Long-Read Genome Assembly Service — Telomere-to-Telomere & Haplotype-Resolved

Complete genome assembly by PacBio HiFi and Oxford Nanopore PromethION — de novo assembly, reference-guided assembly, and hybrid strategies for microbial, fungal, plant, animal, and human genomes — producing chromosome-level, telomere-to-telomere assemblies with comprehensive structural and functional annotation

CD Genomics provides genome assembly services on PacBio HiFi (Revio/Sequel II), Oxford Nanopore PromethION, and Illumina NovaSeq platforms — from bacterial single-contig chromosomes and fungal telomere-to-telomere assemblies to plant and animal genomes with Hi-C chromosome-scale scaffolding, comprehensive structural and functional annotation, and NCBI submission support.

Genome assembly is the computational process of reconstructing complete genome sequences from the short DNA fragments (reads) produced by sequencing instruments. From the first bacterial genomes assembled by Sanger sequencing in the 1990s to the telomere-to-telomere (T2T) human genome completed in 2022, genome assembly has been transformed by advances in sequencing technology — most profoundly by the emergence of long-read sequencing platforms that produce reads tens to hundreds of kilobases in length, capable of spanning repetitive regions and structural complexities that were intractable to short-read methods.

At CD Genomics, we provide comprehensive genome assembly services across all major long-read and short-read platforms — PacBio HiFi (Sequel II / Revio), Oxford Nanopore (PromethION R10.4.1), Illumina NovaSeq, and Hi-C proximity ligation — supporting the full range of genome assembly projects: from bacterial and fungal genomes (1–100 Mb) to plant, animal, and human genomes (100 Mb to 20+ Gb). Our platform-agnostic approach ensures that each project receives the optimal sequencing and assembly strategy for its specific genome size, complexity, ploidy, and research objectives.

Whether you need a complete bacterial genome in a single contig, a haplotype-resolved assembly of a highly heterozygous plant, or a telomere-to-telomere reference genome for a non-model organism, our team of experienced bioinformaticians and molecular biologists will design and execute a tailored genome assembly solution — from DNA extraction and sequencing library preparation through assembly, curation, and comprehensive annotation — delivering publication-ready genome assemblies with full documentation.

Why Choose Our Genome Assembly Service

• Multi-Platform Assembly Pipeline — Integration of PacBio HiFi (Q30+ accuracy), ONT ultra-long reads (50–100+ kb), Illumina polishing, and Hi-C scaffolding — selecting the optimal combination for each genome’s unique characteristics
• Telomere-to-Telomere Quality — Our assembly pipelines are designed to produce complete chromosomal pseudomolecules with telomeric repeats at both ends, zero spanned gaps, and resolved centromeric and rDNA arrays — assemblies that meet or exceed the quality standards of published reference genomes
• Haplotype-Resolved & Diploid Assembly — For heterozygous, diploid, and polyploid genomes, our PacBio HiFi + hifiasm pipeline produces phased assemblies that separate haplotypes and resolve allelic variation — essential for accurate gene annotation, evolutionary genomics, and breeding applications
• Cross-Kingdom Expertise — Dedicated assembly protocols for bacteria, fungi, plants, animals, and viruses — each optimized for the specific genome size, repeat content, GC composition, and ploidy of the target taxonomic group
• Comprehensive Annotation — Beyond genome assembly, we provide structural and functional annotation including repeat annotation, gene prediction, non-coding RNA identification, and kingdom-specific functional databases (CAZy, antiSMASH, PHI-base for fungi; PLAZA/Ensembl Plants for plants; RefSeq/VEGA for animals)
• End-to-End Project Management — From initial project consultation and experimental design through sequencing, assembly, annotation, and NCBI submission support — a dedicated project scientist manages every stage with regular progress updates

Genome Assembly Progressed from Fragmented Drafts to Telomere-to-Telomere Completeness Through Long-Read Sequencing

Genome assembly — the process of reconstructing a complete genome sequence from sequencing reads — is the foundational step in nearly all genomics research. The quality of the genome assembly determines virtually everything that follows: the accuracy of gene prediction, the completeness of repeat annotation, the reliability of comparative genomics, and the interpretability of population genetic analyses. A fragmented or error-prone assembly propagates uncertainty through every downstream analysis; a complete, accurate assembly enables confident biological discovery.

The transition from short-read to long-read sequencing has been the single most transformative advance in genome assembly. Short-read platforms (Illumina) produce reads of 150–300 bp with very high accuracy (Q30+), but the fundamental limitation of read length means that repetitive regions longer than the read length cannot be uniquely resolved. Since most eukaryotic genomes contain 30–80% repetitive sequence — including transposable elements, tandem repeats, rDNA arrays, and centromeric satellites — short-read assemblies are necessarily fragmented into thousands or tens of thousands of contigs, with collapsed repeats, missing telomeres, and misassembled structural variants.

Long-read sequencing overcomes this limitation fundamentally. PacBio HiFi reads (15–25 kb, Q30+) provide the accuracy of short reads with 100× the read length, enabling single-copy resolution of most repetitive elements. Oxford Nanopore ultra-long reads (50–100+ kb, and up to 2 Mb) can span entire centromeric repeat arrays, large rDNA clusters, and complex structural rearrangements in a single contiguous read. When combined — HiFi for base-level accuracy and haplotype phasing, ultra-long ONT reads for bridging the largest repeats, and Hi-C for chromosome-scale scaffolding — modern long-read assembly pipelines routinely produce telomere-to-telomere genomes with zero gaps, fully resolved centromeres, and complete haplotype information.

At CD Genomics, we offer genome assembly services for organisms across the tree of life. Our service is organized by genome size and complexity tier — from microbial genomes (which we can complete in a single contig per chromosome) to large plant and animal genomes (where we integrate HiFi, ultra-long ONT, and Hi-C for chromosome-scale T2T assemblies) — with dedicated pipelines and bioinformatic expertise for each category.

Our Genome Assembly Pipelines Cover Bacterial, Fungal, Plant, Animal, and Viral Genomes with Taxon-Specific Optimization

Beyond reference-grade assemblies for individual organisms, long-read genome assembly is increasingly applied in pangenome construction and comparative genomics across populations, where complete, haplotype-resolved assemblies from multiple individuals of the same species enable discovery of gene presence-absence variation, structural variant landscapes, and accessory genome content that single-reference approaches systematically miss. International initiatives such as the Vertebrate Genomes Project (VGP), Earth BioGenome Project (EBP), and species-specific pangenome consortia (human, plant, livestock) increasingly rely on long-read assembly as the foundational technology for producing the reference-quality genomes that power cross-species comparative analysis and population genomics.

We provide dedicated genome assembly pipelines optimized for each major taxonomic group. Click on each service for detailed information.

Microbial Genome Assembly (Bacteria & Archaea)

Complete bacterial genome assembly from single-contig circular chromosomes to multi-replicon genomes with plasmid reconstruction. Our bacterial WGS de novo sequencing service delivers complete, circularized genomes with automatic plasmid detection and comprehensive functional annotation (COG, KEGG, GO, CARD, VFDB, CAZy, antiSMASH). For strains with an existing reference, our bacterial WGS resequencing service provides cost-effective variant discovery.

Fungal Genome Assembly

Telomere-to-telomere genome assembly for yeasts, filamentous fungi, and mushrooms — from compact 10 Mb yeast genomes to large repeat-rich filamentous fungal genomes exceeding 100 Mb. Our fungal WGS de novo sequencing service features fungal-specific annotation including CAZy carbohydrate-active enzymes, antiSMASH secondary metabolite clusters, and PHI-base effector prediction.

Plant & Animal Genome Assembly

Chromosome-scale genome assembly for plants and animals, integrating PacBio HiFi, ultra-long ONT, and Hi-C proximity ligation for T2T pseudomolecules. Our services cover genome size estimation, repeat characterization, genome assembly, and comprehensive annotation — from small invertebrate genomes to large vertebrate and plant genomes exceeding 10 Gb. Contact us to discuss your project.

Viral Genome Assembly

Complete genome sequencing and assembly for DNA and RNA viruses, combining short-read and long-read platforms for maximum coverage of viral genomes, quasispecies detection, and identification of recombinant or novel viral strains. Genome assembly from direct RNA sequencing available for RNA viruses without reverse transcription bias.

Our Bioinformatics Delivers Assembly QC, Repeat and Gene Annotation, Epigenome Analysis, and Comparative Genomics

Our bioinformatics pipeline supports comprehensive genome analysis from raw read processing through assembly, quality assessment, and multi-level annotation — organized across two service tiers.

Sample Input Requirements for Genome Assembly — DNA Quality, Coverage Targets, and Multiplexing Options for Each Platform

Case Study: Telomere-to-Telomere Genome Assembly of Sorghum Using PacBio HiFi, ONT Ultra-Long, and Hi-C

Wang F, Bao J, Zhang H, Zhai G, Song T, Liu Z, Han Y, Yu F, Zou G, Zhu Y. A telomere-to-telomere genome assembly of Chinese grain sorghum 654. Scientific Data. 2025;12:460. doi:10.1038/s41597-025-04791-6.

1. Background

Sorghum (Sorghum bicolor) is the world’s fifth most important cereal crop and a key model for C4 photosynthesis research, drought tolerance, and bioenergy production. The existing reference genome for sorghum (BTx623) was a landmark assembly but contained approximately 4,000 spanned gaps — unresolved regions including complex centromeric repeats, rDNA arrays, and telomeric sequences — that limited comprehensive genomic analysis. The Chinese grain sorghum inbred line 654 is an elite variety with important agronomic traits, but no complete genome sequence was available for this or any sorghum line.

In this study, Wang et al. generated a complete, gap-free T2T genome assembly of sorghum line 654 using an integrated multi-platform long-read sequencing strategy — demonstrating the current state of the art in plant genome assembly.

2. Methods

The genome was assembled using three complementary sequencing technologies: PacBio HiFi (Revio, 36.4× coverage, 15–20 kb reads), ONT ultra-long reads (PromethION R10.4.1, 24.3× coverage, N50 read length >50 kb), and Hi-C proximity ligation (95.7× coverage). Initial assembly was performed with hifiasm using HiFi reads, scaffolded to chromosome-level with Hi-C data, and gaps filled using ONT ultra-long reads. Assembly quality was validated by multiple complementary methods including k-mer completeness (Merqury), BUSCO gene completeness, Hi-C contact map consistency, and manual inspection of all telomere and centromere junctions.

3. Results

Figure 2. Telomere-to-telomere genome assembly of sorghum line 654 using integrated PacBio HiFi + ONT ultra-long + Hi-C strategy. The assembly (728.81 Mb) achieved zero gaps, all 10 centromeres and 20 telomeres fully resolved, with a QV score of 64.72 and 44,399 annotated protein-coding genes. Adapted from Wang et al. (2025), Scientific Data, CC BY 4.0.

Key Findings

• Zero-gap T2T assembly: The final genome assembly (728.81 Mb) was completely gap-free — all 10 centromeres and all 20 telomeres were fully resolved with no spanned gaps, a milestone not achieved in any previous sorghum genome assembly.
• Centromere resolution: All 10 centromeric regions were fully assembled, ranging from 135 to 574 kb, revealing the complete structure of sorghum centromeric satellite arrays and centromere-specific retrotransposons.
• Telomere completion: All 20 chromosome ends were capped with canonical plant telomere repeats (TTTAGGG)n, with telomere lengths ranging from 2.5 to 18.8 kb — information completely absent from the fragmented reference assembly.
• Annotation quality: The assembly contained 44,399 protein-coding genes with a BUSCO completeness of 97.5%, QV score of 64.72 (error rate <3.4 × 10⁻⁷), and 68.0% repeat content fully characterized.

4. Conclusions

This study demonstrates that integrated multi-platform long-read sequencing — combining the accuracy of PacBio HiFi with the repeat-spanning capability of ONT ultra-long reads and the chromosome-scale information of Hi-C — can produce complete, telomere-to-telomere plant genome assemblies of the highest quality. The approach and coverage specifications serve as a benchmark for plant genome assembly projects: ~36× HiFi for accuracy, ~24× ONT ultra-long for gap closure, and ~100× Hi-C for scaffolding. The methods are directly applicable to other plant and animal genomes of similar or larger size.

Interpretation Boundaries for Long-Read Genome Assembly

• Assembly contiguity and completeness depend on genome complexity and data quality. While long-read assemblies dramatically outperform short-read assemblies, the achievable contiguity remains genome-dependent. Genomes with extremely high repeat content (>70%), large segmental duplications, or very high heterozygosity may still contain regions that cannot be fully resolved regardless of sequencing depth. The assembly quality achievable for each genome is assessed during project design and documented in the project report
• Coverage requirements are genome-specific and should be validated empirically. Coverage recommendations (30–100× HiFi, 40–100× ONT) are based on published benchmarks across diverse genome types. Specific genomes may require different coverage levels depending on their unique characteristics. Pilot sequencing at lower coverage can provide empirical data for optimizing the final sequencing strategy
• Annotation quality is limited by available evidence. Gene prediction accuracy depends on the availability of species-matched transcript evidence (RNA-seq, Iso-Seq), protein homology databases, and ab initio training sets. For novel or deeply divergent species with limited existing genomic resources, annotation completeness may be lower than for well-characterized model organisms. Annotation quality metrics are provided with all deliverables
• Reference genome choice influences comparative results. Alignment-based analyses using a long-read assembly as the reference will produce different results than analyses using a short-read or alternate assembly as the reference. Any reference-specific effects on variant detection or comparative analyses should be considered when designing downstream studies
• All genome assembly data are generated for research use only. Our sequencing and assembly services are intended for research applications and are not validated for clinical diagnosis, treatment decisions, or clinical variant interpretation in individual patients or specimens

Genome Assembly Deliverables Include Complete Chromosome Sequences, Annotated GenBank Files, and Publication-Ready Reports

Deliverable Examples for Genome Assembly Projects

1. Complete genome assembly file (FASTA) with chromosome-level pseudomolecules — one sequence per chromosome with telomeric repeats at both ends, plus mitochondrial genome and any accessory or extrachromosomal elements.

2. Genome annotation file (GFF3/GenBank format) with structural and functional annotations — protein-coding genes, tRNA, rRNA, ncRNA, repeat annotations, and predicted functional domains with cross-references to kingdom-specific databases.

3. Hi-C contact map and chromosome-scale scaffolding validation — interactive heatmap showing chromatin interaction patterns and evidence supporting chromosomal pseudomolecule assignments.

4. Comprehensive project report PDF documenting experimental methods, sequencing QC, assembly statistics, annotation summary, and detailed methods text suitable for manuscript preparation.

Figure 3. Representative deliverable formats for genome assembly projects. Left: chromosome-scale genome map with multi-track annotation. Center: Hi-C contact map validating chromosome-scale scaffolding. Right: assembly quality metrics and functional annotation summary. AI-generated representative data.