Sequencing by Expansion (SBX) — a novel, high-throughput single-molecule sequencing technology

Mark Kokoris
Robert McRuer
Melud Nabavi
Aaron Jacobs
Marc Prindle
Cynthia Cech
Kendall Berg
Taylor Lehmann
Cara Machacek
John Tabone
Jagadeeswaran Chandrasekar
Lacey McGee
Matthew Lopez
Tommy Reid
Cara Williams
Salka Barrett
Alex Lehmann
Michael Kovarik
Robert Busam
Scott Miller
Brent Banasik
Brittany Kesic
Anasha Arryman
Megan Rogers-Peckham
ALAN KIMURA
Megan LeProwse
Mitchell Wolfin
Svetlana Kritzer
Joanne Leadbetter
Majid Babazedeh
Majid Babazadeh
John Chase
Greg Thiessen
William Lint
Drew Goodman
Dylan O’Connell
Nadya Lumanpauw
John Hoffman
Samantha Vellucci
Kendra Collins
Jessica Vellucci
Amy Taylor
Molly Murphy
Michael Lee
Matthew Corning
Publication typePosted Content
Publication date2025-02-24
Abstract

Remarkable advances in high-throughput sequencing have enabled major biological discoveries and clinical applications, but achieving wider distribution and use depends critically on further improvements in scale and cost reduction. Nanopore sequencing has long held the promise for such progress, but has had limited market penetration. This is because efficient and accurate nanopore sequencing of nucleic acids has been challenged by fundamental signal-to-noise limitations resulting from the poor spatial resolution and molecular distinction of nucleobases. Here, we describe Sequencing by Expansion (SBX), a single-molecule sequencing technology that overcomes these limitations by using a biochemical conversion process to encode the sequence of a target nucleic acid molecule into an Xpandomer, a highly measurable surrogate polymer. Expanding over 50 times longer than the parent DNA templates, Xpandomers are engineered with high signal-to-noise reporter codes to enable facile, high-accuracy nanopore sequencing. We demonstrate the performance of SBX and present the specialized molecular structures, chemistries, enzymes and methods that enable it. The innovative molecular and systems engineering in SBX create a transformative technology to address the needs of existing and emerging sequencing applications.

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