CN115735700A - Method for improving heat resistance of carnation by using melatonin - Google Patents
Method for improving heat resistance of carnation by using melatonin Download PDFInfo
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Abstract
The invention relates to the technical field of improving plant resistance, in particular to a method for improving heat resistance of carnation by using melatonin. The method provided by the invention comprises the following steps: spraying melatonin solution on the surfaces of carnation leaves; the concentration of the melatonin solution is 25-200 mu mol/L. By spraying the melatonin solution with proper concentration on the leaf surfaces of the carnation, the heat damage condition of the carnation can be obviously relieved, the chlorophyll content is improved, the electrolyte leakage rate is reduced, and the accumulation of osmotic substances is adjusted; the stomata can be promoted to be opened, and the structural integrity of chloroplast is protected; the expression of heat-resistant related genes can be also up-regulated on a molecular level, so that the heat resistance of the carnation is improved.
Description
Technical Field
The invention relates to the technical field of improving plant resistance, in particular to a method for improving heat resistance of carnation by using melatonin.
Background
Dianthus caryophyllus (Dianthus caryophyllus) is an important flower crop, is one of the four cut flowers in the world, and has great commercial value. However, because carnations like cold and cool climates and are not resistant to heat, high temperature can affect the growth and development of the carnations, thereby causing adverse effects on the yield and quality of the carnations and seriously affecting the aesthetic value and the commercial value of the carnations.
Many studies indicate that high temperature can accelerate the aging of plant leaves, chlorophyll degradation can cause oxidative damage to plants, stomata closing, chloroplast ultrastructural damage and finally plant yield reduction. The adverse effect of high temperature on plants can be relieved by means of covering a sunshade net, spraying exogenous substances, genetic engineering and the like, wherein the covering of the sunshade net wastes time and labor, the genetic engineering period is long, and the spraying of the exogenous substances is a method with relatively low cost and quick effect.
However, at present, the study on carnation mostly focuses on cut flower preservation; the technology for improving the heat resistance of the carnation has not been reported yet, which substances can be sprayed on the leaf surfaces to relieve the high-temperature damage of the carnation.
Disclosure of Invention
In order to solve the problems, the invention provides a method for improving the heat resistance of carnation by using melatonin. The method provided by the invention can relieve the high-temperature damage of the carnation by spraying the melatonin on the leaf surfaces of the carnation, thereby improving the heat resistance of the carnation.
In order to achieve the above purpose, the invention provides the following technical scheme:
the invention provides a method for improving the heat resistance of carnation by using melatonin, which comprises the following steps:
spraying melatonin solution on the leaf surfaces of carnation; the concentration of the melatonin solution is 25-200 mu mol/L.
Preferably, the concentration of the melatonin solution is 100 mu mol/L.
Preferably, the spraying standard is as follows: the method is based on that the carnation leaves are wet and the melatonin solution does not drip.
Preferably, the spraying frequency is 1 time per day, and the spraying days are 6 days.
Preferably, the improving heat resistance of carnation includes: improving carnation biomass, reducing oxidation damage, improving chlorophyll content, reducing electrolyte leakage rate, regulating osmotic substance accumulation, promoting stomatal opening, protecting chloroplast structural integrity, inhibiting expression of chlorophyll degradation gene DcpPH, and up-regulating expression of heat-resistant related gene.
Preferably, the heat-resistance-associated genes include: one or more of DcHsfA1d, dcHsfA2, dcHsfA4, dcHSP17.8 and DcMBF1 c.
Has the advantages that:
the invention provides a method for improving the heat resistance of carnation by using melatonin, which comprises the following steps: spraying melatonin solution on the surfaces of carnation leaves; the concentration of the melatonin solution is 25-200 mu mol/L. By spraying the melatonin solution with proper concentration on the leaf surfaces of the carnation, the heat damage condition of the carnation can be obviously relieved, the chlorophyll content is improved, the electrolyte leakage rate is reduced, and the accumulation of osmotic substances is adjusted; the stomata can be promoted to be opened, and the structural integrity of chloroplast is protected; the expression of heat-resistant related genes can be up-regulated on a molecular level, so that the heat resistance of the carnation is improved.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the embodiments will be briefly described below.
FIG. 1 is a diagram illustrating grading of heat injury of carnation; wherein, from top to bottom, the number of the stages is 0,1, 2, 3 and 4;
FIG. 2 is a bar graph of carnation plant height, total chlorophyll, relative electrolyte leakage rate and proline content in different treatment groups; different lower case letters indicate that significant differences were seen for different treatment groups (P < 0.05);
FIG. 3 is a graph of carnation phenotype and DAB, NBT staining in different treatment groups;
FIG. 4 is a micrograph of carnation porosity in different treatment groups;
FIG. 5 is an ultra-micro structural drawing of carnation chloroplasts and thylakoids in different treatment groups;
FIG. 6 is a bar graph of relative expression of chlorophyll degradation and heat resistance related genes in carnauba of different treatment groups; different lower case letters indicate that significant differences were seen for different treatment groups (P < 0.05).
Detailed Description
The invention provides a method for improving the heat resistance of carnation by using melatonin, which comprises the following steps:
and spraying the melatonin solution on the surfaces of the carnation leaves.
The concentration of the melatonin solution is 25-200 mu mol/L, preferably 25 mu mol/L, 50 mu mol/L, 100 mu mol/L, 150 mu mol/L or 200 mu mol/L, and more preferably 100 mu mol/L.
By spraying the melatonin solution with proper concentration on the carnation leaf surfaces, the heat damage condition of the carnation can be obviously relieved, particularly the high-temperature stress environment with the daytime temperature of about 42 ℃ and the night temperature of about 35 ℃ can be relieved, the chlorophyll content is improved, the electrolyte leakage rate is reduced, and the osmotic substance accumulation is adjusted; the stomata can be promoted to be opened, and the structural integrity of chloroplast is protected; the expression of heat-resistant related genes can be also up-regulated on a molecular level, so that the heat resistance of the carnation is improved.
In the present invention, the spraying criteria are preferably: the melatonin solution is not dropped because the carnation leaves are wet, and is preferably sprayed by 5 to 7mL per plant, and more preferably sprayed by 6mL per plant.
In the present invention, the frequency of spraying is preferably 1 time per day, and the number of spraying days is preferably 6 days.
In the present invention, the improving heat resistance of carnation preferably includes: improving carnation biomass, reducing oxidation damage, improving chlorophyll content, reducing electrolyte leakage rate, regulating osmotic substance accumulation, promoting stomatal opening, protecting chloroplast structural integrity, inhibiting expression of chlorophyll degradation gene DcpPH, and up-regulating expression of heat-resistant related gene.
The heat-resistance-associated gene of the present invention preferably includes: one or more of DcHsfA1d, dcHsfA2, dcHsfA4, dcHSP17.8 and DcMBF1 c.
In order to further illustrate the present invention, the method for improving the heat resistance of carnation using melatonin provided in the present invention will be described in detail with reference to the following examples, which should not be construed as limiting the scope of the present invention.
Example 1
A method for improving the heat resistance of dianthus caryophyllus by using melatonin comprises the following steps:
planting the purchased carnation seedlings 'Carlin wave' in a matrix, wherein the matrix consists of turfy soil and perlite, and the volume ratio of the turfy soil to the perlite is 6:4, setting the temperature at 21 ℃ in the daytime, setting the temperature at 19 ℃ at night, setting the illumination time at 16h, setting the illumination intensity at 7200LX, performing conventional culture at the humidity of 70%, slowing seedling, spraying a melatonin aqueous solution with the leaf spraying concentration of 25 mu mol/L to the carnation when the carnation grows to 6 pairs of true leaves, spraying at 8 o' clock every night, spraying once every day for 6 days continuously, wherein the leaf spraying amount is suitable for moistening the carnation leaves without dripping, and the spraying amount of each plant is 6mL.
Example 2
A process similar to that of example 1, the only difference being: the concentration of the melatonin aqueous solution was 50. Mu. Mol/L.
Example 3
A process similar to that of example 1, the only difference being: the concentration of the melatonin aqueous solution was 100. Mu. Mol/L.
Example 4
A method similar to example 1, the only difference being: the concentration of the melatonin aqueous solution was 150. Mu. Mol/L.
Example 5
A method similar to example 1, the only difference being: the concentration of the melatonin aqueous solution was 200. Mu. Mol/L.
Comparative example 1
A method similar to example 1, the only difference being: the melatonin solution was replaced with distilled water.
Test example 1
After melatonin or distilled water is sprayed on leaf surfaces, high-temperature stress treatment is carried out on carnations which are sprayed for 6 days in comparative example 1 (marked as HT) and examples 1-5 (marked as MT 1-MT 5 in sequence), wherein the high-temperature stress treatment temperature is 42 ℃ in the daytime, 35 ℃ in the evening, the illumination time is 16h, the illumination intensity is 7200LX, and the humidity is 70%; additionally arranging a normal temperature culture control group (marked as CK), culturing carnation of the CK group according to the method of the comparative example 1, wherein except the high temperature stress treatment temperature, the other treatment conditions are consistent with the comparative example 1, the treatment temperature of the CK group is 21 ℃ in the day, and 19 ℃ in the night; 7 treatment groups (HT, MT 1-MT 5 and CK) 9 carnations were performed per group in 3 replicates of 3 experiments.
After 12 days of high-temperature stress treatment or normal-temperature culture treatment, the relevant indexes of the carnations of 7 treatment groups are detected, and the method specifically comprises the following steps:
1) Specifically, the heat damage of carnations was classified as shown in table 1 and fig. 1, and then the heat damage index was calculated according to the formula heat damage index = Σ (number of plants at each stage × number of stages)/(maximum number of stages × total number of plants), and the results are shown in table 2.
TABLE 1 Classification of heat injury of Dianthus caryophyllus
TABLE 2 Dianthus caryophyllus heat damage index statistics
Note: the value in parentheses is a relative index of the control group HT of 100, calculated from the following formula: heat injury index in treatment group/heat injury index in HT group × 100.
As can be seen from Table 2, melatonin at each concentration can relieve high temperature injury of carnation, wherein MT3 is 100 mu mol/L, and the relieving effect is the best.
2) The plant height, the total chlorophyll content, the relative electrolyte leakage rate and the proline content of 7 treated carnations were measured, and the measurement results are shown in table 3 and fig. 2;
wherein the total chlorophyll content and the relative electrolyte leakage rate are determined according to the methods described in [ cost M, zhang W, zhang Q, et al, exogenous pharmaceutical acid amides stress-induced data and improvements growth and photosynthetic efficacy in alfa (medical go sativa L.) [ J ]. Ecotoxology and Environmental Safety,2020,191, 110206 ];
proline content was determined according to the method described in [ Tiwari J K, munshi A D, kumar R, et al. Effect of salt stress on focus: na + -K + ratio, osmolyte concentration, phenols and chlorophyl content [ J ]. Acta Physiologiae Plantarum,2010,32 (1): 103-114 ].
Table 3 measurement results of plant height, total chlorophyll content, relative electrolyte leakage rate and proline content of carnations of different treatment groups
As can be seen from table 3 and a in fig. 2, the height of the carnation plant was significantly reduced by the high-temperature treatment. Compared with a normal temperature Control (CK), the plant height under the high temperature treatment (HT) is obviously reduced by 29.74%, the plant height under the MT3 treatment is reduced by 26.57%, but the MT3 change is not obvious.
As shown in Table 3 and B in FIG. 2, the total chlorophyll content of the carnauba leaf significantly decreased under high temperature stress, and the chlorophyll content of HT significantly decreased by 41.07% relative to CK. MT3 was increased significantly by 31.82% compared to HT, with no significant difference with other treatments.
As can be seen from table 3 and C in fig. 2, the relative electrolyte leakage rate is an index reflecting the permeability of the plant cell membrane, and the relative electrolyte leakage rate of carnation under HT treatment is significantly increased, and compared to HT, MT1, MT2, MT3, MT4 and MT5 are respectively decreased by 31.07%, 17.35%, 37.38%, 29.28% and 19.72%, wherein the sustained release effect of MT3 is better.
As can be seen from table 3 and D in fig. 2, the proline content of carnation leaves after HT treatment was significantly increased by 192.85% compared to CK, and the proline content of carnation leaves after MT3 treatment was most significantly decreased by 60.83% compared to HT. The growth physiological results show that the melatonin can relieve the high temperature of the carnation under the treatment, wherein the MT3 is 100 mu mol/L, and the relieving effect is the best.
Test example 2
Carnation phenotype and oxidative damage status: after the melatonin with different concentrations is sprayed, the test example 1 shows that the MT3 relieving effect is the best, so the MT3 treatment is further analyzed, and the specific steps are as follows:
after melatonin or distilled water is sprayed on leaf surfaces, the carnation sprayed for 6 days in comparative example 1 (marked as HT) and example 3 (marked as MT 3) is subjected to high-temperature stress treatment, wherein the high-temperature stress treatment temperature is 42 ℃ in the daytime, 35 ℃ in the evening, the illumination time is 16h, the illumination intensity is 7200LX, and the humidity is 70%; a normal temperature culture control group (marked as CK) is additionally arranged, the CK group carnation is obtained by culturing according to the method of the comparative example 1, except the high temperature stress treatment temperature, the other treatment conditions are consistent with the comparative example 1, the treatment temperature of the CK group is 21 ℃ in the day, and 19 ℃ in the evening; 3 treatment groups (HT, MT3 and CK) 15 carnations were performed per group, 3 replicates per group, 5 for each.
After 12 days of high-temperature stress treatment or normal-temperature culture treatment, the relevant indexes of the carnation of 3 treatment groups are detected, and the method specifically comprises the following steps:
the phenotype of carnation in the CK, HT and MT3 groups was photographed and subjected to DAB and NBT staining according to the methods described in [ Su Y A, zu B, ams C, et al, salicylic acid underslung silicon in ameliorative chlorine toxicity in rice by modulating antioxidant defenses, ion homenosis and cellular architecture [ J ]. Plant Physiology and Biochemistry,2021 ], and the results are shown in FIG. 3.
As can be seen from FIG. 3, the damage of the plants is severe at high temperature, DAB and NBT staining is obviously deepened, and the damage is obviously relieved after MT3 treatment.
The above-ground fresh weight, the above-ground dry weight, the root fresh weight and the root dry weight of carnations in the CK, HT and MT3 groups in test example 2 were measured, and the results are shown in Table 4.
TABLE 4 fresh and Dry weight changes of carnation
Note: different lower case letters indicate significant differences (P < 0.05) between treatment groups.
As can be seen from table 4, the above-ground dry weight and the underground dry weight of the carnation treated by HT were significantly lower than CK, and the above-ground dry weight, the root dry weight and the root dry weight of the carnation treated by MT3 were increased by 73.77%, 33.33%, 123.53% and 50.00%, respectively, compared to HT. The results can show that the melatonin with proper concentration can obviously improve the biomass of the carnation under high-temperature treatment, reduce the oxidative damage of high temperature to plants and improve the heat resistance of the plants.
Microscopic changes in the pores, pore lengths, pore widths, pore diameters and pore areas of the carnations in CK, HT and MT3 group in test example 2 were observed and determined according to the methods described in the literature [ Jiangnan, huang Di, zhang Qian, lvxiao, ping-Lu-Xuan ] and the taxonomic significance of the classification [ J ]. Modern horticulture, 2022,45 (19): 33-35 ] and [ Ahmad S, muhammad I, wang G Y, et al.
TABLE 5 Dianthus caryophyllus pore characteristics variation
As can be seen from FIG. 4, the pores under CK were normally open, the pores under HT treatment were substantially closed, and the pores after MT3 treatment were open.
As can be seen from Table 5, HT significantly reduced the pore length (18.20%), width (22.57%), pore diameter (83.76%) and area (32.63%) compared to CK. However, compared with HT, the length, width, aperture and area of stomata after MT3 treatment are respectively increased by 14.66%, 21.76%, 163.16% and 34.10%, and it can be seen that melatonin with appropriate concentration can regulate stomata change of carnation at high temperature, thereby improving physiological metabolism and photosynthetic efficiency of plants.
The chloroplast and thylakoid ultrastructure changes of carnation in CK, HT and MT3 groups in test example 2 were observed according to the method described in the literature [ Jiang D, lu B, liu L, et al, exogenous genes expressed in the expression of the salt tolerance of cotton by removing active oxygen and protective photosynthetic organisms [ J ]. BMC Plant Biology,2021,21 (1) ], and the results are shown in FIG. 5.
As can be seen from FIG. 5, under CK treatment, the green body membrane of the carnauba leaf has a complete structure, is spindle-shaped as a whole, contains abundant white starch grains, has a complete and clear thylakoid lamellar structure, and has a small number of osmyl bodies. Under HT treatment, chloroplasts swell and even burst, starch granules disappear, thylakoids lamella are blurred and disordered, and a large number of osmyl bodies appear (B and E in figure 5). However, MT3 treated plants showed slight swelling, clear and tidy thylakoid lamellae and less accumulation of osmyl bodies (C, F in fig. 5). It can be seen that the high temperature stress destroys the chloroplast structure of carnation, and the melatonin with proper concentration reduces the damage of carnation leaf chloroplast and thylakoid.
According to literature [ Huang, c.; tian, Y.; zhang, b.; hassan, m.j.; li, z; zhu, Y.Chitosan (CTS) leaves Heat-Induced Leaf sensing in growing branched chromatographic methods, antibiotic defenses, and the Heat Shock pathway. Molecules 2021,26,5337. The results of the assay for the degradation of Chlorophyll and the relative expression of Heat resistance related genes in carnation in the CK, HT and MT3 groups of test example 2 are shown in FIG. 6 and Table 6.
TABLE 6 chlorophyll degradation and heat resistance related gene relative expression of carnation
As can be seen from fig. 6 and table 6, the chlorophyll degradation gene DcPPH was significantly increased under HT, whereas MT3 treatment was significantly 45.13% relative to HT treatment. The relative expression quantity of heat-resistant related genes is obviously increased at high temperature, and after MT3 treatment, the relative expression quantities of DcHsfA1d, dcHsfA2, dcHsfA4, dcHSP17.8 and DcMBF1c are respectively obviously increased by 2.66, 11.00, 2.14, 0.86 and 3.13 times. Melatonin improves heat resistance of carnation by reducing chlorophyll degradation gene expression and improving heat resistance related gene expression.
Comprehensive analysis of the heat damage condition, plant height, chlorophyll, electrolyte leakage rate and proline indexes of the carnation shows that 100 mu mol/L melatonin has the best effect of relieving high-temperature stress damage to the carnation, spraying the melatonin with the optimal concentration can improve the biomass of the carnation, reduce oxidation damage, promote stomata opening, protect the structural integrity of chloroplast, and regulate the expression of genes, so that the damage of high temperature to the carnation is obviously relieved.
Although the present invention has been described in detail with reference to the above embodiments, it is only a part of the embodiments of the present invention, not all of the embodiments, and other embodiments can be obtained without inventive step according to the embodiments, and the embodiments are within the scope of the present invention.
Claims (6)
1. A method for improving the heat resistance of dianthus caryophyllus by using melatonin is characterized by comprising the following steps of:
spraying melatonin solution on the surfaces of carnation leaves; the concentration of the melatonin solution is 25-200 mu mol/L.
2. The method of claim 1 wherein the concentration of the melatonin solution is 100 μmol/L.
3. The method of claim 1, wherein the spraying criteria are: the method is based on that the carnation leaves are wet and the melatonin solution does not drip.
4. The method according to claim 1 or 3, wherein the frequency of spraying is 1 time per day and the number of spraying days is 6 days.
5. The method of claim 1, wherein the increasing the heat resistance of the carnation comprises: improving carnation biomass, reducing oxidation damage, improving chlorophyll content, reducing electrolyte leakage rate, regulating osmotic substance accumulation, promoting stomatal opening, protecting chloroplast structural integrity, inhibiting expression of chlorophyll degradation gene DcpPH, and up-regulating expression of heat-resistant related gene.
6. The method of claim 5, wherein the heat-resistance-associated genes comprise: one or more of DcHsfA1d, dcHsfA2, dcHsfA4, dcHSP17.8 and DcMBF1 c.
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| CN106105660A (en) * | 2016-06-28 | 2016-11-16 | 四川农业大学 | A kind of it is obviously enhanced the method for Fructus Solani melongenae resistance under high temperature stress |
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| CN116458423A (en) * | 2023-03-31 | 2023-07-21 | 浙江大学 | Application of exogenous melatonin in improving heat resistance of Chinese cabbage and relieving high-temperature bitter of Chinese cabbage |
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