Ncan Genomics, Inc.

University of British Columbia

Boreal Genomics Corp.

Quantum-si Incorporated

Joel Pel

Andrea Marziali

Milenko Despotovic

Gosuke Shibahara

Jason Donald Thompson

David John Broemeling

Matthew Wiggin

Peter Jason Eugster

Dylan Corey Gunn

The invention generally relates to capturing, amplifying, and sequencing nucleic acids. In certain embodiments, copies of the sense and antisense strands of a duplex template nucleic acid are captured using linked capture probes and multiple binding and extension steps to improve specificity over traditional single binding target capture techniques. Methods of seeding sequencing clusters with sense and antisense strands of a target nucleic acid are also disclosed including identifying the strands using sense-specific barcodes and confirming base calls using two sense-specific sequencing reads. Linked adapters may be used to increase adapter ligation selectively or efficiency and yield.

Linked ligation

The invention generally relates to sequencing library preparation methods. In certain embodiments, two or more template nucleic acids are joined together by a linking molecule, such as a PEG derivative. Identical copies of a nucleic acid fragment or both strands of a duplex fragment may be linked together. The linked nucleic acids are amplified, creating linked amplicons. Emulsion PCR with linked primers creates linked template nucleic acids for seeding sequencing clusters and errors can be readily identified by their presence on only one of the linked fragments.

Linked target capture

The invention generally relates to sequencing library preparation methods. In certain embodiments, two template nucleic acids are joined together by a linking molecule, such as a PEG derivative. The linked template nucleic acids is amplified, creating linked amplicons.

Methods of sequencing linked fragments

The invention generally relates to capturing, amplifying, and sequencing nucleic acids. In certain embodiments, linked capture probes and multiple binding and extension steps improve specificity over traditional single binding target capture techniques. Methods of seeding sequencing clusters with captured target nucleic acids are also disclosed. Linked adapters may be used to increase adapter ligation selectively or efficiency and yield. Ligation adapters and primers can be linked to various sequence-specific or feature-specific molecules to selectively bind targets for ligation or amplification with universal adapters or primers.

Linked target capture and ligation

The invention includes methods and apparatus for separating mutations, especially rare and unknown mutations, using heteroduplex binding proteins. Nucleic acids may optionally be nicked at or near the mutation in order to promote heteroduplex binding protein recognition and binding. In particular, using the disclosed methods, it is possible to separate heteroduplexed nucleic acid strand pair from homoduplexed nucleic acid strand pairs having similar sequences and being at a much higher concentration. Once the heteroduplexed nucleic acids are isolated and recovered, it is straightforward to analyze the sequences of the heteroduplexed nucleic acids, e.g., using sequencing or hybrid assays.

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Vancouver, Canada

Joel Pel