What is long-read
sequencing?
Long-read sequencing provides more complete genomic context
Long-read sequencing is a DNA and RNA sequencing approach that reads long, continuous molecules, often thousands of bases at a time. By preserving more molecular context in each read, long-read sequencing can help researchers build more complete genomes, resolve complex variants, and study biology that short reads may miss.
Most sequencing workflows fall into two broad categories: short-read and long-read sequencing. The main difference is read length. Short-read sequencing analyzes smaller fragments, usually a few hundred bases long. Long-read sequencing reads much longer fragments, preserving more context from the original DNA or RNA molecule.
That added context can make genomes easier to assemble and interpret. Long reads can span repeats, structural variants, and other complex regions that are difficult to resolve with short reads alone.
PacBio HiFi sequencing combines long reads with high accuracy, giving researchers a clearer view of complex biology across human genomics, cancer research, microbial genomics, plant and animal genomics, biodiversity, and transcriptomics
Comparison of short-read sequencing and PacBio HiFi long-read sequencing mapped to a reference genome. HiFi reads span longer regions with continuous, accurate coverage, helping researchers detect variants in complex regions.
If you’re studying rare disease, cancer, complex genomes, microbial evolution, or biodiversity, better context can lead to better answers. PacBio HiFi sequencing is designed to deliver accurate long reads that help researchers see more of the genome in a single workflow.
Long-read sequencing was named 2022 Method of the Year by Nature Methods, reflecting growing recognition of its ability to resolve regions and variant types that short reads can miss. Today, HiFi sequencing continues to advance long-read sequencing for genome, transcriptome, methylome, and epigenome research.
Watch how long-read sequencing works and how HiFi technology generates highly accurate long reads by sequencing the same DNA molecule multiple times.
Video: How long-read sequencing works using HiFi sequencing to produce accurate, high-quality long reads.
Benefits of long-read sequencing
Long-read sequencing provides more context for genome assembly
With long-read sequencing, you can capture continuous stretches of DNA that are tens of thousands of bases long. PacBio HiFi sequencing routinely delivers read lengths of up to 25 kb. These long reads make it easier to see how different parts of the genome fit together, which simplifies assembly and reveals patterns that shorter reads might miss.
High-accuracy long-read sequencing improves data quality
The strongest long-read sequencing data combines length with accuracy. PacBio HiFi reads deliver long reads with up to 99.95% accuracy, helping researchers generate confident results while reducing the need for downstream error correction or polishing.
Long-read sequencing helps resolve repeats, GC-rich regions, and structural variants
Certain parts of the genome like GC-rich regions, tandem repeats, and complex variants, are tough to analyze with short reads. PacBio HiFi reads can span the full length of challenging genomic regions that short reads struggle to resolve. PureTarget panels target clinically relevant genes buried in these regions with a single amplification-free assay and shine a light on previously “dark” regions of the genome that might be missed with traditional hybrid capture or amplicon short-read approaches.
Long-read sequencing supports research across rare disease, cancer, public health, and biodiversity
Highly accurate long-read sequencing can support research and public health programs by revealing biology that short reads may miss. In cancer research, long reads can uncover structural variants and gene fusions. In rare disease research, they can improve variant detection and phasing. In public health, long reads support pathogen surveillance and antimicrobial resistance tracking. In biodiversity, they help build high-quality reference genomes for conservation and population studies.
Want to brush up on long-read sequencing fundamentals?
Curious about what’s next for HiFi technology?
Long-read sequencing vs. short-read sequencing
Historically, researchers often had to choose between read length and accuracy. Short-read sequencing offered high accuracy but limited molecular context, while older long-read approaches captured larger genomic regions with lower precision.
HiFi sequencing changes that trade-off. PacBio HiFi reads deliver long-read lengths of up to 25 kb with >99.9% accuracy, helping researchers detect SNVs, indels, structural variants, methylation, and transcript isoforms with greater context in a single sequencing approach.
Comparison of sequencing technologies by read length and accuracy. HiFi reads are both long and highly accurate, outperforming short reads and Nanopore reads.
| PacBio HiFi sequencing | Illumina SBS | |
| Average read length | 500 bp-20kb | 2x150 bp |
| Average read accuracy | 99.95% | 99.92% (Q31) |
| Coverage | Unbiased | Reduced at low and high [GC] |
| Variant calling: SNVs | ||
| Variant calling: Indels | ||
| Variant calling: SVs | X | |
| Genome assembly: contiguity | X | |
| Genome assembly: accuracy | ||
| Epigenetics: 5mC, 5hmC, and 6mA methylation | X | |
| Isoform repertoire | Complete view | Partial view |
| Precision | recall: SNVs | 99.9 | 99.9 | 99.8 | 99.4 |
| Precision | recall: Indels | 99.4 | 99.4 | 99.7 | 99.5 |
| Precision | recall: SVs | 96.1 | 96.0 | 94.7 | 62.4 |
Learn how HiFi sequencing works any why accurate long reads can change what researchers see.
Long-read sequencing for multiomics
The benefits of long reads extend beyond the genome. Because long-read sequencing combines extended read length with high accuracy, it unlocks multiomic insights across the genome, transcriptome, methylome, and epigenome, delivering richer, more reliable biological information in a single assay.
Genome
Long-read sequencing delivers accurate, contiguous reads that reduce gaps, resolve structural variants, and span complex regions that short reads struggle to analyze. This helps researchers assemble high-quality reference genomes across humans, plants, animals, and microbes.
Transcriptome
With full-length long reads, researchers can explore complete transcript isoforms, alternative splicing events, and novel transcripts. Kinnex RNA sequencing kits use HiFi sequencing to capture full-length transcripts, providing a more detailed view of isoform structure than gene expression alone.
Methylome
HiFi long-read sequencing enables direct detection of base modifications in native DNA without bisulfite conversion. Researchers can capture sequence, methylation, and full-molecule context in the same experiment.
Epigenome
Fiber-seq is a long-read whole genome sequencing assay that maps genetic variation, chromatin accessibility, nucleosome position, CpG methylation, and bound transcription factors on individual chromatin fibers.
Proof points of long-read sequencing
These real-world examples show how HiFi long-read sequencing is being used in population genomics, rare disease research, clinical genomics research, public health, and microbial genomics. Together, they show how accurate long reads can improve variant detection, reduce assembly gaps, shorten project timelines, and expand what researchers can see.
All of Us study: HiFi sequencing helps uncover disease-relevant variants missed by short reads
A major population genomics program used HiFi sequencing to uncover disease-relevant genetic variation that had been overlooked by short-read methods. This study highlights how long-read sequencing can improve representation, resolution, and equity in genomics research across diverse populations.
Berry Genomics uses HiFi sequencing for complex single-gene disorder research
Berry Genomics adopted HiFi sequencing to address the complexity of single-gene disorders. By revealing previously undetected variants, long-read technology helped improve clinical research outcomes and demonstrated clear advantages over traditional approaches in public health.
HiFi Solves EMEA study shows the clinical research potential of a complete HiFi genome
A collaborative European study shows how HiFi sequencing delivers complete, interpretable genome data, including challenging variant types, for clinical genomics. The findings point to the growing role of long-read sequencing as a comprehensive solution for genome analysis in translational research and precision medicine.
HiFi sequencing fast-tracks antimicrobial resistance research with complete plasmid assemblies
Researchers at Charles University increased their annual publications by adopting a HiFi long-read workflow that replaced lengthy fragmented assemblies with complete, circularized plasmid sequences. This long-read workflow reduced project timelines from months to a single sequencing run, demonstrating the practical advantages of long-read sequencing for public health and microbial genomics.
Read more customer success stories to see how researchers use HiFi long-read sequencing to resolve complex genomes, shorten project timelines, and generate more complete insights.
