CN114793675A - Method for rapidly screening optimal concentration of methyl jasmonate for relieving salt stress of corn seedlings - Google Patents
Method for rapidly screening optimal concentration of methyl jasmonate for relieving salt stress of corn seedlings Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G22/00—Cultivation of specific crops or plants not otherwise provided for
- A01G22/20—Cereals
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- G—PHYSICS
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- G01D—MEASURING NOT SPECIALLY ADAPTED FOR A SPECIFIC VARIABLE; ARRANGEMENTS FOR MEASURING TWO OR MORE VARIABLES NOT COVERED IN A SINGLE OTHER SUBCLASS; TARIFF METERING APPARATUS; MEASURING OR TESTING NOT OTHERWISE PROVIDED FOR
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- G01D21/02—Measuring two or more variables by means not covered by a single other subclass
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/62—Detectors specially adapted therefor
- G01N30/72—Mass spectrometers
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/02—Food
- G01N33/10—Starch-containing substances, e.g. dough
Abstract
A method for rapidly screening exogenous brassinosteroids to relieve the optimal concentration of salt stress of corn seedlings relates to the field of development and utilization of saline-alkali soil and production guarantee of corn. The content comprises the steps of collecting materials, preparing an EBR solution, pretreating seeds, carrying out a seedling stress experiment, collecting characters, applying optimal concentration and analyzing results. According to the method, a membership function method is adopted, the alleviation effects of exogenous MeJA with different concentrations on the salt stress of the corn are comprehensively evaluated, technical references are provided for the rapid screening of the MeJA concentration of the corn and the efficient application of the MeJA to alleviate the salt stress, and the method has high application value.
Description
Technical Field
The invention relates to the technical field of agriculture, in particular to a method for rapidly screening an optimal concentration of methyl jasmonate for relieving salt stress of corn seedlings.
Background
The corn is used as the main energy, economy and food crops in China, and the production safety of the corn plays an extremely important role in the stable production of food in China. Meanwhile, corn is also a salt-sensitive crop, and the tolerance of the whole growth period to salt stress is poor. Research shows that when the salt content of the soil is 30% of the total salt content, the whole growth period of the corn is slightly stressed, and the phenomenon of slow growth is shown; when the soil content is 40% of the total salt content, the growth of corn plants is severely inhibited, and the phenomena of light inhibition and PS pi repair inhibition are generated, so that the photosynthesis is seriously reduced, and the corn cannot be harvested due to serious salinization stress. In recent years, in order to improve the tolerance of corn varieties to soil salinization, different scholars develop a series of related researches from different research directions, but mainly focus on two aspects, on one hand, the existing germplasm resources are screened under different salt stress conditions, so that some corn inbred lines with better salt tolerance are screened out, and excellent materials are provided for field assembly and molecular marker-assisted breeding in the future; another aspect is the alleviation of salt damage stress to corn by the addition of some exogenous regulatory substances. The new salt-tolerant corn variety cultivated by the salt-tolerant inbred line not only needs longer time, but also needs to be researched only by utilizing salt-tolerant materials to combine the salt tolerance of the new variety. Some recent studies indicate that methyl jasmonate (MeJA) plays an important role in relieving stress resistance of plants.
Methyl jasmonate is a novel growth regulating substance, widely exists in higher plants and participates in a plurality of important reaction processes in the plants. In recent years, studies for alleviating salt stress tolerance of plants using methyl jasmonate have been reported successively. Yoon and other researches indicate that applying exogenous MeJA can effectively improve the chlorophyll content in the soybean body under salt stress and promote photosynthesis. At the same time, the application of exogenous MeJA also promotes the change of other endogenous hormones in the soybean body. Wang et al have shown that exogenous MeJA application alters the SOD, CAT, POD activity and amino acid levels in strawberries, thereby improving drought tolerance. AMOR and the like indicate that the salt tolerance of the broccoli can be improved by spraying MeJA on the leaf surfaces through researches on the broccoli. In addition, Liu, SALIMI and the like find that the MeJA with proper concentration can promote the growth of plant plants and reduce the harm of salt stress by researching the plants of the ammopiptanthus nanus and the chamomile. Although different researchers have shown the role of exogenous MeJA in plant salt stress relief in different crops, there have been relatively few studies on salt stress relief in corn, and screening methods have yet to be perfected. Therefore, the method for rapidly screening the optimal concentration of the methyl jasmonate for relieving the salt stress of the corn seedlings is provided by researching the relieving effect of MeJA with different concentrations on the corn seedlings under the salt stress.
Disclosure of Invention
The invention aims to provide a method for rapidly screening the optimal concentration of exogenous methyl jasmonate for relieving salt stress of corn seedlings. The method can effectively improve the salt tolerance of the corn in the seedling stage, and can provide a certain reference for developing and utilizing saline-alkali soil in China, enlarging the corn planting area and improving the corn planting area.
In order to solve the problems in the prior art, the invention provides a method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of corn seedlings, which comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, item and method determination, result analysis and the like, and comprises the following specific steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
Preferably, the specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution to dissolve, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
Preferably, in the step (2), 5 kinds of treatment liquids with different concentrations, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1:180Mmol/LNaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3, 180Mmol/L NaCl +5.0 umol/L; m4:180Mmol/LNaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
Preferably, the specific operation steps in the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
Preferably, the specific operation step in the step (4) is that after the corresponding treatment solution and vermiculite are uniformly stirred, the soaked seeds are planted in small flowerpots respectively, each treatment is repeated for 3 times, the seeds are placed in a human illumination incubator for cultivation after being planted, the day/night temperature is (27 +/-1)/(25 +/-1) ° c, the light cycle is set to 14 hours of illumination, and the dark is 10 hours. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling is quickly washed away by distilled water, and attached water is sucked dry by a sterilized filter paper to carry out index determination.
Preferably, in the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system specifically comprises the following steps: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
Preferably, the seedling form in the step (5) includes data indexes such as seedling length, root length, fresh weight of overground part, fresh weight of underground part, projection area of root system, surface area of root system and volume of root system.
Preferably, the relative content of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves is determined by HPLC tandem mass spectrometry.
Compared with the prior art, the invention has the beneficial effects that: according to the method, 1 part of salt-sensitive corn inbred line is taken as a test material, 5 exogenous MeJAs with different concentrations are added, and the influence of the MeJA treatment with different concentrations on the morphological index and the physiological index of the corn seedling under salt stress is researched; on the basis, the effect of relieving the salt stress of different exogenous MeJAs is objectively and comprehensively evaluated by adopting a membership function method, the method provided by the invention is simple and easy to operate, has good repeatability, can provide reference for knowing the relieving effect of the exogenous MeJAs on the salt stress of different crops in the future, provides technical reference for quickly screening the MeJA concentration of the corn and efficiently applying the MeJA to relieve the salt stress, and has high application value.
Drawings
FIG. 1 is a schematic diagram showing the influence of different concentrations of MeJA treatment on seedling length, root length, fresh weight of overground part and fresh weight of underground part of a maize inbred line under salt stress;
FIG. 2 is a schematic diagram showing the influence of MeJA treatment with different concentrations on the total root length, surface area, projection area and root volume of a maize inbred line under salt stress;
FIG. 3 is a graph showing the effect of different concentrations of MeJA treatment on POD activity, SOD activity and MDA content of maize inbred lines under salt stress.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example one
The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items and measurement methods, result analysis and the like, and specifically comprises the following steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
The specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
In the step (2), 5 treatment liquids with different concentrations need to be prepared, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1, 180Mmol/L NaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3:180Mmol/LNaCl +5.0 umol/L; m4, 180Mmol/L NaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
The specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
And (4) uniformly stirring the corresponding treatment solution and vermiculite, planting the soaked seeds in small flowerpots respectively, repeating the treatment for 3 times, placing the seeds in a human illumination incubator for cultivation after planting, wherein the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination and 10h of darkness. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
In the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system is specifically as follows: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
And (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
Relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
During the experiment, MeJA treating solution with the concentration of 180Mmol/LNaCl +0.1umol/L prepared in the step (2) is adopted.
Example two
The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items and measurement methods, result analysis and the like, and specifically comprises the following steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing the result.
The specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
In the step (2), 5 treatment liquids with different concentrations need to be prepared, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1, 180Mmol/L NaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3:180Mmol/LNaCl +5.0 umol/L; m4, 180Mmol/L NaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
The specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
And (4) uniformly stirring the corresponding treatment solution and vermiculite, planting the soaked seeds in small flowerpots respectively, repeating the treatment for 3 times, placing the seeds in a human illumination incubator for cultivation after planting, wherein the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination and 10h of darkness. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
In the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system is specifically as follows: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
And (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
Relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
During the experiment, MeJA treatment solution with the concentration of 180Mmol/L NaCl +1.0umol/L prepared in the second step is adopted.
EXAMPLE III
The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items and measurement methods, result analysis and the like, and specifically comprises the following steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
The specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
In the step (2), 5 treatment liquids with different concentrations need to be prepared, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1, 180Mmol/L NaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3:180Mmol/LNaCl +5.0 umol/L; m4, 180Mmol/L NaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
The specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
And (4) uniformly stirring the corresponding treatment solution and vermiculite, planting the soaked seeds in small flowerpots respectively, repeating the treatment for 3 times, placing the seeds in a human illumination incubator for cultivation after planting, wherein the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination and 10h of darkness. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
In the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system is specifically as follows: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
And (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
Relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
And in the experiment, the treatment solution with the concentration of 180Mmol/LNaCl +5.0umol/L prepared in the step two is adopted.
Example four
The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items, measurement method, result analysis and the like, and specifically comprises the following steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
The specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
In the step (2), 5 treatment liquids with different concentrations need to be prepared, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1, 180Mmol/L NaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3:180Mmol/LNaCl +5.0 umol/L; m4, 180Mmol/L NaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
The specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
And (4) uniformly stirring the corresponding treatment solution and vermiculite, planting the soaked seeds in small flowerpots respectively, repeating the treatment for 3 times, placing the seeds in a human illumination incubator for cultivation after planting, wherein the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination and 10h of darkness. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
In the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system is specifically as follows: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
And (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
Relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
The concentration prepared in the second step is 180Mmol/LNaCl +10.0umol/L during the experiment.
EXAMPLE five
The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items and measurement methods, result analysis and the like, and specifically comprises the following steps:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
The specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
In the step (2), 5 treatment liquids with different concentrations need to be prepared, specifically CK +: distilled water; CK-: 180Mmol/LNaCl solution; m1, 180Mmol/L NaCl +0.1umol/L MeJA; m2, 180Mmol/L NaCl +1.0 umol/L; m3:180Mmol/LNaCl +5.0 umol/L; m4, 180Mmol/L NaCl +10.0 umol/L; m5:180Mmol/LNaCl +20.0 umol/L.
The specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
And (4) uniformly stirring the corresponding treatment solution and vermiculite, planting the soaked seeds in small flowerpots respectively, repeating the treatment for 3 times, placing the seeds in a human illumination incubator for cultivation after planting, wherein the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination and 10h of darkness. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
In the step (5), 5 plants are selected for each treatment of each material, each treatment is repeated for 3 times, and the determination of the physiological indexes of the root system is specifically as follows: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
And (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
Relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
And in the experiment, the treatment solution with the concentration of 180Mmol/LNaCl +20.0umol/L prepared in the step two is adopted.
After analyzing the data, the comparison shows that 0.1 umol.L is applied -1 -5.0umol·L -1 The exogenous MeJA of (1) can relieve salt stress, but the relieving effect is best when the concentration is 1.0 umol/L.
In the above experiments, a membership function method is adopted to calculate membership function values of indexes under different hormone treatments, and comprehensively evaluate each treatment, wherein the membership function calculation formula is as follows:
Uij＝(Xij-Xijmin)/(Xijmax-Xijmin)
in the formula, Uij represents a membership value of j index under i processing, Xij represents a measured value of j index under i processing, and Xijmin and Xijmax represent minimum and maximum values of j index under all processing. And respectively accumulating membership function values of the traits under corresponding treatment, calculating the arithmetic mean of the membership function values, and comparing, wherein the larger the value is, the better the salt stress relieving effect of the exogenous MeJA is, all test data are processed and plotted by adopting Microsoft Excel2019 software, and Duncan multiple comparative analysis of variance and correlation analysis are carried out on the data by adopting SPSS22.0 software.
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, and the preferred embodiments of the present invention are described in the above embodiments and the description, and are not intended to limit the present invention. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (8)
1. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings comprises the steps of material collection, MeJA treatment fluid preparation, seed selection and disinfection, seed soaking, salt stress test, measurement items and measurement methods, result analysis and the like, and is characterized by comprising the following specific steps of:
(1) collecting materials: selecting a part of salt-sensitive inbred line N192;
(2) preparing a MeJA solution and a treatment solution;
(3) pretreating seeds;
(4) a hormone concentration screening test;
(5) measuring the form of the seedling;
(6) and (6) analyzing results.
2. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: the specific method for preparing the MeJA solution in the step (2) is to absorb 22.92ul of a small amount (5-10ml) of 95% ethanol of methyl jasmonate stock solution for dissolution, and then to fix the volume to 1L by using double distilled water to prepare 100umol/L MeJA mother solution.
3. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: in the step (2), 5 treatment solutions with different concentrations, specifically CK +: distilled water; CK-: 180Mmol/L NaCl solution; m1:180Mmol/LNaCl +0.1umol/L MeJA; m2:180Mmol/LNaCl +1.0 umol/L; m3, 180Mmol/L NaCl +5.0 umol/L; m4:180Mmol/LNaCl +10.0 umol/L; m5:180Mmol/L NaCl +20.0 umol/L.
4. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: the specific operation steps of the step (3) are as follows: selecting corn seeds with uniform size, full seeds and no damage, soaking the seeds in 70% ethanol for 1Min30s, washing with double distilled water for 2-3 times, sterilizing with 0.5% NaClO solution for 5-10Min, washing with double distilled water for 3 times, adsorbing water with sterilizing filter paper, and soaking the seeds in distilled water for 12 h.
5. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: the specific operation step of the step (4) is that after corresponding treatment liquid and vermiculite are uniformly stirred, the soaked seeds are respectively planted in small flowerpots, each treatment is repeated for 3 times, the seeds are placed in a human illumination incubator for cultivation after being planted, the day/night temperature is (27 +/-1)/(25 +/-1) DEG C, the light cycle is set to 14h of illumination, and 10h of darkness is set. Pouring 50mL of corresponding desalting treatment solution every 1d, and adding 50mL of corresponding treatment solution every 3 d; after about 12 days of treatment, the vermiculite at the root of the seedling was quickly washed away with distilled water, and the adhering water was blotted up with sterilized filter paper for index measurement.
6. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: in the step (5), 5 plants are selected for each treatment of each material, 3 repetitions are set for each treatment, and the determination of the physiological indexes of the root system specifically comprises the following steps: after about 12 days of treatment, the superoxide dismutase activity, the peroxidase activity and the malondialdehyde content of the corn seedling root system are measured, each material is measured for 3 times respectively, and the average value is taken.
7. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: and (5) the seedling form in the step (5) comprises data indexes such as seedling length, root length, fresh weight of the overground part, fresh weight of the underground part, projection area of the root system, surface area of the root system, volume of the root system and the like.
8. The method for rapidly screening the optimal concentration of methyl jasmonate for relieving the salt stress of the corn seedlings according to claim 1, wherein the optimal concentration of methyl jasmonate in the salt stress relieving corn seedlings is as follows: relative contents of auxin (IAA), gibberellin (GA3), abscisic acid (ABA), methyl jasmonate (MeJA) and 2, 4-Epibrassinolide (EBR) in maize leaves were determined by HPLC tandem mass spectrometry.
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