Element Biosciences, Inc.

Illumina Corporation

Illumina Cambridge Limited

Illumina Singapore Pte. Ltd.

Matthew William Kellinger

Michael Previte

Molly Min He

Sinan Arslan

Junhua Zhao

Chunhong Zhou

Tyler Lopez

Jonathan Mark Boutell

Erin Bomati

Samantha Snow

William Light

Michael Klein

Virginia Saade

Mark Ambroso

Lei Sun

Hua Yu

Cheng-Yao Chen

Yu-Hsien Hwang-Fu

Michael Kim

Hui Zhen Mah

Marco Tjioe

Andrew Boddicker

Kevin L Gunderson

Boyan Boyanov

Roberto Rigatti

Maria Candelaria Rogert Bacigalupo

Jingwei Bai

John M Beierle

Sergio Peisajovich

Klaus Maisinger

Misha Golynskiy

Adeline Huizhen Mah

Minghao Guo

Lorenzo Berti

Christian Gloeckner

Steve Xiangling Chen

Saurabh Nirantar

Seth McDonald

Lea Pickering

Leon Zilun Zhang

Jendrik Hentschel

Jake Levieux

Su Zhang

Xiaodong Qi

Jendrik Boothby-Hentschel

Kelly Wiseman

Hui Zhen Ma

Compositions and methods comprising the use of polymerase-nucleotide conjugates for nucleic acid sequencing and analysis applications are described.

Polymerase-nucleotide conjugates for sequencing by trapping

The present disclosure relates to the field of molecular biology and more specifically to microarrays and methods. In particular, the present disclosure relates to a method for amplifying a nucleic acid on a substrate comprising at least one well, the method includes: depositing a plurality of first capture primer pairs and a plurality of second capture primer pairs in a layer at least partially covering the inner well surface, wherein the primer density of first capture primer pairs is higher than the primer density of second primer pairs; contacting a sample comprising a plurality of target polynucleotides with the substrate under conditions sufficient for a single target polynucleotide per well to hybridize with a capture primer of the second capture primer pair; and performing a kinetic exclusion assay (KEA) to produce a monoclonal population of amplicons from the single target polynucleotide.

Methods and arrays for producing and sequencing monoclonal clusters of nucleic acid

The present disclosure provides compositions comprising nucleic acid double-stranded splint adaptors, including kits, and methods that employ the double-stranded splint adaptors. The double-stranded splint adaptors () can be used in a one-pot, multi-enzyme reaction to introduce one or more new adaptor sequences into a library molecule. The double-stranded splint adaptor () comprises a first splint strand (long splint strand ()) and a second splint strand (short splint strand ()), where the first and second splint strands are hybridized together to form the double-stranded splint adaptor () having a double-stranded region and two flanking single-stranded regions. The second splint strand () carries the new adaptor sequence(s) to be introduced, such as for example a universal binding sequence, an index sequence and/or a random sequence.

Double-stranded splint adaptors with universal long splint strands and methods of use

Presented herein are polymerase-linked nucleotides for improved distinguishing nucleotide sequences for different nucleic acid molecules. Also presented are methods and systems using the polymerase-linked nucleotides for improved distinguishing nucleotide sequences for different nucleic acid molecules.

Enzyme-linked nucleotides

Multivalent binding compositions including a particle-nucleotide conjugate having a plurality of copies of a nucleotide attached to the particle are described. The multivalent binding compositions allow one to localize detectable signals to active regions of biochemical interaction, e.g., sites of protein-protein interaction, protein-nucleic acid interaction, nucleic acid hybridization, or enzymatic reaction, and can be used to identify sites of base incorporation in elongating nucleic acid chains during polymerase reactions and to provide improved base discrimination for sequencing and array based applications.

Multipart reagents having increased avidity for polymerase binding

Provided herein are engineered variants of archaeal, prokaryotic, and eukaryotic polymerases that exhibit enhanced thermostability, enhanced incorporation of 3' modified nucleotides, and improved uracil-tolerance, in polymerase-catalyzed nucleotide extension reactions relative to wild type polymerase enzymes. Also provided are uses of the engineered polymerases for forming complexed polymerases, forming binding complexes and forming ternary complexes, and uses for conducting nucleic acid sequencing reactions.

Engineered polymerases

Multivalent binding composition for nucleic acid analysis

Provided herein are engineered variants of archaeal polymerases that exhibit exonuclease-minus activity, enhanced thermostability, enhanced incorporation of 3' modified nucleotides, improved uracil-tolerance and/or reduce sequence-specific errors in polymerase-catalyzed nucleotide binding and extension reactions relative to wild type polymerase enzymes. Also provided are uses of the engineered polymerases for forming complexed polymerases and forming binding complexes, and uses for conducting nucleic acid sequencing reactions.

Engineered polymerases with reduced sequence-specific errors

Provided herein are methods for generating circular nucleic acid molecules and circular nucleic acid libraries. The methods can be used to generate clonal populations of target nucleic acid molecules for downstream applications such as sequencing.

Methods for generating circular nucleic acid molecules

Provided herein are compositions and methods for the incorporation of unnatural nucleotides using mutant polymerases, such as reverse transcriptases. Further provided herein are methods of detection and sequencing of polynucleotide sequences. In some aspects, the compositions and methods are used enhance the efficiency and speed of detecting nucleotide bases. The methods and compositions described herein may further reduce time, cost, or scale of devices for next generation sequencing platforms.

Reverse transcriptase for nucleic acid sequencing

The present disclosure provides compositions and methods that employ the compositions for conducting pairwise sequencing and for generating concatemer template molecules for pairwise sequencing. The concatemers can be generated using a rolling circle amplification reaction which is conducted either on-support, or conducted in-solution and then distributed onto a support. The rolling circle amplification reaction generates concatemers containing tandem copies of a sequence of interest and at least one universal adaptor sequence. An increase in the number of tandem copies in a given concatemer increases the number of sites along the concatemer for hybridizing to multiple sequencing primers which serve as multiple initiation sites for polymerase-catalyzed sequencing reactions. When the sequencing reaction employs detectably labeled nucleotides and/or detectably labeled multivalent molecules (e.g., having nucleotide units), the signals emitted by the nucleotides or nucleotide units that participate in the parallel sequencing reactions along the concatemer yields an increased signal intensity for each concatemer.

Compositions and methods for pairwise sequencing

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San Diego, CA, United States of America

Matthew William Kellinger