Modification of ubiquitin binding peptidase genes in plants for yield trait improvement

Patent metadata

Field	Value
Publication number	US-12359215-B2
Application number	US-202217822822-A
Country	US
Kind code	B2
Filing date	Aug 29, 2022
Priority date	Aug 30, 2021
Publication date	Jul 15, 2025
Grant date	Jul 15, 2025

Abstract

This invention relates to compositions and methods for modifying Ubiquitin Binding Peptidase (DA1) genes in plants, optionally to improve yield traits. The invention further relates to plants having increased improved yield traits produced using the methods and compositions of the invention.

First claim

That which is claimed is:

1. A soybean plant or part thereof comprising at least one non-natural mutation in an endogenous Ubiquitin Binding Peptidase (DA1) gene encoding a ubiquitin binding peptidase (DA1) polypeptide, wherein the endogenous DA1 gene comprises a nucleotide sequence having at least 80% sequence identity to SEQ ID NO:109, wherein the at least one non-natural mutation comprises a mutation in a region of the endogenous DA1 gene having at least 90% sequence identity to a nucleotide sequence of any one of SEQ ID NOs:112-120 or 127-135, and
wherein the soybean plant comprising the at least one non-natural mutation has a phenotype of one or more improved yield traits as compared to a soybean plant devoid of the at least one non-natural mutation.

2. The soybean plant or part thereof of claim 1 , wherein the endogenous DA1 gene (a) comprises a nucleotide sequence having at least 80% sequence identity to SEQ ID NO:110; (b) comprises a region having at least 90% sequence identity to a nucleotide sequence of any one of SEQ ID NOs:112-139; (c) encodes the DA1 polypeptide having at least 80% sequence identity to SEQ ID NO:111; and/or (d) encodes a region having at least 90% sequence identity to any one of SEQ ID NOs:140-146.

3. The soybean plant or part thereof of claim 1 , wherein the at least one non-natural mutation in the endogenous DA1 gene results in a mutation of one or more amino acid residue(s) located in a region of the encoded DA1 polypeptide having at least 90% sequence identity to any one of SEQ ID NOs:140-142.

4. The soybean plant or part thereof of claim 1 , wherein the at least one non-natural mutation results in a substitution, insertion, or deletion of one or more amino acid residue(s) located in a region having at least 90% sequence identity to any one of SEQ ID NOs:140-142.

5. The soybean plant or part thereof of claim 1 , further comprising a substitution of an amino acid residue located at position 379 with reference to amino acid position numbering of SEQ ID NO:111.

6. The soybean plant or part thereof of claim 1 , wherein the at least one non-natural mutation results in a mutated DA1 gene in the soybean plant or part thereof, the mutated DA1 gene having at least 90% sequence identity to any one of SEQ ID NOs:154-156, or 158.

7. The soybean plant or part thereof of claim 1 , wherein the at least one non-natural mutation results in a modified DA1 polypeptide in the soybean plant or part thereof, the modified DA1 polypeptide comprising a mutation in an amino acid residue located in a region of a DA1 polypeptide comprising an amino acid sequence having at least 90% sequence identity to any one of SEQ ID NOs:140-142.

8. A soybean plant cell comprising at least one non-natural mutation within an endogenous Ubiquitin Binding Peptidase (DA1) gene, wherein the endogenous DA1 gene comprises a nucleotide sequence having at least 80% sequence identity to SEQ ID NO:109, wherein the at least one non-natural mutation is a substitution, insertion, or deletion that is introduced using an editing system that comprises a nucleic acid binding domain that binds to a target site in the endogenous DA1 gene, and wherein the at least one non-natural mutation comprises a mutation in a region of the endogenous DA1 gene having at least 90% sequence identity to a nucleotide sequence of any one of SEQ ID NOs:112-120 or 127-135.

9. The soybean plant cell of claim 8 , wherein the target site is within a region of the endogenous DA1 gene, the region encoding an amino acid sequence having at least 90% sequence identity to any one of SEQ ID NOs:140-142.

10. The soybean plant cell of claim 9 , wherein the editing system further comprises a nuclease, wherein the nucleic acid binding domain binds to the target site in a sequence having least 80% sequence identity to any one of SEQ ID NOs:109 or 110 and/or in a sequence having at least 90% sequence identity to the nucleotide sequence of any one of SEQ ID NOs:112-120 or 127-135, and wherein the at least one non-natural mutation within the DA1 gene is made following cleavage by the nuclease.

11. The soybean plant cell of claim 8 , wherein the at least one non-natural mutation within the DA1 gene is an insertion and/or a deletion.

12. The soybean plant cell of claim 8 , further comprising a substitution of an amino acid residue located: at position 379 with reference to amino acid position numbering of SEQ ID NO:111.

13. The soybean plant cell of claim 8 , wherein the at least one non-natural mutation results in a mutated DA1 gene comprising a nucleotide sequence having at least 90% sequence identity to any one of SEQ ID NOs:154-156 or 158.

14. A method for producing a soybean plant or part thereof comprising at least one cell having a mutated endogenous Ubiquitin Binding Peptidase (DA1) gene, the method comprising;
contacting a target site in an endogenous DA1 gene in the soybean plant or part thereof with a nuclease comprising a cleavage domain and a nucleic acid binding domain, wherein the nucleic acid binding domain binds to a target site in the endogenous DA1 gene, wherein the endogenous DA1 gene comprises a nucleotide sequence having at least 80% sequence identity to SEQ ID NO: 109, thereby producing the soybean plant or part thereof comprising the at least one cell having the mutated endogenous DA1 gene;
and
wherein the mutated endogenous DA1 gene comprises a non-natural mutation in a region of the endogenous DA1 gene having at least 90% sequence identity to a nucleotide sequence of any one of SEQ ID NOs:112-120 or 127-135.

15. The method of claim 14 , wherein the non-natural mutation is a substitution, an insertion, and/or a deletion.

16. The method of claim 14 , wherein the non-natural mutation is an insertion and/or a deletion that results in an amino acid substitution and/or a premature stop codon.

17. The method of claim 14 , wherein the non-natural mutation results in a modified amino acid residue located in a region having at least 90% sequence identity to any one of SEQ ID NOs:140-142.

18. The method of claim 14 , wherein the mutated endogenous DA1 gene further comprises a substitution at position 379 with reference to amino acid position numbering of SEQ ID NO:111.

19. A guide nucleic acid that binds to a target site in a Ubiquitin Binding Peptidase (DA1) gene, wherein the target site is in a region of the DA1 gene having at least 90% sequence identity to any one of SEQ ID NOs:112-120 or 127-135.

20. The soybean plant or part thereof of claim 1 , wherein the at least one non-natural mutation is a deletion.

Assignees

• Pairwise Plants Services Inc

Inventors

• Mathew Lolita George
• Zhang Xiaoyu

Classifications

• C12N15/8213

  Targeted insertion of genes into the plant genome by homologous recombination · CPC title

• C12N15/11

  DNA or RNA fragments; Modified forms thereof (DNA or RNA not used in recombinant technology, C07H21/00); {Non-coding nucleic acids having a biological activity} · CPC title

• C12N2310/20

  involving clustered regularly interspaced short palindromic repeats [CRISPR] · CPC title

• C12N9/63

  derived from plants · CPC title

• Y02A40/146

  Genetically Modified [GMO] plants, e.g. transgenic plants · CPC title