Pioneer Hi-bred International, Inc.

E.i. Dupont De Nemours and Company

Dupont Nutrition Biosciences Aps

Vilnius University

Universite Laval (canada)

Corteva Agriscience LLC

Joshua K Young

Phillip A Patten

Cathy Xiaoyan Zhong

Andrew Mark Cigan

Virginijus Siksnys

Christopher Jay Scelonge

Zhenglin Hou

Huirong Gao

Sergei Svitashev

James C Register, Iii

Erin L Crowgey

M Alejandra Pascual

Tautvydas Karvelis

Albert Laurence Lu

Saverio Carl Falco

Scott H Diehn

James J English

Timothy M Nowatzki

Douglas S Nubel

Lu Liu

Ericka Bermudez

Zhan-Bin Liu

Azalea S Ong

William James Gordon-Kamm

Mark Jacob Gadlage

Keith S Lowe

Zhongsen Li

David J Peterson

David Hondred

Robert B Meeley

Kent Brink

Matthew Curtis Harmon

Stephane Deschamps

Giedrius Gasiunas

Clara Maria Alarcon

Nina Dietrich

Stephen Lawrence Gasior

Sandeep Kumar

Christophe Fremaux

Philippe Horvath

Jinrui Shi

Dennis A Romero

Zhan-bin Liu

Douglas Stuart Nubel

Victor Llaca

Alexander Hynes

Geneviève Rousseau

L Aleksander Lyznik

Marie-Laurence Lemay

Mindaugas Zaremba

M Alejandra Pascual Villanelo

Iii James Calvin Register

Maria Alejandra Pascual

James C Iii Register

Clara M Alarcon

M Alejandra Villanelo

James Calvin Register

James C Register

Compositions and methods are provided for genome modification of a target sequence in the genome of a plant or plant cell. The methods and compositions employ a guide RNA/Cas endonuclease system to provide an effective system for modifying or altering target sites within the genome of a plant, plant cell or seed. Also provided are compositions and methods employing a guide polynucleotide/Cas endonuclease system for genome modification of a nucleotide sequence in the genome of a cell or organism, for gene editing, and/or for inserting or deleting a polynucleotide of interest into or from the genome of a cell or organism. Once a genomic target site is identified, a variety of methods can be employed to further modify the target sites such that they contain a variety of polynucleotides of interest. Breeding methods and methods for selecting plants utilizing a two component RNA guide and Cas endonuclease system are also disclosed. Compositions and methods are also provided for editing a nucleotide sequence in the genome of a cell.

Plant genome modification using guide RNA/Cas endonuclease systems and methods of use

Compositions and methods are provided for genome modification of a target sequence in the genome of a cell, using a novel Cas endonuclease. The methods and compositions employ a guide polynucleotide/endonuclease system to provide an effective system for modifying or altering target sequences within the genome of a cell or organism. Also provided are novel effectors and endonuclease systems and elements comprising such systems, such as guide polynucleotide/endonuclease systems comprising an endonuclease. Compositions and methods are also provided for guide polynucleotide/endonuclease systems comprising at least one endonuclease, optionally covalently or non-covalently linked to, or assembled with, at least one additional protein subunit, and for compositions and methods for direct delivery of endonucleases as ribonucleotide proteins.

CRISPR-Cas systems for genome editing

Methods and compositions are provided for the improvement of double-strand-break-inducing agent activity in eukaryotic cells, through the usage of one or more morphogenic factors or developmental genes. The morphogenic factor may be provided to the same cell or to a different cell than that comprising or receiving the double-strand-break-inducing agent. The morphogenic factor may be provided to a cell as a polynucleotide composition on a recombinant vector, and may be placed on the vector outside of a T-DNA border. The morphogenic factor may be provided via an upregulation of an endogenous gene. The morphogenic factor, or the double-strand-break-inducing agent, may further comprise a cell penetrating peptide. The morphogenic factor may be co-introduced with a vector comprising RepA.

Use of morphogenic factors for the improvement of gene editing

CRISPR-cas systems for genome editing

Novel engineered non-natural deaminases are provided. Methods and compositions are provided for modifying the genome of a cell, including a plant cell. Genome modification through guided CRISPR polypeptides and multiple deaminases are provided. Engineered non-natural novel deaminases, including adenosine deaminases are provided. These novel deaminases have improved activity in cells including plant cells.

Engineered deaminases

Compositions and methods are provided for novel Cas9 orthologs, including, but not limiting to, novel guide polynucleotide/Cas9 endonucleases complexes, single or dual guide RNAs, guide RNA elements, and Cas9 endonucleases. The present disclosure also describes methods for creating a double strand break in a target polynucleotide, methods for genome modification of a target sequence under various in vivo and in vitro conditions, in the genome of a cell, for gene editing, and for inserting a polynucleotide of interest into the genome of a cell. Also provided are nucleic acid constructs and cells having a modified target site or altered polynucleotide of interest produced by the methods described herein.

Cas9 orthologs

Methods and compositions are provided for the identification, detection, characterization, and/or utilization of double strand breaks in a target polynucleotide; the identification, detection, characterization, and/or utilization of cutting sites for double-strand-break-inducing agents; and the identification, detection, characterization, and/or utilization of double-strand-break-inducing agents.

Methods for the identification and characterization of double-strand break sites and compositions and uses thereof

The present disclosure involves the production of Waxy maize. Compositions and methods are provided for knocking out expression of the Waxy (Wx1) gene in maize by making double strand breaks at one or more target sites in an endogenous WX1 encoding sequence. Some methods employ one or more guide polynucleotides and a Cas endonuclease, wherein Cas endonuclease is guided by the one or more guide polynucleotides to recognize and introduce double strand breaks at specific target sites in and around the Wx1 gene. Also provided are compositions and methods for the production of Waxy maize plant cells, plant explants, seeds and grain.

Waxy corn

Methods and compositions are provided for the use of anti-CRISPR (ACR) proteins in plants, including modulation of Cas endonuclease activity, improvement of frequency of homologous recombination, control of Cas endonuclease activity during various cell cycles, spatial and/or temporal regulation of Cas endonuclease activity in plants, usage in gene activation or repression, as well as reduction of off-target polynucleotide cleavage.

Anti-CRISPR polynucleotides and polypeptides and methods of use

Compositions and methods are provided for rapid characterization of Cas endonuclease systems and the elements comprising such systems, including, but not limiting to, rapid characterization of PAM sequences, guide RNA elements and Cas endonucleases. Type II Cas9 endonuclease systems originating fromMlc3,DSM 15814,SL4,JCM 14932,sp. SCADC,DSM 20210,NX02,DSM 16811,are described herein. The present disclosure also describes methods for genome modification of a target sequence in the genome of a cell, for gene editing, and for inserting a polynucleotide of interest into the genome of a cell.

Rapid characterization of Cas endonuclease systems, PAM sequences and guide RNA elements

The invention provides DNA compositions that relate to transgenic insect resistant maize plants. Also provided are assays for detecting the presence of the maize DP-004114-3 event based on the DNA sequence of the recombinant construct inserted into the maize genome and the DNA sequences flanking the insertion site. Kits and conditions useful in conducting the assays are provided.

Maize event DP-004114-3 and methods for detection thereof

CRISPR-CAS systems for genome editing

Growing community of inventors

Johnston, IA, United States of America

Joshua K Young