# US 9,434,953 B2 · Methods and compositions for modified ethanol inducible promoter systems

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- Number: US 9,434,953 B2
- Document ID: US9434953B2
- Status: Granted
- Kind: B2
- Application number: 13/978,133
- Filed: 2011-12-15
- Published: 2013-10-31
- Granted: 2016-09-06
- Earliest claimed: 2011-01-18
- Assignees: Syngenta Participations AG; Queensland University of Technology
- Applicants: Manuel Benito Sainz; Mark David Kinkema
- Inventors: Manuel Benito Sainz; Mark David Kinkema
- Web page: https://patentgrep.com/patent/US9434953B2

## CPC

- C12N 15/8238: {chemically inducible, e.g. tetracycline}
- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- C12N 15/8241: {Phenotypically and genetically modified plants via recombinant DNA technology}
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

The present invention provides nucleic acid molecules comprising one or more nucleotide sequences selected from the group consisting of the nucleotide sequence of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, and/or SEQ ID NO:10 that can be operably linked to a promoter, thereby making the promoter inducible by a chemical compound that can induce the expression of the alcohol dehydrogenase system of Aspergillus nidulans. Methods for making an inducible promoter and for making the expression of a nucleotide sequence of interest inducible are also provided. Further provided are plants, plant parts, and plant cells, comprising the compositions of the present invention.

## Claims

### Claim 1

An isolated nucleic acid molecule comprising five to nine nucleotide sequences selected from the group consisting of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 10, wherein the selected nucleotide sequences can be in any combination, in any orientation, and/or in any order, and wherein the five to nine nucleotide sequences are operably linked to a heterologous promoter.

### Claim 2 (depends on 1)

The isolated nucleic acid molecule of claim 1, wherein the heterologous promoter is further operably linked to a nucleotide sequence of interest.

### Claim 3

A method of making a promoter inducible, comprising:
a) constructing a nucleic acid molecule comprising five to nine nucleotide sequences selected from the group consisting of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 10, in any combination, in any orientation, and/or in any order; and
b) operably linking the nucleic acid molecule of (a) with the promoter, wherein the promoter is heterologous to the five to nine nucleotide sequences of the nucleic acid molecule.

### Claim 4 (depends on 3)

The method of claim 3, wherein the heterologous promoter is made inducible to a chemical compound selected from the group consisting of a primary alcohol, a primary monoamine, a ketone, a methyl ketone, a hydrolysable ester, an aliphatic aldehyde, and any combination thereof.

### Claim 5 (depends on 3)

The method of claim 3, wherein the heterologous promoter is made inducible to a chemical compound selected from the group consisting of ethanol, acetaldehyde, ethyl methyl ketone, acetone, ethylamine, cyclohexanone, butan-2-ol, 3-oxobutyric acid, propan-2-ol, propan-1-ol, butan-2-ol, threonine, and any combination thereof.

### Claim 6 (depends on 3)

The method of claim 3, wherein the heterologous promoter that is made inducible is a constitutive promoter, a tissue specific promoter, a developmentally regulated promoter, and/or a chemically inducible promoter that is inducible by compounds that do not induce the expression of the alcohol dehydrogenase system of Aspergillus nidulans.

### Claim 7 (depends on 3)

The method of claim 3, wherein the heterologous promoter is a minimal promoter.

### Claim 8 (depends on 3)

The method of claim 3, wherein the five to nine nucleotide sequences are at the 5′ end of the nucleic acid molecule.

### Claim 9 (depends on 1)

A method of making the expression of a nucleotide sequence of interest inducible comprising operably linking the heterologous promoter of claim 1 to the nucleotide sequence of interest.

### Claim 10 (depends on 3)

A promoter made by the method of claim 3.

### Claim 11 (depends on 2)

A plant, plant part or plant cell comprising the isolated nucleic acid molecule of claim 2.