Pangenome graph from assemblies

Built by aligning high-quality genomes, saved as paths through the pangenome.

Human Pangenome Reference Consortium (HPRC)

Annotations could be paths through the pangenome

GAM format

• Reads aligned to a pangenome graph.
• Used by vg, GraphAligner, …
• Binary/protobuf format, indexable but unwieldy.

{
    "mapping_quality": 60,
    "name": ">44878920>44878957",
    "path": {
        "mapping": [
            {"edit": [{"from_length": 1, "to_length": 1}],
             "position": {"node_id": "44878920"}, "rank": "1"},
            {"edit": [{"from_length": 1, "to_length": 1}],
             "position": {"node_id": "44878957"}, "rank": "2"}
        ]
    },
    "score": 2,
    "sequence": "AC"
}

Graph Alignment Format (GAF)

• Text format describes mapping between sequences and a graph.

read_name_6 100 0   100 +   <394<393<392<391    128 25  124 100 100 60 cs:Z::100

!-----query info----------!-----target info----------------!--------------------!
  name size range strand         path size range              align + opt. tags

Indexing genomic files with HTSlib

https://github.com/samtools/htslib

Indexing using minimum/maximum node IDs

Assumes node IDs are sorted integers.

read_name_9 100 0   100 +   <3222<3221<3220<3219    128 18  117 100 100 0

New commands available

Sorting a GAF file

vg gamsort -G reads.gaf.gz | bgzip > reads.sorted.gaf.gz

Indexing sorted bgzipped GAF

tabix -p gaf reads.sorted.gaf.gz

Extracting a node interval

tabix reads.sorted.gaf.gz {node}:20034-20158

Sorting short sequencing reads

Illumina 30x short reads for HG002, aligned to the HPRC pangenome with vg giraffe.

Projecting annotation to the pangenome

Input annotation relative to one haplotype (BED or GFF), present in the pangenome (path)

vg annotate -x pg.gbz -f genes.hapH.gff -F > genes.hapH.gaf

Visualization with the sequenteTubemap

Interactively query a subgraph and aligned reads.

Recently modified to accept indexed GAF files.

https://github.com/vgteam/sequenceTubemap

Coding sequence (CDS) for two haplotypes in the HPRC pangenome.

Visualization with Bandage?

BandageNG can color nodes by paths, present in the input graph (GFA).

https://github.com/asl/BandageNG

Mobile element insertion

Projecting variants to the pangenome

Custom scripts to (try to) find and annotate known variants in the pangenome.

Annotating the GWAS catalog and eQTLs

Annotating genotype calls

E.g. to investigate supporting reads.

Summarizing read coverage

E.g. for epigenomics tracks.

On the linear genome:

Summarizing read coverage

• Re-analyzing ENCODE ATAC-seq data across a few tissues.
• Implemented a naive approach to bin read coverage.
  1. Split the coverage in bins, e.g. (0, 5, 30, +)
  2. Extend them to create paths of contiguous bins.

Coverage of ATAC-seq data from ENCODE

Reads from 7 cell types aligned with vg giraffe to the HPRC pangenome.

Conclusion

GAF files can now be sorted/indexed, and then queried fast.

Acknowledgments

Adam M. Novak, Dickson Chung, Glenn Hickey, Sarah Djebali, Toshiyuki T. Yokoyama, Erik Garrison, Benedict Paten.

Compact sorted node IDs through graph sorting

Graph sorting aims to find the best node order for a 1D and 2D layout.