Agilent Technologies, Inc.

Hewlett-Packard Company

University of Colorado

Leland Stanford Junior University

University and Agilent Technologies, Inc.

Joel Myerson

Douglas J Dellinger

Geraldine F Dellinger

Zoltan Timar

Jeffrey Robert Sampson

Marvin H Caruthers

Agnieszka Sierzchala

Daniel E Ryan

Robert Kaiser

Brian Jon Peter

Michel G M Perbost

Matthew Hebden Porteus

Zohar H Yakhini

Brian Phillip Smart

Rasmus O Bak

Ayal Hendel

Joe Clark

Nicholas Michael Sampas

Peter G Webb

Karla M Robotti

Zoltan Kupihar

Paige Anderson

John Turner

Fernando Ferreira

Anna M Tsalenko

Daniel Bernardo Roitman

Siyuan Chen

Bo U Curry

Holly Hogrefe

Arthur Schleifer

Reid Alyn Brennen

Robert A Ach

Rolfe C Anderson

May Tom-Moy

Javier E Satulovsky

Michael Thomas Barrett

Andrew Stefanski

Geraldine Fulcrand

Diane D Ilsley

John Mannion

H Bryan Bente

Philip W Barth

Paul F Bente, Iii

Michel G M Perobost

Craig Daniel Wenger

Alan C Herman

Benjamin G Shaw

Agnieska Sierzchala

Geraldine F Dellingen

Provided herein are methods for inducing CRISPR/Cas-based gene regulation (e.g., genome editing or gene expression) of a target nucleic acid (e.g., target DNA or target RNA) in a cell. The methods include using modified single guide RNAs (sgRNAs) that enhance gene regulation of the target nucleic acid in a primary cell for use in ex vivo therapy or in a cell in a subject for use in in vivo therapy. Additionally, provided herein are methods for preventing or treating a genetic disease in a subject by administering a sufficient amount of a modified sgRNA to correct a mutation in a target gene associated with the genetic disease.

Compositions comprising chemically modified guide RNAs for CRISPR/Cas-mediated editing of HBB

Compounds and methods for purifying oligonucleotides such as RNA and DNA. A target oligonucleotide is reacted with an orthoester linker comprising an affinity tag to form an orthoester oligonucleotide-orthoester linker conjugate which is subjected to a purification technique to separate the target oligonucleotide from impurities such as truncated oligonucleotides. The orthoester linker can be then removed under mild conditions to generate the target oligonucleotide in high purity.

Orthoester compositions for affinity purification of oligonucleotides

Chemically modified guide RNAS for CRISPR/Cas-mediated gene regulation

Chemically modified guide RNAs for CRISPR/CAS-mediated gene regulation

The present invention relates to modified guide RNAs and their use in clustered, regularly interspaced, short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems.

Guide RNA with chemical modifications

Described herein, among other things, is a method of estimating efficiency of an oligonucleotide synthesis reaction. In some embodiments, the method comprises subjecting the products of one or more oligonucleotide synthesis reactions to LC-MS to produce a series of mass spectra, analyzing the mass spectra, and estimating the overall efficiency of an oligonucleotide synthesis reaction and/or the efficiency of addition of one or more of G, A, T or C individually in an oligonucleotide synthesis reaction.

Method of oligonucleotide analysis

One or more liquids are transferred from a source array to one or more remotely positioned destination sites such as chambers by utilizing one or more movable transfer elements, such as contact pins or capillaries. The source array may include a predetermined organization of addresses at which materials are positioned. One or more materials may be selected for transfer. Based on the selection, one or more addresses may be accessed by the transfer element(s). The addresses may correspond to spots on a surface of the source array. Each spot may be a feature containing one or more (bio)chemical compounds. At the chamber(s), the material(s) may be processed, such by reaction with one or more reagents. The reaction(s) may entail synthesis of one or more desired products. Alternatively, reaction(s) may be performed at the source array, and the product(s) then transferred to the chamber(s).

Systems and methods for transferring liquids

A method of deprotecting a solid support bound polynucleotide comprising at least one 2'-protected ribonucleotide in which a step of contacting the polynucleotide with a composition comprising a diamine is performed under conditions sufficient to deprotect and cleave the polynucleotide which remains retained on the solid support.

Protected monomer and method of final deprotection for RNA synthesis

A method for sequencing a nucleic acid is provided. In certain embodiments the method comprises obtaining a duplex comprising a nucleic acid and a primer, wherein the primer has a nuclease resistant 3' end, combining the duplex with a chain terminator nucleotide and a proof-reading polymerase to produce a reaction in which the polymerase idles on the added chain terminator nucleotide, identifying the chain terminator nucleotide added to the end of the primer; and adding a nuclease-resistant nucleotide to the end of the primer after the polymerase has idled on and removed the added chain terminator nucleotide, thereby producing a duplex comprising the template and an extended primer that has a nuclease resistant 3&#x2032; end.

Polymerase idling method for single molecule DNA sequencing

This disclosure provides a method comprising: a) clamping the top and bottom strands of a double stranded DNA molecule to produce a duplex in which the top and bottom strands are linked; b) denaturing the duplex to produce a denatured product; and c) renaturing the denatured product in the presence of a labeled oligonucleotide that is complementary to a sequence of nucleotides in the double stranded DNA molecule, thereby producing a D-loop-containing product. Kits for performing the method and products made by the method are also provided.

Target enrichment and labeling for multi-kilobase DNA

Aspects of the present disclosure include methods for double coupling a nucleoside phosphoramidite during synthesis of an oligonucleotide. The method can include coupling a free hydroxyl group of a nucleoside residue with a first sample of a protected nucleoside phosphoramidite via an internucleoside P(III) linkage, followed by exposure to an oxidizing agent prior to a second coupling step with a second sample of the protected nucleoside phosphoramidite, and further exposure to an oxidizing agent. The method finds use in synthesizing an oligonucleotide on a solid phase support, such as a planar surface. The double coupling method can be utilized at one or more nucleotide positions during oligonucleotide synthesis thereby reducing single base deletion rates. Oligonucleotide containing compositions synthesized according to the disclosed methods are also provided.

Double coupling method for oligonucleotide synthesis

A method of deprotecting a solid support bound polynucleotide includes the step of contacting the polynucleotide with a composition comprising a diamine under conditions sufficient to deprotect the 2'-protected ribonucleotide residue. The solid support bound polynucleotide has at least one 2&#x2032;-protected ribonucleotide residue, which has the following structure: wherein Bis a protected or unprotected heterocycle; Ris a protecting group selected from a hydrocarbyl, a substituted hydrocarbyl, an aryl, and a substituted aryl; X is O or S; and PG is a thionocarbamate protecting group.

Growing community of inventors

Berkeley, CA, United States of America

Joel Myerson