000"

Coverage Analysis

Regional coverage analysis:

bash

# Download BAM region
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -f "bam,bam.bai"

# Calculate coverage for region
REGION_BAM="sample_001.chr1_1000000_2000000.bam"

# Per-base coverage
samtools depth "$REGION_BAM" > coverage.chr1_1000000_2000000.txt

# Coverage statistics
samtools depth "$REGION_BAM" | awk '{sum+=$3; count++} END {print "Average coverage:", sum/count}'

# Coverage histogram
samtools depth "$REGION_BAM" | cut -f3 | sort -n | uniq -c | sort -nr > coverage_histogram.txt

Research Annotation Workflows

Region-specific research annotation:

bash

# Download VCF region
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" -f "vcf.gz,vcf.gz.tbi"

# Annotate variants in region
REGION_VCF="sample_001.chr1_1000000_2000000.vcf.gz"

# VEP annotation
vep --input_file "$REGION_VCF" \
    --output_file "annotated.chr1_1000000_2000000.vcf" \
    --format vcf \
    --vcf \
    --symbol \
    --terms SO \
    --tsl \
    --hgvs \
    --fasta /data/reference/hg38.fa \
    --offline \
    --cache

Troubleshooting Range Downloads

Common Issues

Tool not found errors:

Error: samtools not found or version too old
Required: samtools v1.17+, Found: v1.10

Solution:

bash

# Update samtools
conda install -c bioconda samtools=1.17
# or
sudo apt install samtools=1.17

Index file issues:

Error: Could not load index for sample_001.bam

Solution:

bash

# The tool should automatically download indexes
# If manual intervention needed:
samtools index sample_001.bam

Region format errors:

Error: Invalid region format: chr1:1000000_2000000

Solution:

bash

# Use colon and dash: chr1:1000000-2000000
# Not underscore: chr1:1000000_2000000

Debug Range Downloads

Enable debug logging:

bash

./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000000-2000000" --loglevel debug

Test region validity:

bash

# Test with a small region first
./varvis-download.cjs -t mytarget -a 12345 -g "chr1:1000-2000" --list

# Verify chromosome naming
./varvis-download.cjs -t mytarget -a 12345 --list | grep -i bam
samtools view -H original.bam | grep "@SQ"  # Check chromosome names

Manual tool testing:

bash

# Test samtools with region
samtools view original.bam "chr1:1000000-2000000" | head

# Test tabix with region
tabix original.vcf.gz "chr1:1000000-2000000" | head

Best Practices

Region Selection

Use specific regions for focused retrieval
Combine related regions in single downloads
Document the coordinate source and reference genome build
Use standard coordinate systems (0-based or 1-based consistently)

File Management

Organize by region and analysis identifier
Use descriptive directory names
Keep original and extracted files separate
Document region coordinates and purposes

Quality Control

Always verify extracted files
Check read/variant counts in regions
Validate chromosome naming consistency
Test with small regions before large downloads

Next Steps

Archive Management - Handle archived files
Batch Operations - Large-scale processing
Examples - Real-world download scenarios

Range Downloads ​

Overview ​

Prerequisites ​

Tool Installation ​

Verification ​

Basic Range Syntax ​

Genomic Coordinates ​

BED File Regions ​

File Type Behavior ​

BAM Files ​

VCF Files ​

Advanced Range Examples ​

Region-Specific Downloads ​

Unmapped Read Extraction ​

Unmapped Only ​

Combined with Range Downloads ​

Exome Regions ​

Whole Chromosome Downloads ​

Range Download Workflow ​

Complete Workflow Example ​

Quality Control Integration ​

Performance Considerations ​

File Size Impact ​

Network Optimization ​

Storage Optimization ​

Integration with Downstream Research Tools ​

Optional Downstream Processing ​

Coverage Analysis ​

Research Annotation Workflows ​

Troubleshooting Range Downloads ​

Common Issues ​

Debug Range Downloads ​

Best Practices ​

Region Selection ​

File Management ​

Quality Control ​

Next Steps ​

Range Downloads

Overview

Prerequisites

Tool Installation

Verification

Basic Range Syntax

Genomic Coordinates

BED File Regions

File Type Behavior

BAM Files

VCF Files

Advanced Range Examples

Region-Specific Downloads

Unmapped Read Extraction

Unmapped Only

Combined with Range Downloads

Exome Regions

Whole Chromosome Downloads

Range Download Workflow

Complete Workflow Example

Quality Control Integration

Performance Considerations

File Size Impact

Network Optimization

Storage Optimization

Integration with Downstream Research Tools

Optional Downstream Processing

Coverage Analysis

Research Annotation Workflows

Troubleshooting Range Downloads

Common Issues

Debug Range Downloads

Best Practices

Region Selection

File Management

Quality Control

Next Steps