Telomere-to-Telomere · HiFi + Ultra-Long ONT

Telomere to telomere. Centromeres, duplications, rDNA — resolved.

The first truly complete human genome (T2T-CHM13, published 2022) required the union of two technologies: highly accurate HiFi reads for base-level correctness and ultra-long nanopore reads for the largest repeats. Nucleome Informatics brings this same recipe to your organism — plant, animal, fungal, or human.

Plan Your Project → Learn the Basics
HiFi + ultra-long ONT
combined
hifiasm / Verkko
assembly
QV, BUSCO, Merqury
QC
Any organism
— plant, animal, fungal
About the Service

T2T Genome Sequencing at Nucleome Informatics

Most "complete" genomes are not complete: draft assemblies contain hundreds of gaps, concentrated in centromeres, ribosomal DNA arrays, and segmental duplications — precisely the regions driving genome function, evolution, and disease. Telomere-to-telomere (T2T) assembly closes these gaps by combining:

Assemblies are built with hifiasm (with HiFi + ultra-long ONT integration) or Verkko, the two assemblers that produced the T2T-CHM13 human genome and the current generation of complete genomes.

  • Gapless (or gap-minimal, honestly reported) chromosome-scale assembly
  • Haplotype-resolved (diploid) output where data supports it
  • Organellar genomes (mitochondrial/chloroplast) assembled and circularised where present
  • Comprehensive QC: QV (quality value), BUSCO gene completeness, Merqury k-mer spectra, contig N50, gap assessment
  • Structural annotation support: repeat annotation, gene prediction-ready outputs
  • Full methods documentation for publication

What T2T unlocks

  • Centromeres: the last unexplored frontier of most genomes — essential for chromosome biology and genome evolution studies.
  • Segmental duplications: gene conversions and copy-number evolution visible only in complete assemblies.
  • rDNA arrays and satellite landscapes: complete characterisation of the repeat universe.
  • Structural variant discovery: a complete reference makes downstream resequencing projects dramatically more powerful.
Learning Path

From first principles to expert-level analysis

New here? Start with Foundations. Planning a study or reviewing results? Jump to Practitioner or Expert. Each level builds on the one before it, so you can read this page as a short course or dip in where you need to.

Start here if you are new to this service — six ideas that explain what it is and why it matters.

LEVEL 1

What does "telomere to telomere" mean?

A genome assembly where every chromosome is represented by a single continuous sequence from one end to the other — no gaps, no "N-stretches" standing in for unknown regions.

LEVEL 1

Why do most assemblies have gaps?

Repetitive regions (centromeres, rDNA arrays, segmental duplications) are longer than the reads used to sequence them. Short reads cannot bridge them at all; even standard long reads often cannot.

LEVEL 1

What makes T2T possible now?

Two things: HiFi reads (accurate enough to assemble correctly) and ultra-long nanopore reads (long enough to span the biggest repeats). The T2T-CHM13 human genome proved the recipe in 2022.

LEVEL 1

What organisms can this be done for?

Any organism with sufficient DNA: mammals, fish, insects, plants (including polyploids — see our plant genome page), fungi, and protists. Genome size and repeat content determine the data requirements.

LEVEL 1

How much DNA is needed?

T2T projects are DNA-hungry: typically micrograms of high-molecular-weight DNA for HiFi plus a separate ultra-long nanopore extraction. We provide extraction guidance or accept HMW DNA.

LEVEL 1

What is the deliverable?

A chromosome-scale, gapless (or gap-minimal) assembly with QC metrics, plus all supporting data and methods.

For scientists planning a project or reading a report: how each step works and how it can mislead.

LEVEL 2

Data generation

HiFi libraries from HMW DNA (Revio) plus ultra-long nanopore libraries (PromethION, optimised for read length). Optional Hi-C library for scaffolding, or parental short reads for triad-based phasing.

LEVEL 2

Assembly strategy

hifiasm integrates HiFi and ultra-long reads natively, building a repeat graph that resolves centromeric and ribosomal arrays. Verkko offers an alternative graph-based approach. Assembler choice is made per genome based on repeat content and haploidy.

LEVEL 2

Polishing and validation

HiFi-based polishing where needed; QV estimation from k-mers (Merqury); BUSCO gene completeness; manual inspection of gap-closing evidence at each former gap.

LEVEL 2

Haplotype resolution

hifiasm's built-in phasing produces haplotype-resolved (diploid) assemblies; Hi-C or parental data strengthens chromosome-scale phasing. For polyploids, see our plant genome page for subgenome-aware strategies.

LEVEL 2

Organelle assembly

Mitochondrial (and chloroplast, for plants) genomes are assembled separately and circularised — included in the deliverable.

LEVEL 2

QC standards

T2T-grade assemblies are judged on: QV (≥40 is the common benchmark, i.e. <1 error per 10 kb), BUSCO completeness, Merqury spectra consistency, contig N50, and the number of chromosomes assembled telomere-to-telomere.

For study design: the decisions that determine whether the data answers your question.

LEVEL 3

The centromere problem

Centromeric satellites are the hardest regions. Success depends on ultra-long read yield and repeat structure; some centromeres may remain partially resolved — we report per-chromosome status honestly.

LEVEL 3

Polyploid and highly heterozygous genomes

Allopolyploids need subgenome-aware assembly and phasing; high heterozygosity requires careful haplotype separation. See our plant genome page for the extended strategy.

LEVEL 3

Reference-grade vs T2T

A "reference-grade" assembly (contiguous, accurate, chromosome-scale) is achievable for most organisms; full T2T (every chromosome gapless) depends on repeat architecture. We define the target with you before starting.

LEVEL 3

Downstream value

A complete reference transforms subsequent resequencing: short-read projects against a T2T reference recover structural variants and repeat-region variation invisible against gappy references.

LEVEL 3

Pangenome programmes

T2T assemblies are the foundation of pangenome projects; we support multi-individual T2T pipelines for cohort-scale reference building.

LEVEL 3

Data management

T2T projects generate large datasets (HiFi + UL-ONT + Hi-C). We handle storage, transfer, and NCBI submission-ready formatting (including full methods for the assembly publication).

T2T
telomere-to-telomere; a gapless chromosome-level assembly.
QV
quality value; base-level consensus accuracy (QV 40 ≈ 1 error per 10,000 bases).
Merqury
k-mer-based assembly QC without a reference.
BUSCO
Benchmarking Universal Single-Copy Orthologs; gene completeness metric.
Segmental duplication
duplicated genomic segments (>1 kb, ≥90% identity) that complicate assembly.
Hi-C
chromatin conformation capture; used for scaffolding and phasing.
The Science

How it works, in two figures

The T2T-CHM13 human genome (Nurk et al., Science 2022) established the HiFi + ultra-long ONT + hifiasm paradigm; Verkko (Rautiainen et al., Nature Methods 2023) is the complementary assembler. See references below.

Figure 1 The gap landscape. schematic
Draft assembly T2T assembly Hundreds of gaps at repeats Centromeres and rDNA unresolved Segmental duplications collapsed Gapless chromosomes where achievable Centromeric arrays assembled QV, BUSCO and Merqury validated
Figure 1 | The gap landscape. (Illustrative schematic, not sample data.)
Figure 2 The T2T recipe. schematic
Genome scoping HMW DNA HiFi + ultra-long ONT hifiasm / Verkko assembly Hi-C scaffolding & phasing QV / BUSCO / Merqury QC
Figure 2 | The T2T recipe. (Illustrative schematic, not sample data.)
Technology Highlights

Built for the questions you are asking

The platforms, pipelines and standards behind this service.

PacBio HiFi on Revio

≥99.9% accurate reads at 15–20+ kb — the accuracy backbone of every modern T2T assembly.

Ultra-long Oxford Nanopore on PromethION

Reads of 50–100+ kb (N50 configurable by input DNA quality) span the tandem repeats and arrays that HiFi alone cannot bridge.

hifiasm with UL-ONT integration

The assembler that produced the first T2T human genome, integrating HiFi accuracy with ultra-long read span, with built-in haplotype phasing.

Verkko

The alternative T2T assembler (from the Canu team), with graph-based resolution of complex repeats — we run both where assembly difficulty warrants.

Merqury k-mer QC

Reference-free consensus quality and completeness assessment from k-mer spectra — the standard for T2T-grade validation.

Hi-C scaffolding options

Chromosome-scale scaffolding and haplotype phasing via Hi-C (or parental short reads for triad phasing) where chromosome-length confirmation is required. ---

Scope Note

What this service does and does not cover

Scope note. T2T assembly is the most demanding genome assembly class. Outcomes depend on DNA quality, genome size, repeat architecture, and ploidy. We commit to defined QC targets (QV, BUSCO, contiguity) agreed at project design, and report per-chromosome completion status honestly — including any regions that remain unresolved and why. Full T2T (every chromosome gapless) is the goal but is not guaranteed for every genome; reference-grade assemblies are achievable for the large majority of organisms. ---
Why Nucleome

Service highlights

What you can expect from our team in Hyderabad.

Both technologies under one roof

PacBio Revio HiFi and ONT PromethION ultra-long reads from one laboratory — no cross-vendor coordination.

The proven assembler stack

hifiasm and Verkko, the tools behind the T2T-CHM13 human genome.

Honest QC

QV, BUSCO, Merqury, and per-chromosome gap status — reported, not asserted.

Polyploid-capable

Subgenome-aware strategies for plant genomes (see our plant genome page).

NCBI submission-ready

Formatted assemblies and full methods documentation for publication and database submission.

India's long-read assembly expertise

South Asia's first PacBio Sequel II and Revio service provider. ---

Questions

Frequently asked, in plain language

From the basics to the finer points of design and interpretation.

basicWhat is a T2T assembly?
A genome assembly with every chromosome as one continuous gapless sequence, from telomere to telomere — the standard set by the first complete human genome in 2022.
basicWhy can't short reads do this?
Short reads (150–300 bp) are far shorter than the repeats they need to bridge. Even perfect short-read data cannot resolve a 10 Mb centromeric array.
basicWhat do I need to provide?
High-quality high-molecular-weight DNA — typically micrograms. We provide extraction protocols or accept extracted HMW DNA with QC.
intermediateWhich assemblers do you use?
hifiasm (HiFi + ultra-long ONT integration, built-in phasing) and Verkko, chosen per genome. Both produced the current generation of complete genomes.
intermediateHow is assembly quality measured?
QV (base accuracy), BUSCO (gene completeness), Merqury (k-mer consistency), contig N50, and per-chromosome gap status. All are reported.
intermediateCan you assemble polyploid plant genomes?
Yes — with subgenome-aware phasing strategies (HiFi + Hi-C, sometimes parental data). See our plant genome sequencing page.
advancedIs full T2T guaranteed?
No honest provider can guarantee gapless status for every chromosome — centromere architecture varies. We agree QC targets at design stage and report per-chromosome status transparently.
advancedCan you phase the assembly into haplotypes?
Yes — hifiasm produces haplotype-resolved assemblies; Hi-C or parental sequencing strengthens chromosome-scale phasing.
advancedWhat data volume does a T2T project need?
Typically ~30× HiFi plus ~30–60× ultra-long ONT (genome-sized), plus optional Hi-C. Exact depths are scoped per genome size and repeat content during design.
advancedCan this support a pangenome project?
Yes — multi-individual T2T assemblies are the foundation of pangenomes; we support cohort-scale pipelines and submission formatting.
References

Key literature & resources

Selected sources behind the methods and reference values on this page. Figures labelled "schematic" or "illustrative" are generated for this page, not client data.

  1. Nurk S, et al. The complete sequence of a human genome. Science. 2022;376(6588):eabj6987. (T2T-CHM13.)
  2. Cheng H, et al. Haplotype-resolved de novo assembly with phased assembly graphs (hifiasm). Nature Methods. 2021;18(2):170–175. (HiFi + ultra-long ONT integration: Cheng H, et al. Nature Methods. 2024.)
  3. Rautiainen M, et al. Telomere-to-telomere assembly of diploid chromosomes with Verkko. Nature Methods. 2023;20(10):1544–1552.
  4. Alonge M, et al. Automated assembly scaffolding using RaGOO / Hi-C: see also Dudchenko O, et al. De novo assembly of the Aedes aegypti genome using Hi-C. Science. 2017;356(6333):92–95.
  5. Formenti G, et al. Merqury: reference-free assessment of assembly quality: Rhie A, et al. Merqury. Genome Biology. 2020;21(1):245. (VGP quality standards: Rhie A, et al. Towards complete and error-free genome assemblies of vertebrate genomes. Science. 2021;374(6566):367–373.)
  6. Simão FA, et al. BUSCO: assessing genome assembly and annotation completeness. Bioinformatics. 2015;31(10):1670–1676.
  7. Hoyt SJ, et al. From telomere to telomere: the transcriptional and epigenetic state of human repeat elements. Science. 2022;376(6588):eabk3112.