# US 2005/0097640 A1 · Methods for enhancing stress tolerance in plants and compositions thereof

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- Number: US 2005/0097640 A1
- Document ID: US20050097640A1
- Status: Published application
- Kind: A1
- Application number: 10/953,856
- Filed: 2004-09-29
- Published: 2005-05-05
- Earliest claimed: 2003-09-29
- Assignees: Monsanto Technology LLC
- Inventors: Mary Fernandes
- Web page: https://patentgrep.com/patent/US20050097640A1

## CPC

- C12N 15/8273: {for drought, cold, salt resistance}
- C12N 15/8261: {with agronomic (input) traits, e.g. crop yield}
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Increased tolerance to abiotic stress in a plant is provided by introducing DNA expressing a cold shock protein, e.g. bacterial cold shock protein.

## Claims

### Claim 1

A recombinant DNA molecule that comprises, in the 5′ to 3′ direction:
a) a first DNA polynucleotide comprising a promoter that functions in plants and which is operably linked to;
b) a second DNA polynucleotide that encodes a cold shock protein, operably linked to;
c) a 3′ transcription termination DNA polynucleotide that functions as a polydenylation sequence.

### Claim 2 (depends on 1)

A recombinant DNA molecule of claim 1 wherein a plant DNA intron is inserted between said first DNA polynucleotide and said second DNA polynucleotide.

### Claim 3 (depends on 1)

A recombinant DNA molecule of claim 1 wherein said second DNA polynucleotide encodes a protein comprising the amino acid motif of SEQ ID NO: 3.

### Claim 4 (depends on 3)

A recombinant DNA molecule of claim 3 wherein said second DNA polynucleotide encodes a protein selected from the group consisting of
(a) A protein with an amino acid sequence of substantial identity to an amino acid sequence of a cold shock protein from gram positive bacteria,
(b) a cold shock protein from Bacillus subtilis,
(c) a homologue of Bacillus subtilis cold shock protein B (CspB),
(d) a protein with an amino acid sequence of substantial identity to SEQ ID NO: 2,
(e) a protein with an amino acid sequence of substantial identity to an amino acid sequence of a cold shock protein from a gram negative bacteria,
(f) a protein comprising a cold shock protein from Escherichia coli
(g) a homologue of Escherichia coli cold shock protein A (CspA),
(h) a protein with an amino acid sequence that has substantial identity to SEQ ID NO: 1,
(i) a cold shock protein from Agrobacterium tumefaciens, and
(j) a protein having an amino acid sequence with substantial identity to any of SEQ ID NO: 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, or 65.

### Claim 5 (depends on 1)

A recombinant DNA molecule according to claim 1, wherein the promoter is selected from the group consisting of inducible promoters, constitutive promoters, temporal-regulated promoters, developmentally-regulated promoters, tissue-preferred promoters, cold enhanced promoters, cold-specific promoters, stress enhanced promoters, stress specific promoters, drought inducible promoters, water deficit inducible promoters, and tissue-specific promoters.

### Claim 6

An abiotic stress-tolerant, transgenic plant that has been transformed with a DNA molecule that expresses a cold shock protein.

### Claim 7 (depends on 6)

A propagule of a plant of claim 6.

### Claim 8 (depends on 6)

Progeny of a plant of claim 6.

### Claim 9 (depends on 6)

A plant of claim 6 which is a crop plant.

### Claim 10 (depends on 6)

A plant of claim 6 which is a monocot plant.

### Claim 11 (depends on 6)

A plant of claim 6 which is a dicot plant.

### Claim 12 (depends on 6)

A plant of claim 6 which is a plant is selected from the group consisting of soybean, corn, canola, rice, cotton, barley, oats, turf grass, cotton, and wheat.

### Claim 13 (depends on 6)

A plant of claim 6 that has
(a) a higher growth rate under conditions where cold temperature would be limiting for growth for a non-transformed plant of the same species,
(b) a higher growth rate under conditions where high temperature would be limiting for growth for a non-transformed plant of the same species,
(c) a higher growth rate under conditions where water would be limiting for growth for a non-transformed plant of the same species,
(d) a higher growth rate under conditions where increased salts or ions in the soil and/or water would be limiting for growth of a non-transformed plant of the same species,
(e) has a greater percentage of plants surviving after a cold shock than a non-transformed plant of the same species,
(f) an increased yield when compared to a non-transformed plant of the same species, or
(g) resistance to drought compared to a non-transformed plant of the same species.

### Claim 14 (depends on 13)

A propagule of a plant of claim 13 which is a seed.

### Claim 15 (depends on 14)

A method wherein a seed of claim 14 is planted in soil and allowed to grow.

### Claim 16 (depends on 1)

A method of producing a transgenic plant comprising the steps of:
a) inserting into the genome of plant cells a recombinant DNA molecule of claim 1:
b) obtaining transformed plant cell containing said recombinant DNA;
c) regenerating plants from said plant cells; and
d) selecting a plant for increased abiotic stress tolerance or increased root growth.

### Claim 17 (depends on 16)

A stress tolerant plant produced by the method of claim 16.

### Claim 18 (depends on 16)

A method of claim 16 wherein said abiotic stress is selected from the group consisting of heat tolerance, salt tolerance, drought tolerance, and survival after cold shock.

### Claim 19

An isolated protein which
(a) is at least 40% identical to at least one protein selected from the group consisting of SEQ ID NOS: 5, 7, 9, 29, 31, 33, 35, 37, 39, 41, 43, 53, 55, 57, 59, 61, 63, and 65,
(b) hybridizes under stringent conditions to a nucleic acid selected from the group consisting of SEQ ID NOs: 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 90, and 92.
(c) has an amino acid sequence which is substantially identical to any of SEQ ID NOS: 5, 7, 9, 29, 31, 33, 35, 37, 39, 41, 43, 53, 55, 57, 59, 61, 63, and 65.

### Claim 20

A field crop comprising at least 50% of plants germinated from a propagule comprising a prokaryotic cold shock protein.