# US 2023/0265448 A1 · Methods and compositions for the introduction and regulated expression of genes in plants

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- Number: US 2023/0265448 A1
- Document ID: US20230265448A1
- Status: Published application
- Kind: A1
- Application number: 18/047,714
- Filed: 2022-10-19
- Published: 2023-08-24
- Earliest claimed: 2009-12-30
- Assignees: E. I. du Pont de Nemous and Company; Pioneer Hi-Bred International, Inc.
- Applicants: E. I. Du Pont De Nemours and Company; Pioneer Hi-Bred International, Inc.
- Inventors: WILLIAM JAMES GORDON-KAMM; THEODORE M KLEIN; KEITH S LOWE; KEVIN E MCBRIDE; CHRISTOPHER J SCELONGE; NING WANG
- Web page: https://patentgrep.com/patent/US20230265448A1

## CPC

- C12N 15/8241: {Phenotypically and genetically modified plants via recombinant DNA technology}
- C12N 15/8213: {Targeted insertion of genes into the plant genome by homologous recombination}
- C12N 15/8216: {Methods for controlling, regulating or enhancing expression of transgenes in plant cells}
- C12N 15/8262: {involving plant development}
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Compositions and methods are provided for the introduction and the regulated expression of genes in plants. Compositions include promoter constructs that provide a level of activity useful for the regulated expression of site-specific recombinases, while avoiding premature excision. Further provided are isolated polynucleotides encoding novel babyboom polypeptides, expression cassettes, and plants comprising the same. Methods for the introduction of genes into plants are provided, including methods for plastid transformation and methods for the transformation of tissues from mature seeds and leaves.

## Claims

### Claim at position 1

1-50. (canceled)

### Claim 51

A method for introducing a polynucleotide of interest into a monocot leaf explant, said method comprising:
a) providing a monocot leaf tissue explant;
b) introducing into said monocot leaf explant:
i) a heterologous polynucleotide encoding a babyboom cell proliferation factor and a heterologous polynucleotide encoding a Wuschel cell proliferation factor; and expressing said heterologous polynucleotide encoding said babyboom cell proliferation factor and said heterologous polynucleotide encoding said Wuschel cell proliferation factor; and
ii) a polynucleotide of interest; and
c) regenerating a monocot plant comprising the polynucleotide of interest.

### Claim 52 (depends on 51)

The method of claim 51, further comprising excising said heterologous polynucleotide encoding said babyboom cell proliferation factor and said heterologous polynucleotide encoding said Wuschel cell proliferation factor.

### Claim 53 (depends on 51)

The method of claim 51, wherein the monocot is selected from the group consisting of maize, sorghum, sugarcane, rice, and wheat.

### Claim 54

A method for introducing a polynucleotide of interest into a monocot leaf tissue explant, said method comprising:
a) excising a leaf segment from a leaf above the first leaf base node;
b) dissecting said leaf segment into a leaf tissue explant;
c) introducing into said leaf tissue explant:
i) a heterologous polynucleotide encoding a babyboom cell proliferation factor and a heterologous polynucleotide encoding a Wuschel cell proliferation factor; and expressing said heterologous polynucleotide encoding said babyboom cell proliferation factor and said heterologous polynucleotide encoding said Wuschel cell proliferation factor; and
ii) a polynucleotide of interest; and
d) regenerating a monocot plant comprising the polynucleotide of interest.

### Claim 55 (depends on 54)

The method of claim 54, further comprising excising said heterologous polynucleotide encoding said babyboom cell proliferation factor and said heterologous polynucleotide encoding said Wuschel cell proliferation factor.

### Claim 56 (depends on 54)

The method of claim 54, wherein the monocot is selected from the group consisting of maize, sorghum, sugarcane, rice, and wheat.