Method for inducing heat stress tolerance in plants

Abstract

The present invention is directed to a method for improving heat stress tolerance in plants, comprising applying an effective amount of 1-amino-1-cyclopropanecarboxylic acid to the plants.

Description

Technical Field

[0001] The present invention is directed to a method for improving heat stress tolerance in plants, which comprises applying an effective amount of 1-amino-1-cyclopropanecarboxylic acid to the plants.

Background Art

[0002] Temperatures are rising worldwide and are expected to continue to rise for the remainder of this century. This increase in temperature will cause heat stress in crops such as wheat. See Tack J., et al., Effect of warming temperatures on US wheat yields Proc Natl Acad Sci U S A. 2015 Jun 2;112(22):6931-6. Heat stress is particularly threatening to wheat production when stress occurs during the reproductive and ripening stages. See Ababaei B. and Chenu K., Heat shocks increasingly impede grain filling but have little effect on grain setting across the Australian wheatbelt, Agr. For. Meterol., 2020 Jan; 284, 107889.

[0003] Heat stress is thought to reduce a plant's ability to photosynthesize by limiting the plant's metabolic rate and damaging the chloroplasts. This reduction in photosynthetic ability results in a decrease in grain production. The decrease in grain production is costly to both farmers and the entire population. With the increase in world population, an increase in crop production is needed. Therefore, improving heat stress tolerance in crops such as wheat is beneficial and necessary for the survival of the world population.

[0004] 1-Amino-1-cyclopropanecarboxylic acid ("ACC") is a product of the enzyme ACC synthase and acts as a biosynthetic precursor of ethylene in plants. Ethylene has been shown to be involved in several plant responses such as stress, fruit set, leaf abscission, flowering, and senescence. Because of its role as an ethylene precursor, ACC is used in agriculture to induce ethylene response phenomena. However, ACC has not been shown to induce heat stress tolerance in various plants such as wheat, corn, soybean, cotton, tomato, legumes, lettuce, oilseed rape (e.g., canola, rapeseed, etc.), turfgrass, and ornamental plants. In fact, heat stress-induced ethylene production in developing wheat grains causes kernel abortion and a decrease in yield. See Hays D.B. et al., Heat stress induced ethylene production in developing wheat grains induces kernel abortion and increased maturation in a susceptible cultivar, Plant Sci, 2007, 172, 1113-1123 and Valluru R., et al., Phenotypic and genome-wide association analysis of spike ethylene in diverse wheat genotypes under heat stress, New Phytologist, 2016, doi:10.1111 / nph. 14367, 1-13.

[0005] Therefore, there is a need in the art for methods to improve heat stress tolerance in crops. SUMMARY OF THE INVENTION

[0006] The present invention is a method for improving heat stress tolerance in plants, characterized by applying an effective amount of 1-amino-1-cyclopropanecarboxylic acid ("ACC") to the plants, wherein the plants are selected from the group consisting of wheat, corn, soybean, cotton, tomato, rape, bean, lettuce, turfgrass and ornamental plants.

BEST MODE FOR CARRYING OUT THE INVENTION

[0007] The applicant unexpectedly found that the application of 1-amino-1-cyclopropanecarboxylic acid ("ACC") to plants improves heat stress tolerance and results in an increase in grain production.

[0008] Once planted, plants go through life stages such as the vegetative stage, reproductive stage, maturity stage and senescence stage. Plants are generally subjected to heat stress during the developmental stage, vegetative stage and reproductive stage, especially during the flowering stage. Plants may also be subjected to heat stress during the maturity stage.

[0009] Wheat undergoes several life cycle stages, starting particularly from germination, proceeding through tillering, heading, flowering, and finally maturity. See Miller T.D., Growth Stages of Wheat: Identification and Understanding Improve Crop Management, Texas A&M Agrilife Ext. SCS-1999-16. The Feekes scale was developed to systematically describe the life cycle of the wheat plant. Feekes stage 1 describes the emergence of the wheat plant from the soil. Feekes stages 2-3 describe the tillering stage. The tillering stage is the time when the plant produces axillary or lateral buds known as tillers. The wheat plant begins to produce tillers at Feekes stage 2 and stops producing tillers at Feekes stage 3. Feekes stage 4 describes the start of the upright growth of the wheat plant. Feekes stage 5 occurs after a period of low temperature and determines the number of spikelets per ear. The ear grows from the tip of each tiller, bears multiple spikelets, and each of them sets seeds. Tillers formed after stage 5 do not lead to the harvest of the grain. Between Feekes stages 6-9, nodes occur and leaves appear. Heading and flowering begin at Feekes stage 10. Feekes stage 10 is divided into multiple sub-stages. Feekes stage 10.1 describes the appearance of the tip of the ear. Feekes stage 10.3 describes the appearance of half of the ear. Feekes stage 10.4 describes the appearance of three-quarters of the ear. Feekes stage 10.5 describes the complete appearance of the ear. Flowering begins at Feekes stage 10.5.1. Thereafter, pollination and the maturation of the grain (i.e., the cereal) continue from Feekes stage 10.5.3 to Feekes stage 11.4.

[0010] Throughout this application, the applicant refers to the soybean growth stage as the "V" stage. The "V" stage is designated by numbers such as V1, V2, V3, etc. In this V(n) identification system, (n) represents the number of trifoliate leaves that are open. Each leaf stage is defined by the uppermost leaf that is fully expanded.

[0011] As used herein, the term "heat stress" is defined as the exposure of a plant to an air temperature of 24 °C or higher. In a preferred embodiment, the term "heat stress" may include the exposure of a plant to an air temperature of 25 °C or higher, 26 °C or higher, 27 °C or higher, 28 °C or higher, 29 °C or higher, or 30 °C or higher. Heat stress may occur either during the day or at night. For the same plant, nocturnal heat stress may occur at a lower temperature than diurnal heat stress.

[0012] As used herein, the term "improve" is defined as an increase in the growth and / or yield of a plant exposed to heat stress. Measurements of growth and / or yield include, but are not limited to, measurements taken during the vegetative growth stage such as fresh weight of biomass, total number of tillers, rate of tillering, population density, and total dry weight of biomass, and measurements taken during the reproductive stage such as total number of spikes, weight of spikes, yield per spike, grain weight, and harvest index, but are not limited thereto.

[0013] As used herein, all numerical values relating to amounts, ratios, weight percentages, and the like are defined as "about" each particular value, plus or minus 10%. For example, the phrase "at least 5.0% by weight" should be understood to mean "from at least 4.5% to 5.5% by weight". Accordingly, amounts within 10% of the numerical values in the claims are encompassed by the claims.

[0014] The term "effective amount" means the amount of a formulation that will improve heat stress tolerance. The "effective amount" will vary depending on, among other factors, the type of plant being treated, the severity of the heat stress, the desired result, and the life stage of the plant during treatment. Therefore, it is not always possible to specify the exact "effective amount".

[0015] The articles "a", "an" and "the" are intended to include the plural as well as the singular, unless the context clearly indicates otherwise. For example, some methods of the present invention are directed to improving heat stress in "wheat", which may include the control of multiple wheat plants (e.g., two or more wheat plants or two or more wheat species, etc.).

[0016] In one embodiment, the present invention is a method for improving heat stress tolerance in a plant, characterized by applying an effective amount of ACC to the plant, wherein the plant is selected from the group consisting of wheat, corn, soybean, cotton, tomato, Brassica napus, legume, lettuce, turfgrass and ornamental plants.

[0017] Brassica napus includes all its varieties, subspecies and variants, such as, but not limited to, annual rape, Argentine canola, canola, colza, Hanover-salad, oilseed rape, rape, rapeseed, rape kale, rutabaga, Siberian kale, summer rape, swede, Swede rape, Swedish turnip and winter rape. In a preferred embodiment, the variant of Brassica napus is canola.

[0018] In a preferred embodiment, the plant is selected from the group consisting of wheat, corn, soybean, cotton, tomato, Brassica napus, legume, lettuce, and turfgrass. In a more preferred embodiment, the plant is selected from the group consisting of wheat, corn, soybean, cotton, tomato, Brassica napus, legume, and lettuce. In an even more preferred embodiment, the plant is wheat, lettuce, soybean, or Brassica napus.

[0019] In a preferred embodiment, the effective amount of ACC is from about 1 to about 1,000 parts per million ("ppm"), more preferably from about 1 to about 500 ppm, even more preferably from about 10 to about 300 ppm, even more preferably from about 30 to about 300 ppm, and most preferably from about 30 to about 100 ppm.

[0020] In another preferred embodiment, ACC is applied to the plant at a rate of from about 0.001 to about 1,000 grams per hectare ("g / HA"), more preferably from about 0.028 to about 281 g / HA, even more preferably from about 0.28 to about 28 g / HA.

[0021] The method of the present invention contemplates applying ACC to the plant at any growth stage of the plant. In a preferred embodiment, ACC is applied to the plant during the development, growth and / or reproductive stages of the plant, including the flowering stage.

[0022] In another preferred embodiment, ACC is applied to the wheat plant from Feekes stage 2 to Feekes stage 11, even more preferably from Feekes stage 2 to Feekes stage 5, or from Feekes stage 10 to Feekes stage 11, even more preferably at Feekes stage 5, or from Feekes stage 10.4 to 10.5.

[0023] In another preferred embodiment, ACC is applied to the lettuce plant during the growth stage, more preferably during the head formation stage, also known as the rosette stage.

[0024] In another preferred embodiment, ACC is applied to the soybean plant during the growth stage, more preferably from V1 to V4 stage.

[0025] In another preferred embodiment, ACC is applied to the Brassica napus plant during the reproductive stage, more preferably during the flowering stage.

[0026] The ACC of the present invention can be applied by any convenient means. Those skilled in the art are familiar with application methods such as, but not limited to, spraying, brushing, dipping, infurrow treatment, drip irrigation, drenching, sprenching, dusting, powdering, granulating, seed treatment, pressurized liquid (aerosol), fogging, or side-dressing. In a preferred embodiment, the ACC is applied to the plant as a spray, and even more preferably as a foliar spray.

[0027] These representative embodiments are in no way limiting and are described only for the purpose of illustrating some aspects of the present invention.

[0028] Furthermore, the following examples are provided as examples only and are not intended to be limiting.

Examples

[0029] Example 1 - Heat stress tolerance in wheat after ACC application at the flowering stage Method Twenty-four-pot Apogee wheat was cultivated in a greenhouse under three consecutive growth conditions. First, the wheat was planted in ProMix® growing medium and cultivated in a growth cabinet for approximately six weeks under a 24 / 18 °C day / night temperature and a 16 / 8 hour light / dark photoperiod. After the first series of growth conditions, the plants were at the Feekes stage 10.4 - 10.5 (flowering stage). Next, using a tank sprayer, spray applications of 0, 30, or 100 ppm ACC (n = 8) were performed. Then, two days after the treatment, the plants were transferred to a growth chamber for seven days under a 36 / 30 °C day / night temperature and a 16 / 8 hour light / dark photoperiod. These conditions constitute significant heat stress. Finally, the plants were then returned to the growth chamber for an additional approximately five weeks under a 24 / 18 °C day / night temperature and a 16 / 8 hour light / dark photoperiod. The plants were then destructively harvested, and the total number of spikes, spike weight, grain weight, yield per spike, Δ spike number, and harvest index were measured. The Δ spike number is calculated by subtracting the number of tillers at the time of treatment from the number of tillers at harvest. An increase in tillers indicates an increase in reproductive success. The harvest index is calculated by dividing the number of pounds of grain by the total number of pounds of above-ground biomass. The results are shown in Table 1 below.

Table 1

[0030] Results As shown in Table 1 above, the application of ACC prior to heat stress resulted in an increase in the number of spikes, spike weight, grain weight, yield per spike, tiller number, and harvest index. These increases were dose-dependent. Specifically, the application of 30 ppm ACC increased the number of spikes by 6.3%, spike weight by 8.5%, grain weight by 15.7%, yield per spike by 11.1%, tiller number by 16.7%, and harvest index by 10.3% compared to the control. The application of 100 ppm ACC increased the number of spikes by 10.5%, spike weight by 14.5%, grain weight by 25.7%, yield per spike by 17.8%, tiller number by 42.0%, and harvest index by 17.2% compared to the control. Therefore, the application of ACC at the flowering stage of wheat protected wheat from subsequent heat stress and increased yield and reproductive success compared to wheat not treated with ACC.

[0031] Example 2 - Heat stress tolerance in wheat after ACC application at Feekes stage 5 Method Apogee wheat was sown in ProMix BX (Premier Horticulture) and cultivated in a growth cabinet under the following conditions: a day / night temperature of 24 / 18 °C and a light / dark photoperiod of 16 / 8 hours. After the first set of growth conditions on November 15, 2019, the plants were evaluated at the Feekes stage 5. Next, a spray application of 0, 0 or 100 ppm ACC (n = 8) was performed using a track sprayer. Then, two days after the treatment, one set of control plants ("treatment control") and the 100 ppm ACC plants were transferred to a second growth cabinet for 4 days under a day / night temperature of 34 / 28 °C and a light / dark photoperiod of 16 / 8 hours. These conditions constitute significant heat stress. Another set of control plants ("stress control") was left in the first growth cabinet. Finally, the plants were then returned to the first growth cabinet for approximately 14 days until the plants reached the Feekes stage 10.1 (heading). The plants were then destructively harvested and the total number of spikes, fresh weight, total number of tillers, percentage of tillers, Δ population density and total dry weight were measured. The percentage of tillers is calculated by dividing the number of tillers by the total number of spikes at harvest. The Δ population density is calculated by subtracting the population density at the time of treatment from the population density at harvest. The results are shown in Table 2 below.

Table 2

[0032] Results As shown in Table 2 above, the application of ACC prior to heat stress resulted in an increase in fresh weight, total tiller number, % tiller, Δ community density, and total dry weight, compared to wheat plants that were subjected to heat stress but not treated with ACC. The % reproductive tiller was returned to within 1.1% of that of wheat plants grown under ideal conditions (i.e., stress control). Furthermore, the Δ community density increased by 42.1% in ACC-treated wheat plants compared to wheat plants grown under ideal conditions. Therefore, the application of ACC during the vegetative growth stage of wheat protected the wheat from subsequent heat stress and increased yield and reproductive success compared to heat-stressed wheat that was not treated with ACC.

[0033] Example 3 - Heat stress tolerance in wheat after ACC application at Feekes stage 5 Method Apogee wheat was sown into ProMix BX (Premier Horticulture) and grown in a growth cabinet under the following conditions: 24 / 18 °C day / night temperature, 16 / 8 h light / dark photoperiod. After the first set of growth conditions, the plants were at the Feekes stage 5. Next, on October 4, 2019, a spray application of 0, 0, 100, or 300 ppm ACC (n = 8) was performed using a track sprayer. Then, two days after the treatment, one set of control plants (“treatment control”) and the 100 ppm ACC plants were transferred to a second growth cabinet for 4 days under a 34 / 28 °C day / night temperature, 16 / 8 h light / dark photoperiod. These conditions constitute significant heat stress. Another set of control plants (“stress control”) was left in the first growth cabinet. Finally, the plants were then returned to the first growth cabinet for approximately 14 days to allow the plants to reach the Feekes stage 10.1 (heading). The plants were then destructively harvested and the total number of spikes, fresh weight, total reproductive tiller number, proportion of reproductive tillers, Δ community density, and total dry weight were measured. The proportion of reproductive tillers was calculated by dividing the number of reproductive tillers by the total number of spikes at harvest. The Δ community density was calculated by subtracting the community density at the time of treatment from the community density at harvest. The results are shown in Table 3 below.

Table 3

[0034] Results As shown in Table 3 above, the application of 100 ppm ACC prior to heat stress resulted in increases in the number of ears, fresh weight, total tiller number, Δ community density, and total dry weight, compared to wheat plants that were exposed to heat stress but not treated with ACC. The application of 300 ppm ACC prior to heat stress resulted in increases in the number of ears and total dry weight. Therefore, the application of ACC during the vegetative growth stage of wheat protected the wheat from subsequent heat stress and increased the yield compared to heat-stressed wheat that was not treated with ACC.

[0035] Example 4 - Heat stress tolerance in lettuce after ACC application at the rosette stage Method Butterhead lettuce was sown in ProMix BX (Premier Horticulture) and grown in a greenhouse. Three weeks after sowing, the lettuce was transferred to a growth cabinet under the following conditions: a day / night temperature of 24 / 18 °C and a light / dark photoperiod of 16 / 8 hours. After the first set of growth conditions, the plants were in the head-forming (rosette) stage. Four days after transfer, the community density was calculated. Next, on September 21, 2020, a spray application of 0, 10, 30, or 100 ppm ACC (n = 5) was performed using a track sprayer. Next, two days after the treatment, half of the plants were transferred to a second growth cabinet for 9 days under a day / night temperature of 34 / 30 °C and a light / dark photoperiod of 16 / 8 hours. These conditions constitute significant heat stress. The plants were then destructively harvested, and the fresh weight, size, community density, and Δ community density were measured. The Δ community density was calculated by subtracting the community density at the time of treatment from the community density at harvest. The results are shown in Table 4 below.

Table 4

[0036] Results As shown in Table 4 above, the application of 10, 30 or 100 ppm ACC prior to heat stress resulted in an increase in fresh weight, canopy density and Δ canopy density, compared to lettuce plants that were exposed to heat stress but not treated with ACC. Therefore, the application of ACC during the vegetative growth stage of lettuce protected the lettuce from subsequent heat stress and increased the yield compared to heat-stressed lettuce that was not treated with ACC.

[0037] Example 5 - Heat stress tolerance in soybean after ACC application Method Williams 82 soybeans were sown in ProMix BX (Premier Horticulture) and cultivated in a greenhouse. When the soybean plants showed fully expanded first trifoliate leaves (V1 growth stage), the canopy density of the soybeans was measured and grouped for treatment and replication based on relative size. Next, on October 9, 2020, spray applications of 0, 10, 30 or 100 ACC (n = 5) were performed using a track sprayer. After spraying, the plants were transferred to a growth cabinet under the following conditions: 24 / 18 °C day / night temperature, 16 / 8 hour light / dark cycle. Next, three days after treatment, half of the plants were transferred to a second growth cabinet for six days under a day / night temperature of 34 / 30 °C and a light / dark photoperiod of 16 / 8 hours. These conditions constitute early heat stress. The plants were then destructively harvested and fresh weight, canopy density, Δ canopy density, total dry weight and height were measured. The results are shown in Table 5 below.

Table 5

[0038] Results As shown in Table 5 above, the application of 10 and 30 ppm ACC prior to heat stress resulted in an increase in fresh weight, colony density, Δ colony density, total dry weight, and height, compared to soybean plants that were subjected to heat stress but not treated with ACC. The application of 100 ppm ACC prior to heat stress resulted in an increase in total dry weight. Therefore, the application of ACC during the vegetative growth stage of soybeans protected the soybeans from subsequent heat stress and increased the yield compared to heat-stressed soybeans that were not treated with ACC.

[0039] Example 6 - Heat stress tolerance in rapeseed after ACC application Method Dwarf varieties of Brassica napus, which are commonly used as experimental models for the canola variety of Brassica napus, were sown in ProMix BX (Premier Horticulture) and cultivated in a greenhouse. Next, using a track sprayer, spray applications of 0, 10, 100, or 300 ppm ACC (n = 10) were performed 6 days after the Brassica napus plants began to flower. After spraying, the plants were transferred to a growth cabinet under the following conditions: a day / night temperature of 24 / 18°C and a light / dark cycle of 16 / 8 hours. Next, 2 days after the treatment, half of the plants were transferred to a second growth cabinet for 7 days under a day / night temperature of 36 / 30°C and a light / dark cycle of 16 / 8 hours. These conditions constitute early heat stress. The flowers and sheaths were always pruned, and the number of sheaths was kept at approximately 10. The plants were then destructively harvested, and the seed yield, number of seeds, seed yield per sheath, number of seeds per sheath, and single-seed weight were measured. The results are shown in Table 6 below.

Table 6

[0040] Results As shown in Table 6 above, the application of 30, 100 or 300 ppm ACC prior to heat stress resulted in an increase in seed yield per sheath and the number of seeds per sheath, even when compared to Brassica napus plants that were exposed to heat stress but not treated with ACC. The application of 10 ppm ACC prior to heat stress resulted in an increase in the number of seeds per sheath. Therefore, the application of ACC at the early stage of reproductive growth of Brassica napus dwarf varieties protected the plants from subsequent heat stress and increased the yield compared to heat-stressed Brassica napus dwarf varieties that were not treated with ACC.

Claims

**Claim 1** A method for improving heat stress tolerance in a plant, comprising applying an effective amount of 1-amino-1-cyclopropanecarboxylic acid (ACC) to the plant, wherein the plant is selected from the group consisting of wheat, corn, soybean, rapeseed, and lettuce, wherein the ACC is applied to the plant during the growth stage of the plant from germination to maturity in wheat, from germination to flowering in corn, from germination to flowering in soybean, from germination to flowering in rapeseed, and from germination to rosette formation in lettuce, method. **Claim 2** The method according to claim 1, wherein the plant is wheat. **Claim 3** The method according to claim 1, wherein the plant is lettuce. **Claim 4** The method according to claim 1, wherein the plant is soybean. **Claim 5** The method according to claim 1, wherein the plant is rapeseed. **Claim 6** The method according to claim 1, wherein the effective amount is from about 1 to about 1000 ppm. **Claim 7** The method according to claim 1, wherein the effective amount is from about 10 to about 300 ppm. **Claim 8** The method according to claim 1, wherein the effective amount is from about 30 to about 300 ppm. **Claim 9** The method according to claim 1, wherein the effective amount is from about 30 to about 100 ppm. **Claim 10** The method according to claim 1, wherein the effective amount is from about 10 to about 100 ppm. **Claim 11** The method according to claim 1, wherein the effective amount is from about 10 to about 30 ppm. **Claim 12** The method according to claim 1, wherein the ACC is applied to the plant as a spray. **Claim 13** The method according to claim 12, wherein the ACC is applied to the plant as a foliar spray. **Claim 14** The method according to claim 1, wherein the ACC is applied to the plant during the growth period of the plant. **Claim 15** The method according to claim 2, wherein the ACC is applied to the wheat between Feekes stage 2 and Feekes stage 11. **Claim 16** The method according to claim 2, wherein the ACC is applied to the wheat at Feekes stage 5. **Claim 17** The method according to claim 2, wherein the ACC is applied to the wheat between Feekes stage 10.4 and Feekes stage 10.

5. **Claim 18** The method according to claim 3, wherein the ACC is applied to the lettuce during the rosette stage of the lettuce. **Claim 19** The method according to claim 4, wherein the ACC is applied to the soybean during the period from V1 to V4 stages. **Claim 20** The method according to claim 5, wherein the ACC is applied to the Brassica napus plant during the flowering period of the Brassica napus plant.

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