Liftover Coordinates

Convert Genomic Coordinates Between Assembly Versions

When to Use

• User needs to convert genomic coordinates between assemblies (hg19↔hg38, mm9↔mm10)
• User asks about "liftover", "coordinate conversion", "assembly mismatch", or "CrossMap"
• User has data in hg19/GRCh37 that needs conversion to GRCh38 (or vice versa) before integration
• User wants to use UCSC liftOver or CrossMap for BED, VCF, bigWig, or BAM files
• Example queries: "convert my hg19 peaks to hg38", "liftover coordinates for integration with ENCODE", "my data is in mm9, how do I convert to mm10?"

Guide coordinate liftover between genome assemblies using UCSC liftOver, CrossMap, Ensembl REST API, and rtracklayer. Assembly conversion is one of the most common pitfalls in genomics — this skill provides the definitive workflow for safe, reproducible liftover with full provenance tracking.

Scientific Rationale

The question: "How do I safely convert my genomic coordinates from one assembly to another without losing data or introducing errors?"

Assembly conversion is referenced as a critical step in 10+ other ENCODE Toolkit skills because ENCODE spans multiple data releases: some experiments were processed against hg19/GRCh37, while most current data uses GRCh38/hg38. Combining data across assemblies without proper liftover is one of the most common and most dangerous errors in computational genomics — coordinates that look valid in both assemblies may refer to completely different genomic locations.

The Core Problem

Genome assemblies are updated to fix errors, fill gaps, add alternative haplotypes, and improve centromeric/telomeric sequence. Between hg19 and hg38, approximately 1,000 sequence gaps were closed, 8% of the genome was modified, and several regions were rearranged. A coordinate like chr17:41,197,694 in hg19 (BRCA1) maps to chr17:43,044,295 in GRCh38 — a shift of nearly 2 Mb. Using the wrong assembly silently produces incorrect results.

When to Liftover

Common scenarios requiring coordinate conversion:

• Combining ENCODE data from different releases: Some hg19, some GRCh38 — must unify before intersection
• Integrating GWAS Catalog results: Many GWAS hits are still reported in hg19/GRCh37 coordinates
• Using gnomAD: gnomAD v4 uses GRCh38; older v2 datasets use GRCh37
• Cross-species comparison: Mouse data across mm9/mm10/GRCm39
• Legacy datasets: Published supplementary files often use older assemblies
• ClinVar integration: Some ClinVar entries reference GRCh37 positions
• GTEx cross-reference: GTEx v8 uses GRCh38, earlier versions used GRCh37

Literature Support

• Kent et al. 2002 (Genome Research, ~5,000 citations): UCSC Genome Browser and the liftOver tool. The original chain/net alignment framework for coordinate conversion between genome assemblies. DOI
• Zhao et al. 2014 (Bioinformatics, ~800 citations): CrossMap — a versatile tool for coordinate conversion between genome assemblies. Handles VCF, BAM, bigWig, GFF, and Wiggle formats that UCSC liftOver cannot process natively. DOI
• Hinrichs et al. 2006 (Nucleic Acids Research, ~1,200 citations): UCSC genome browser chain/net alignment methodology. Defines the reciprocal-best chain alignment that underpins coordinate conversion. DOI
• Kuhn et al. 2013 (Nucleic Acids Research, ~600 citations): Assembly updates and the implications for re-annotation. Documents the biological impact of assembly changes on gene models and regulatory element coordinates. DOI
• Schneider et al. 2017 (Genome Research, ~400 citations): GRCh38 improvements over GRCh37 — gap closures, centromere models, alternative haplotypes. Quantifies what changed and why liftover is necessary. DOI
• Amemiya et al. 2019 (Scientific Reports, ~1,372 citations): ENCODE Blacklist regions — some blacklisted regions are assembly-specific. Liftover of blacklist files must use the correct version. DOI

Assembly Version Mapping

Naming Convention Alert

The same assembly has different names depending on the source:

• UCSC convention: hg19, hg38, mm10 — used in filenames, chromosome prefixes (chr1)
• NCBI/GRC convention: GRCh37, GRCh38, GRCm38 — used in publications, Ensembl
• Ensembl convention: Chromosomes without chr prefix (1 instead of chr1)

Always verify which naming convention your data uses. Mixing chr1 (UCSC) with 1 (Ensembl) causes silent failures in bedtools intersection and peak overlap analysis.

Chain Files

Chain files encode the alignment between assemblies and are the essential input for liftover.

Source: UCSC (Recommended)

https://hgdownload.soe.ucsc.edu/goldenPath/{from}/liftOver/{from}To{To}.over.chain.gz

Common chain files:

Source: Ensembl

ftp://ftp.ensembl.org/pub/assembly_mapping/

Ensembl provides chain files for their coordinate system (without chr prefix). Useful when working with Ensembl VEP output or Ensembl gene annotations.

Source: NCBI Remap

NCBI Genome Remapping Service: https://www.ncbi.nlm.nih.gov/genome/tools/remap

• Web-based and API access
• Handles complex remapping with alignment-based and annotation-based methods
• Useful for non-standard assemblies or patch-level conversions

Chain File Verification

Always verify chain file integrity after download:

wget https://hgdownload.soe.ucsc.edu/goldenPath/hg19/liftOver/hg19ToHg38.over.chain.gz
md5sum hg19ToHg38.over.chain.gz
# Verify against UCSC md5sum.txt in the same directory
gunzip -t hg19ToHg38.over.chain.gz  # Test archive integrity

Tool Guide: UCSC liftOver (BED Files)

The standard tool for BED-format coordinate conversion.

Basic Usage

liftOver input.bed hg19ToHg38.over.chain.gz output.bed unmapped.bed

Parameters

Recommended Settings by Data Type

Handling narrowPeak Format

UCSC liftOver expects standard BED (3–12 columns). For narrowPeak files (BED6+4):

# Step 1: Extract BED6 columns + preserve extra columns as name
awk 'BEGIN{OFS="\t"} {print $1, $2, $3, $4, $5, $6, $7, $8, $9, $10}' input.narrowPeak > input_full.bed

# Step 2: Liftover (liftOver handles extra columns)
liftOver input_full.bed hg19ToHg38.over.chain.gz output.bed unmapped.bed

# Step 3: Verify column count is preserved
awk '{print NF}' output.bed | sort -u

Peak summit recalculation: After liftover, the summit position (column 10 in narrowPeak = offset from start) may no longer accurately represent the signal maximum. For critical analyses, re-calculate summits from signal data in the new assembly rather than relying on lifted summit positions.

Checking Unmapped Regions

# Count unmapped regions
wc -l unmapped.bed  # Note: comment lines start with #

# Calculate loss rate
total=$(wc -l < input.bed)
unmapped=$(grep -v '^#' unmapped.bed | wc -l)
loss_pct=$(echo "scale=2; $unmapped * 100 / $total" | bc)
echo "Lost $unmapped of $total regions ($loss_pct%)"

# Investigate reasons for unmapping
grep '^#' unmapped.bed | sort | uniq -c | sort -rn
# Common reasons:
# "Partially deleted in new" — region spans a deletion
# "Deleted in new" — region fully removed
# "Split in new" — region maps to multiple locations

Tool Guide: CrossMap (VCF, bigWig, BAM, GFF)

CrossMap (Zhao et al. 2014) handles file formats that UCSC liftOver cannot process natively.

VCF Conversion

CrossMap vcf hg19ToHg38.over.chain.gz input.vcf hg38.fa output.vcf

Critical VCF considerations:

• CrossMap updates coordinates AND checks REF alleles against the new reference
• Variants where the REF allele changes between assemblies are flagged
• Always re-validate variant calls after liftover using bcftools norm
• Multi-allelic variants may need special handling

# Post-liftover VCF validation
bcftools norm -f hg38.fa -c ws output.vcf -o output.normalized.vcf 2> norm_warnings.log
# -c ws: warn about and set incorrect REF alleles

bigWig Conversion

CrossMap bigwig hg19ToHg38.over.chain.gz input.bw output.bw

Signal track caveats:

• Resolution is reduced during conversion (interpolation at boundaries)
• Regions that split during liftover lose signal accuracy
• For quantitative analysis, re-generate signal tracks from re-aligned reads when possible

BAM Conversion

CrossMap bam hg19ToHg38.over.chain.gz input.bam output.bam

BAM liftover is generally NOT recommended:

• Read mapping quality is meaningless after coordinate shifting
• Paired-end relationships may break
• Duplicate marking becomes invalid
• Best practice: Re-align from FASTQ to the new reference genome

GFF/GTF Conversion

CrossMap gff hg19ToHg38.over.chain.gz input.gff output.gff

Useful for lifting gene annotations, but prefer downloading the native annotation for the target assembly from GENCODE or Ensembl.

Tool Guide: Ensembl REST API (Single Coordinates)

For programmatic conversion of individual coordinates without installing local tools.

API Endpoint

GET https://rest.ensembl.org/map/human/GRCh37/{region}/GRCh38?content-type=application/json

Example

import requests

def liftover_ensembl(chrom, start, end, source="GRCh37", target="GRCh38", species="human"):
    """Convert coordinates using Ensembl REST API."""
    region = f"{chrom}:{start}..{end}:1"
    url = f"https://rest.ensembl.org/map/{species}/{source}/{region}/{target}"
    headers = {"Content-Type": "application/json"}
    response = requests.get(url, headers=headers)
    if response.status_code == 200:
        mappings = response.json()["mappings"]
        return mappings
    return None

# Example: BRCA1 region
mappings = liftover_ensembl("17", 41197694, 41276113)
for m in mappings:
    mapped = m["mapped"]
    print(f"  {mapped['seq_region_name']}:{mapped['start']}-{mapped['end']}")

Rate Limits

• 15 requests per second (without registered email)
• 50 requests per second (with registered email in User-Agent header)
• For batch conversion, use local liftOver or CrossMap instead

Ensembl Chromosome Naming

Ensembl uses chromosomes WITHOUT chr prefix:

• Ensembl: 17:41197694-41276113
• UCSC: chr17:41197694-41276113

Convert between conventions:

# Add 'chr' prefix (Ensembl to UCSC)
sed 's/^/chr/' input.bed > input_ucsc.bed

# Remove 'chr' prefix (UCSC to Ensembl)
sed 's/^chr//' input.bed > input_ensembl.bed

Tool Guide: R (rtracklayer)

For R-based workflows, rtracklayer provides native liftover support.

library(rtracklayer)
library(GenomicRanges)

# Import chain file
chain <- import.chain("hg19ToHg38.over.chain")

# Create GRanges object from your coordinates
gr <- GRanges(
    seqnames = c("chr17", "chr7", "chr1"),
    ranges = IRanges(
        start = c(41197694, 55086725, 11873),
        end = c(41276113, 55275031, 14409)
    ),
    name = c("BRCA1", "EGFR", "DDX11L1")
)

# Perform liftover
lifted <- liftOver(gr, chain)

# liftOver returns a GRangesList (1:many mapping possible)
# Convert to GRanges (keeping only 1:1 mappings)
lifted_1to1 <- unlist(lifted[elementNROWS(lifted) == 1])

# Check for unmapped
n_unmapped <- sum(elementNROWS(lifted) == 0)
n_multimapped <- sum(elementNROWS(lifted) > 1)
cat(sprintf("Mapped: %d, Unmapped: %d, Multi-mapped: %d\n",
    length(lifted_1to1), n_unmapped, n_multimapped))

Bioconductor Packages for Liftover

Expected Loss Rates

When Loss Rates Are Concerning

• <2% loss: Normal, proceed with analysis
• 2–5% loss: Acceptable for most analyses, document in methods
• 5–10% loss: Investigate — may indicate problematic input regions (many centromeric/repeat-rich regions)
• >10% loss: Something is wrong — check assembly mismatch, chromosome naming, or chain file version

Pitfalls & Edge Cases

• Unmapped regions are expected: 1-5% of coordinates typically fail to lift over. Regions near centromeres, telomeres, and assembly gaps are most affected. Always check the unmapped file and report the loss rate.
• Many-to-one mapping: Some hg19 regions map to multiple hg38 locations due to assembly improvements. UCSC liftOver reports only one mapping by default — use -multiple flag to detect split mappings.
• Peak coordinates may shift asymmetrically: Peak summits can shift by different amounts than peak boundaries after liftover. Re-center peaks on summits after conversion rather than trusting the lifted boundaries.
• Chain file source matters: Only use chain files from UCSC or Ensembl. Third-party chain files may have different coordinate conventions or incomplete mappings. Verify chain file checksums.
• VCF liftover requires reference allele check: After lifting VCF coordinates, the reference allele may no longer match the new assembly. CrossMap handles this with --refgenome but UCSC liftOver does not — always validate.
• Assembly detection is unreliable from filenames: File names like "peaks.bed" give no assembly hint. Check the actual coordinate ranges against known chromosome sizes. chrM length differs between hg19 (16571) and hg38 (16569).

Provenance Integration

Log every liftover operation with encode_log_derived_file for full reproducibility:

encode_log_derived_file(
    file_path="/path/to/lifted_peaks_hg38.bed",
    source_accessions=["ENCSR...", "ENCFF..."],
    description="Lifted [N] narrowPeak regions from hg19 to GRCh38. [X] unmapped ([Y]% loss). Original: [source description]",
    file_type="lifted_coordinates",
    tool_used="UCSC liftOver v377",
    parameters="minMatch=0.95, chain=hg19ToHg38.over.chain.gz (MD5: abc123...), unmapped=[X]/[N] ([Y]%)"
)

Provenance Checklist

Every liftover log entry should include:

• Source file path and assembly
• Chain file used with MD5 checksum
• Tool name and version (e.g., liftOver v377, CrossMap v0.6.4)
• -minMatch or equivalent parameter
• Total input regions/variants
• Successfully mapped count
• Unmapped count and percentage
• Multi-mapped count (if applicable)
• Output file path and assembly

Workflow Summary

1. Identify assemblies:  Check input assembly → check target assembly
2. Get chain file:       Download from UCSC → verify MD5
3. Select tool:          BED → liftOver | VCF → CrossMap | single → Ensembl API | R → rtracklayer
4. Convert:              Run liftover with appropriate parameters
5. Check loss:           Count unmapped, flag if >5%
6. Validate:             Verify output assembly, check chromosome names
7. Post-process:         Re-center peaks, normalize VCF, re-annotate genes
8. Log provenance:       Record all parameters, tools, loss rates

Walkthrough: Converting ENCODE Peak Coordinates Between Genome Assemblies

Goal: Convert ENCODE BED peak files from hg19 to GRCh38 (or vice versa) for cross-study integration when experiments use different genome builds. Context: Older ENCODE experiments may be aligned to hg19, while newer ones use GRCh38. LiftOver enables coordinate conversion for combined analysis.

Step 1: Identify experiments needing liftover

encode_search_experiments(assay_title="Histone ChIP-seq", organ="liver", target="H3K27ac", organism="Homo sapiens")

Expected output:

{
  "total": 8,
  "results": [
    {"accession": "ENCSR100OLD", "assay_title": "Histone ChIP-seq", "assembly": "hg19"},
    {"accession": "ENCSR200NEW", "assay_title": "Histone ChIP-seq", "assembly": "GRCh38"}
  ]
}

Interpretation: ENCSR100OLD uses hg19 — needs liftover before merging with ENCSR200NEW (GRCh38).

Step 2: Download the hg19 peak file

encode_list_files(accession="ENCSR100OLD", file_format="bed", assembly="hg19")

Step 3: Run UCSC liftOver

liftOver ENCSR100OLD_peaks.bed hg19ToHg38.over.chain.gz peaks_GRCh38.bed unmapped.bed

Step 4: Check conversion results

Count converted vs. unmapped:

• If >95% convert successfully → proceed with analysis
• If >5% unmapped → investigate (regions may be in assembly-specific contigs)

Step 5: Log the conversion provenance

encode_log_derived_file(
  source_accessions=["ENCFF100OLD"],
  derived_file="/data/peaks_GRCh38.bed",
  description="Lifted from hg19 to GRCh38 using UCSC liftOver",
  tool="liftOver (UCSC, chain: hg19ToHg38.over.chain.gz)"
)

Integration with downstream skills

• Lifted peaks feed into → histone-aggregation for cross-assembly union merge
• Conversion provenance logged by → data-provenance
• UCSC chain files accessed via → ucsc-browser REST API
• Lifted coordinates used by → variant-annotation for position-dependent annotation

Code Examples

1. Check file assembly before liftover

encode_get_file_info(accession="ENCFF100OLD")

Expected output:

{
  "accession": "ENCFF100OLD",
  "assembly": "hg19",
  "file_format": "bed narrowPeak"
}

2. Find GRCh38 version of same experiment

encode_list_files(accession="ENCSR100OLD", file_format="bed", assembly="GRCh38")

Expected output:

{
  "files": []
}

Interpretation: No GRCh38 files available — liftover is required.

3. Log liftover provenance

encode_log_derived_file(
  source_accessions=["ENCFF100OLD"],
  derived_file="/data/peaks_GRCh38.bed",
  description="hg19→GRCh38 liftOver",
  tool="UCSC liftOver"
)

Expected output:

{"status": "logged", "derived_file": "/data/peaks_GRCh38.bed", "source_count": 1}

Related Skills

• variant-annotation — Variants often need liftover before annotation with ENCODE data (hg19 GWAS variants to GRCh38)
• gwas-catalog — GWAS Catalog coordinates may be in GRCh37; liftover needed for ENCODE GRCh38 integration
• ensembl-annotation — Ensembl REST API provides coordinate mapping; Ensembl uses non-chr chromosome naming
• ucsc-browser — UCSC provides chain files and the liftOver tool; retrieve assembly-specific tracks
• gnomad-variants — gnomAD v4 uses GRCh38; v2 uses GRCh37; liftover needed for cross-version analysis
• histone-aggregation — Aggregating peaks across samples requires all peaks in the same assembly
• accessibility-aggregation — ATAC-seq/DNase-seq peak union requires assembly-consistent coordinates
• data-provenance — Every liftover operation must be logged with chain file, tool version, and loss rate
• publication-trust — Verify literature claims backing analytical decisions

Presenting Results

When reporting liftover results, always present:

• Input summary: Number of regions/variants, source assembly
• Output summary: Number successfully mapped, target assembly
• Loss report: Unmapped count and percentage, with breakdown by reason if available
• Multi-mapping report: Number of regions mapping to multiple locations and how they were handled
• Assembly confirmation: Explicit statement of output assembly (e.g., "All coordinates are now in GRCh38/hg38")
• Flag if loss >5%: Warn the user and investigate the cause (centromeric regions, assembly-specific sequences, or input errors)
• Chain file version: Which chain file was used and its source

Example output summary:

Liftover: hg19 -> GRCh38
Input:    45,231 narrowPeak regions
Mapped:   44,012 (97.3%)
Unmapped: 1,219 (2.7%) — 847 partially deleted, 312 split, 60 fully deleted
Chain:    hg19ToHg38.over.chain.gz (UCSC, MD5: 7a42e...)
Tool:     UCSC liftOver v377, minMatch=0.95

For the request: "$ARGUMENTS"