Introduction
Next-generation sequencing methods, primarily sequencing-by-synthesis (SBS), have advanced significantly over the past 25 years, making it a widely used technique in genomic applications. While Illumina short-read whole-genome sequencing (srWGS) achieves highly accurate coverage over most of the human genome,1 specific regions of the genome have remained challenging to resolve. Additionally, some classes of genomic variation, including large structural rearrangements, are difficult to detect using standard short reads. Longer read lengths have a demonstrated ability to address some of these challenges but are comparatively costly and challenging to scale, with high input amounts and stringent input requirements.
Here, we introduce constellation mapped read technology, an innovative approach that leverages on-flow-cell library prep and informatics that utilize proximity information from clusters in neighboring nanowells to generate long-range genomic insights using standard SBS sequencing. Constellation technology maintains the accuracy, depth of coverage, and scalability of standard SBS while adding the phasing, enhanced mappability, and improved structural variant detection often associated with long-read methods. This novel approach provides a powerful, accessible solution for comprehensive whole-genome analysis.
Consult the end of this post for a glossary of terms.
A novel technology
Constellation technology is unlike other methods that generate long-distance information from standard short-read sequencing (Hi-C or linked-reads, for example). Constellation tech eliminates manual library prep by applying extracted DNA directly to the flow cell surface, where surface-bound transposomes perform tagmentation in situ. This on-flow-cell library prep ensures that adjacent regions in a sample’s genome remain physically proximal on the flow cell. Instead of relying on complex tagging, molecular barcodes, or long contiguous reads, constellation tech leverages the spatial proximity of neighboring clusters to unlock long-range genomic information from a sample’s genome using standard SBS sequencing, resulting in:
- Improved mapping and variant calling in difficult-to-map regions
- Ultra-long phasing, up to several megabases
- Improved calling of large (> 50 bp) structural rearrangements
How it works
Highly simplified, on-flow-cell workflow
The constellation workflow begins with a novel, on-flow-cell library prep that uses the low DNA inputs characteristic of transposome-based library prep workflows and the high sequencing quality of the NovaSeq™ X series. The experimental workflow requires no modifications to the sequencing instrument—only a custom sequencing recipe, making it accessible to a large existing install base.
The constellation experimental workflow:
- Add extracted DNA template to library strip tube
- Add specialized reagents to the custom primer wells of the sequencing cartridge
- Load consumables and initiate run
