Microsoft Technology Licensing, LLC

Advanced Micro Devices Corporation

University of Washington

Karin Strauss

Bichlien Hoang Nguyen

Yuan-Jyue Chen

Luis Henrique Ceze

Sergey M Yekhanin

Douglas Christopher Burger

Jake Smith

John D Davis

Mark Steven Manasse

Hsing-Yeh Parker

Parikshit Santhan Gopalan

Oriana Riva

Robert Nikolaus Grass

Alexander Xavier Christof Kohll

Weida Chen

Siena Dumas Ang

Robert Carlson

Gabriel H Loh

Douglas Patrick Kelley

Jaewoong Chung

Doug Burger

Douglas M Carmean

Kalin Ovtcharov

Gagan Gupta

Richard P Rouse

Adrian Sampson

Henrique Sarmento Malvar

Timothy Lawrence Harris

Eric S Chung

James R Larus

Georg Seelig

Jaron Zepel Lanier

Dimitrios Lymberopoulos

Stuart Schechter

Jeremy P Condit

Timothy Sherwood

Olatunji Ruwase

Joo-Young Kim

Qing Guo

Konstantin Makarychev

Cyrus A Rashtchian

John Mulligan

Lee Organick

Ranveer Chandra

Burton Jordan Smith

Kris Kuppuswamy Ganjam

Alice Jane Bernheim Brush

Richard John Black

Benjamin Zorn

Jeremy Halden Fowers

Dushyanth Narayanan

Peter N Glaskowsky

Kathryn S Mckinley

Henry E Pfister

Orion Tamlin Hodson

Randolph Lopez

Matthew David Turner

Christian Paquin

Sivakanth Gopi

Miklos Zoltan Racz

Eiji Hayashi

Johannes Staffan Anders Linder

Robert Grass

Alexander Steven Crown

Ravindran Kannan

Callie Bee

Jeremy Fowers

Julian Bernhard Koch

Jan Wendelin Stark

Gennady Pekhimenko

Ross Cameron McIlroy

Robert N Grass

Miklos Racz

Cyrus Rashtchian

Peter Glaskowsky

Recycling information is associated with objects through the use of molecular tags. The recycling information may describe the type of material that the object is made from as well as provide instructions for recycling. The molecular tags may be polynucleotides or other types of molecules including inorganic molecules. The molecular tags may be embedded within the object or attached to the surface of the object. At the end of the object's life, the molecular tags are read and the recycling information is used to appropriately recycle the object.

Molecular encoding of recycling information

Polynucleotides such as DNA are stored inside vesicles formed from self-assembling membranes. The vesicles may be protocells, liposomes, micelles, colloidosomes, proteinosomes, or coacervates. The vesicles may include surface functionalization to improve polynucleotide encapsulation and/or to bind polynucleotides having specific sequences. Encapsulation in vesicles provides protection for the polynucleotides. Additional protection is provided by addition of one or more stabilizers. The stabilizer may be nucleic-acid stabilizers that stabilize the polynucleotides or may be a protective structural layer around the vesicles such as a layer of silica. A process for stably storing polynucleotides in vesicles and a process for recovering stored polynucleotides from vesicles are both disclosed. The polynucleotides may be used for storage of digital information.

Polynucleotide encapsulation and preservation using self-assembling membranes

A multiplex assay for nucleic acid detection includes a substrate, a sample, and a fluorophore-labeled oligonucleotide. The substrate has a plurality of physically separated assay locations, each of which includes a nucleotide-targeting enzyme configured to cleave nucleic acids, a guide ribonucleic acid (gRNA), and a quencher-labeled oligonucleotide. A portion of the sample is distributed to each assay location. The gRNA recognizes target nucleic acid in the sample, thereby activating the nucleotide-targeting enzyme to cleave nucleic acids, including the quencher-labeled oligonucleotide. The fluorophore-labeled oligonucleotide is subsequently added to each assay location, which facilitates identification of a presence of the target nucleic acid in the sample via detection of unquenched light emitted by the fluorophore in one or more of the plurality of assay locations.

Multiplex assay for nucleic acid detection

De novo polynucleotide synthesis is performed with a substrate-bound polymerase. The polymerase is attached to a solid substrate such as a microelectrode array. The polymerase adds nucleotides to growing polynucleotides strands that are also attached to the solid substrate. Spatial control of polymerase activity is achieved by changing the rate of nucleotide polymerization at selected locations on the surface of the solid substrate. The rate of polymerization is changed by inhibiting or promoting activity of the polymerase. In some implementations, activation of electrodes in the microelectrode array changes the rate of nucleotide polymerization. Nucleotides are added to the growing polynucleotide strands at areas where the polymerase is active. By varying the locations where the substrate-bound polymerase is active and the species of nucleotide added, a population of polynucleotides with different, arbitrary sequences is synthesized on the surface of the solid substrate.

De novo polynucleotide synthesis with substrate-bound polymerase

Multiplex similarity search can be performed in a DNA data storage context. The described technologies can support a plurality of different DNA data storage queries in a single query run. A linking strand can be used to connect a query to its matching data element. After the query finds a matching data element, a result strand can be sequenced to the reveal the matching data element as well as which of the queries resulted in the match. Thus, in a multiplex similarity search scenario, a plurality of result strands from a single query run can be correlated to a plurality of different queries. Also, the result strand can be of significantly longer length than both the unmatched data strands and the unmatched query strands. Therefore, filtering based on length can provide more accurate results.

Multiplex similarity search in DNA data storage

A method for DNA synthesis using protected nucleosides is disclosed. The nucleosides may be nucleoside triphosphates or nucleoside phosphoramidites with nucleobases attached to electrochemically-cleavable linkers. Removal of a protecting group by application of a voltage in solution triggers a cyclization reaction that cleaves the electrochemically-cleavable linkers. The electrochemically-cleavable linkers may include an amide linkage and an amide that forms a lactam or an ester linkage and a protected alcohol that forms a lactone when the protecting group is removed. The voltage used to cleave the electrochemically-cleavable linkers may be generated by activation of individual electrodes on a microelectrode array. The microelectrode array can be a substrate for solid-phase synthesis of oligonucleotides. Activation of specific electrodes removes the protecting groups at those electrodes and thus enables spatially-controlled extension of the oligonucleotides. Protected nucleosides linked to protecting groups by electrochemically-cleavable linkers are also disclosed.

Regulation of DNA synthesis by nucleotides linked to protecting groups

A data storage medium is disclosed comprising a substrate covered with alternating layers of a polycationic molecule and artificially synthesized DNA molecules encoding digital information. The magnetic substrate may be a metallic nanoparticle formed from a metal such as iron or cobalt. The polycationic molecule may be polyethyleneimine (PEI). The DNA is protected from degradation by encapsulation in silica. A process for stably storing DNA is also disclosed. Stored DNA may be freed from the silica for sequencing or other analysis by washing the silica-coated DNA with a buffered hydrogen fluoride solution. Storage densities of more than 7% DNA by weight are achieved on nanoparticles.

Silica encapsulated DNA on magnetic nanoparticles

This disclosure describes an efficient method to copy all polynucleotides encoding digital data of digital files in a polynucleotide storage container while maintaining random access capabilities over a collection of files or data items in the container. The disclosure further describes a process whereby random-access and sequencing of the polynucleotides are combined in a single step.

Whole pool amplification and in-sequencer random-access of data encoded by polynucleotides

Multiple polynucleotides with random sequences are collectively used as a molecular anti-counterfeiting tag. The polynucleotides are placed on an item as a molecular identifier of authenticity. Each position within the random sequences is synthesized using a predetermined ratio of nucleoside bases. With this technique the sequence of each polynucleotide is random but the ratio of nucleoside bases over the collection of synthetic polynucleotides is not. Verification of authenticity is achieved by sequencing a portion of the polynucleotides collected from the item and calculating the ratio of nucleoside bases at each position. If these ratios are the same or similar to the ratios used for synthesizing the polynucleotides, then the item is identified as authentic. The ratios of nucleoside bases and a description of the item may be stored in an electronic record that is used for validating authenticity of the item.

Anti-counterfeit tags using base ratios of polynucleotides

Molecular anti-counterfeiting taggants are made from a plurality of synthetic polynucleotides that collectively encode a bit sequence using the sequences and hybridization states of the polynucleotides. The polynucleotide taggant is placed on an item as a molecular identifier of authenticity. The bit sequence encoded by the polynucleotide taggant is read out using a substrate which has bound polynucleotides complexes that hybridize with the synthetic polynucleotides in the polynucleotide taggant. A detectable signal is present where hybridization occurs. To prevent a bad actor from reverse engineering and creating a copy of the polynucleotide taggant using the results of hybridization to the substrate, multiple versions of the substrate are created. Each version hybridizes to different subsets of the synthetic polynucleotides in the polynucleotide taggant. A detectable pattern on the substrate that is present when exposed to the polynucleotide taggant is used for validating authenticity of the item.

Anti-counterfeit polynucleotide taggants

This disclosure provides electrochemically-cleavable linkers with cleavage potentials that are less than the redox potential of the solvent in which the linkers are used. In some applications, the solvent may be water or an aqueous buffer solution. The linkers may be used to link a nucleotide to a bound group. The linkers include a cleavable group which may be one of a methoxybenzyl alcohol, an ester, a propargyl thioether, or a trichloroethyl ether. The linkers may be cleaved in solvent by generating an electrode potential that is less than the redox potential of the solvent. In some implementations, an electrode array may be used to generate localized electrode potentials which selectively cleave linkers bound to the activated electrode. Uses for the linkers include attachment of blocking groups to nucleotides in enzymatic oligonucleotide synthesis.

Electrochemically-cleavable linkers

Redundancy information can be included in nucleotide symbol strings encoding underlying data. To avoid propagation of errors during the decoding process, during encoding, a constrained encoding can be performed before the redundancy information is computed. The redundancy information can be an outer encoding across multiple nucleotide symbol strings. An inner coding within nucleotide symbol strings can also be supported. Such redundancy information can be interleaved into the underlying nucleotide symbol strings to which the constrained encoding has been applied, resulting in a relaxed constraint. Insertion/deletion redundancy information can also be included in the resulting strings, and an insertion/deletion-sensitive sequence can be included to assist in recovering accurate sequences during decoding operations.

Reverse concatenation of error-correcting codes in DNA data storage

Selectively controllable cleavable linkers include electrochemically-cleavable linkers, photolabile linkers, thermolabile linkers, chemically-labile linkers, and enzymatically-cleavable linkers. Selective cleavage of individual linkers may be controlled by changing local conditions. Local conditions may be changed by activating electrodes in proximity to the linkers, exposing the linkers to light, heating the linkers, or applying chemicals. Selective cleaving of enzymatically-cleavable linkers may be controlled by designing the sequences of different sets of the individual linkers to respond to different enzymes. Cleavable linkers may be used to attach polymers to a solid substrate. Selective cleavage of the linkers enables release of specific polymers from the solid substrate. Cleavable linkers may also be used to attach protecting groups to the ends of growing polymers. The protecting groups may be selectively removed by cleavage of the linkers to enable growth of specific polymers.

Selectively controllable cleavable linkers

Electrode controlled hybridization is used to change local pH and selectively assemble oligonucleotide complexes on the surface of a microelectrode array. The oligonucleotide complexes have sticky ends that provide locations for subsequent oligonucleotide complexes to hybridize. The order in which specific oligonucleotide complexes are joined together encodes information. Controlled activation of individual electrodes in the microelectrode array creates negative voltages that reduces a buffer solution and raises the pH in proximity to the electrodes. At higher pH levels double-stranded oligonucleotides de-hybridize. Nicks between oligonucleotide complexes and oligonucleotides anchored to the microelectrode array are closed creating covalent attachments. De-hybridized single-stranded oligonucleotides are removed leaving only the oligonucleotides connected to microelectrode array. Thus, during a given round of synthesis, oligonucleotide complexes are added only to the locations on the microelectrode array where the electrodes are not activated.

Oligonucleotide assembly using pH based electrode controlled hybridization

Polymers synthesized by solid-phase synthesis are selectively released from a solid support by reversing the bias of spatially addressable electrodes. Change in the current and voltage direction at one or more of the spatially addressable electrodes changes the ionic environment which triggers cleavage of linkers that leads to release of the attached polymers. The spatially addressable electrodes may be implemented as CMOS inverters embedded in an integrated circuit (IC). The IC may contain an array of many thousands of spatially addressable electrodes. Control circuity may independently reverse the bias on any of the individual electrodes in the array. This provides fine-grained control of which polymers are released from the solid support. Examples of polymers that may be synthesized on this type of array include oligonucleotides and peptides.

Reversing bias in polymer synthesis electrode array

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Seattle, WA, United States of America

Karin Strauss