A method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures
By optimizing inflorescence trimming and plant hormone treatment, combined with shade treatment, the problems of high labor, low efficiency and poor economy in the existing tobacco seed production technology are solved, and the effect of extending the pollination window, improving seed quality and reducing labor costs is achieved.
Patent Information
- Application Number
- CN202310236200.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-03-13
- Publication Date
- 2025-06-17
- Estimated Expiration
- 2043-03-13
AI Technical Summary
The existing tobacco seed production technology has multiple disadvantages, including the need for a large number of skilled workers to determine the empowerability of the stigma, the tearing process is prone to damage the branches and stigma, the pollination window period is short, the flowering period is long, and the flowering period is inconsistent in the maturity period of different parts, which leads to time-consuming and labor-intensive and poor economicality.
The combined methods of optimizing inflorescence trimming, multiple plant hormone compound treatments and shade treatments are adopted, including trimming inflorescences during the budding period of tobacco plants, spraying solutions of turbid, ethylene and brassinin, and shade treatment during the pollination period.
The pollination window period has been extended, the seed quality has been improved, the labor cost has been reduced, and the economic benefits have been significantly improved. The maturity time span of the tobacco plant flower vessel can be reduced to 6 days, the exposure rate of the stigma can reach up to 85.8%, the fruit setting rate can reach up to 96%, the average pollination window period is up to 4 days, and the seed germination rate is up to 98.5%.
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Abstract
Description
Technical Field
[0001] The present invention belongs to the technical field of tobacco seed production, and particularly relates to a method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures. Background Art
[0002] As a self-pollinating plant, the phenotypic traits of tobacco varieties are basically the same among plants, indicating that their genotypes are generally in a homozygous state. Therefore, in the cross-breeding improvement of varieties, only two varieties need to be crossed to obtain the first-generation hybrids with relatively uniform phenotypes. At present, in the cross-breeding of tobacco seeds, male sterile lines are used as female parents. After artificially collecting the pollen of the male parent and storing it at low temperature, when the appropriate pollination period of the female parent stigma (bud stage) is selected, the flower is manually torn open, and then the pollen is manually applied to the stigma with a brush, or a specific device is used to spray pollen or pollen suspension. Such a seed production method has several disadvantages: (1) A large number of skilled workers who can accurately judge the stigma receptivity need to be hired; (2) The process of tearing the flower is likely to damage the flower branches, floral organs and stigmas; (3) The pollination window period is short, and the tearing of the flower and pollination need to be carried out synchronously; (4) The tobacco flowering period is relatively long, and the maturity periods of the flowers in different parts are inconsistent, so the flower needs to be torn and pollinated in batches at different times; (5) It is time-consuming and laborious, with low pollination efficiency and poor economy.
[0003] Predecessors have tried to breed female parent mutants with the characteristic of stigma exsertion (cytoplasmic inheritance) in an attempt to fundamentally avoid the complicated processes such as flower selection, flower tearing and individual pollination in tobacco seed production. At present, 4 improved tobacco female parents with the characteristic of stigma exsertion, namely "Yunyan 87", "K326", "NC89" and "Yuyan 12", have been successfully bred. Through years of seed production practice, it has been found that even when using the stigma-exserted mutant line as the female parent, there are still many problems in the pollination process: for example, (1) Different inflorescence trimming methods have a great influence on the expression of the stigma-exserted trait of the mutant; the time span of the maturity of the floral organs in different parts of the same plant can reach 15 days, and centralized pollination cannot be achieved; (2) Affected by factors such as high temperature and strong light intensity during the tobacco flowering period, the vitality of the exserted stigma is poor and the receptivity is reduced.
[0004] In addition, predecessors have conducted a large number of studies on mutant plants with the characteristic of stigma exsertion in materials such as Brassica napus, cotton, and tomato. It is generally believed that stigma exsertion is an abnormal expression controlled by genes related to the length of the stigma and the length of the petals. The process of stigma exsertion can be understood as the reciprocal change between the length of the petal and the length of the stigma. The elongation of the style is achieved by the overexpression of genes related to the synthesis of plant hormones such as auxin, cytokinin, gibberellin, salicylic acid, abscisic acid, jasmonic acid, and ethylene in the plant; the "shortening" of the corolla is achieved by controlling the silencing or low expression of genes related to the synthesis of auxin, cytokinin, gibberellin, polyamine, stress protein, as well as genes related to the metabolism of reactive oxygen species and carbon-nitrogen metabolism. The research also found that the relatively high metabolic intensity of salicylic acid, abscisic acid and jasmonic acid is the main factor leading to the significant shortening of the corolla length. Summary of the Invention
[0005] In view of the problems existing in the above-mentioned prior art, the present invention provides a method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures.
[0006] To achieve the above object, the present invention provides the following technical solutions:
[0007] A method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures, which uses the tobacco plants of the stigma-exserted mutant line as materials and is processed by combining an optimized inflorescence trimming method, a combined treatment of multiple phytohormones, and a shading treatment.
[0008] Further, the method for regulating the tobacco floral organs specifically includes the following steps:
[0009] (1) At the budding stage of the tobacco plants, trim the inflorescence, remove the apical inflorescence, and retain 8-10 stronger branches;
[0010] (2) Immediately spray a chlormequat solution after trimming the inflorescence, and spray an ethephon solution 3 days later;
[0011] (3) Spray a brassinolide solution at the initial flowering stage of the tobacco plants, and conduct a shading treatment during the same period (pollination period).
[0012] Further, the concentration of the chlormequat solution is 500 mg / L, and the spraying amount is 5 mL / plant.
[0013] Further, the concentration of the ethephon solution is 50 mg / L, and the spraying amount is 5 mL / plant.
[0014] Further, the concentration of the brassinolide solution is 150 mg / L, and the spraying amount is 5 mL / plant.
[0015] Further, the shading treatment refers to shading with a black sunshade net with a light transmittance of 65%.
[0016] Compared with the prior art, the present invention has the following advantages and technical effects:
[0017] By optimizing the inflorescence trimming method of tobacco plants and combining means such as a combined treatment of multiple phytohormones and a shading treatment during the pollination period, the present invention achieves the effects of extending the pollination window period, improving the seed quality, and reducing the labor cost. The minimum time span for the maturity of the tobacco floral organs can be reduced to 6 days, the highest stigma exsertion rate per plant can reach 85.8%, the highest fruit setting rate can reach 96%, the longest average pollination window period is 4 days, and the highest seed germination rate is 98.5%. Detailed Embodiments
[0018] The various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation on the present invention, but rather as a more detailed description of certain aspects, features, and implementation schemes of the present invention.
[0019] It should be understood that the terms described in the present invention are only for describing specific embodiments and are not used to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Each intermediate value within any stated value or stated range, as well as each smaller range between any other stated value or intermediate value within the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range.
[0020] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention pertains. Although the present invention only describes preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the said documents. In case of conflict with any incorporated document, the content of this specification shall prevail.
[0021] Without departing from the scope or spirit of the present invention, various improvements and changes can be made to the specific embodiments of the specification of the present invention, which are obvious to those skilled in the art. Other embodiments obtained from the specification of the present invention are obvious to those skilled in the art. The specification and examples of this application are merely exemplary.
[0022] Regarding the use of "comprising", "including", "having", "containing", etc. herein, they are all open-ended terms, meaning including but not limited to.
[0023] The present invention uses the stigma-exserted mutant line (CHE-01) of "Xiangyan No. 3" selected in the field as the material, and systematically conducts experiments on the effects of tobacco inflorescence trimming methods, compound treatments with various plant hormones, and shading treatments during the pollination period on the flowering consistency, stigma-exserted characteristics, stigma receptivity, fruit-setting characteristics, and economy of CHE-01. To extend the pollination window period, improve the seed quality, and reduce the labor cost, providing technical support for cost reduction and efficiency increase in tobacco seed production.
[0024] Specifically, the present invention provides a method for improving the stigma receptivity and pollination efficiency of a stigma-exserted mutant line of tobacco by using agronomic measures. Using the tobacco plants of the stigma-exserted mutant line as the material, the treatment is carried out by combining the optimized inflorescence trimming method, compound treatment with various plant hormones, and shading treatment. Specifically, the stigma-exserted mutant line "CHE-01" of "Xiangyan No. 3" is selected as the material.
[0025] The method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures specifically includes the following steps:
[0026] (1) During the budding stage of the tobacco plant, trim the inflorescence, and retain 8 - 10 stronger flower branches after removing the top inflorescence.
[0027] (2) Immediately spray a chlormequat solution after trimming the inflorescence, and spray an ethephon solution 3 days later;
[0028] (3) Spray a brassinolide solution during the initial flowering stage of the tobacco plant, and conduct shading treatment during the same period (pollination period).
[0029] In some preferred embodiments, during the flowering stage trimming, retaining 8 - 10 flower branches is to ensure the consistency of the tobacco flowering period as much as possible while maximizing the number of tobacco flowers. If too few, the seed yield per plant will be too low, which is not conducive to improving the economic benefits of seed production. If too many, it will lead to poor development of flower organs and it is difficult to ensure the flowering period consistency.
[0030] In some preferred embodiments, to achieve better effects, the present invention defines the concentration of the chlormequat solution as 500 mg / L. This concentration can ensure the effective regulation of the petal length. If the concentration is too high, the growth of the flower organs of the tobacco plant will be inhibited. If the concentration is too low, the petal length of the tobacco plant cannot be effectively regulated. The present invention further defines the spraying amount as 5 mL / plant. Selecting this spraying amount is because at this spraying amount, the petal length of the whole plant can be effectively regulated.
[0031] In some preferred embodiments, to achieve better effects, the present invention defines the concentration of the ethephon solution as 50 mg / L. This concentration can ensure the early unfolding of the corolla. If the concentration is too high, the tobacco flowers will prematurely senesce. If the concentration is too low, the corolla cannot unfold early or the degree of early unfolding does not meet the requirements. The present invention further defines the spraying amount as 5 mL / plant. Selecting this spraying amount can effectively regulate the concentrated opening of the flower organs.
[0032] In some preferred embodiments, to achieve better effects, the present invention defines the concentration of the brassinolide solution as 150 mg / L. This concentration can ensure the effective growth of the stigma length of the tobacco flowers. If the concentration is too high, both the petals and the stigma of the tobacco plant will overgrow. If the concentration is too low, the growth length of the stigma of the tobacco plant will not reach the protruding effect. The present invention further defines the spraying amount as 5 mL / plant. Selecting this spraying amount is to make the tobacco flower organs larger and the stigma can effectively elongate.
[0033] In some preferred embodiments, in order to achieve better effects, the present invention further defines that the shading treatment specifically uses a black sunshade net with a light transmittance of 65% for shading. Shading is carried out until the artificial pollination ends to ensure that the activity of the stigma remains at a relatively high level during pollination, reducing the influence of high temperature and strong light on the stigma activity.
[0034] All raw materials used in the following examples of the present invention are commercially available.
[0035] The following examples are further illustrations of the technical solutions of the present invention.
[0036] Example 1
[0037] 1. Materials and Methods
[0038] 1.1 Test Materials
[0039] 1) Test variety: The stigma-exposed mutant line "CHE-01" of "Xiangyan No. 3".
[0040] 2) Test reagents: Paclobutrazol, Ethephon, Brassinolide, Distilled water.
[0041] 3) Test tools: Sprayer, Vernier caliper, Micrometer, Scissors, Balance, String, Petri dish, and Constant temperature and humidity light incubator.
[0042] 1.2 Test Design
[0043] 1.2.1 Inflorescence trimming method
[0044] Using CHE-01 as the material, different inflorescence trimming method treatment plots were set up for the experiment, namely no trimming (X1), only removing the central inflorescence (X2), and removing all apical inflorescences (X3). Each treatment only retained 8-10 flower branches, with a total of 3 treatments. Each plot was 1 row with 20 plants, and there were 3 replicates. Conventional fertilizer and water cultivation. After the inflorescence elongated, different trimming methods were used for different treatments to investigate the time span required for the whole plant flower organs to mature.
[0045] 1.2.2 Composite treatment of various plant hormones and shading design
[0046] Using CHE-01 as the material, the effects of different hormones and hormone levels on the development of tobacco flower organs were investigated (the treatment combinations are shown in Table 1). Specifically: For each treatment, 30 healthy tobacco plants with relatively consistent growth were selected. After the budding stage, the flower branches were trimmed. After trimming, each plant removed all apical inflorescences and retained 8-10 flower branches. Immediately after trimming, different hormones were used for spraying treatment, and shading treatment was adopted, and other agricultural operations were kept consistent.
[0047] The specific hormone spraying is as follows: At the budding stage of tobacco plants, chlormequat chloride solution is sprayed with a concentration of 500 mg / L and a spraying amount of 5 mL / plant; 3 days later, ethephon solution is sprayed with a concentration of 50 mg / L and a spraying amount of 5 mL / plant; at the initial flowering stage of tobacco plants, brassinolide solution is sprayed with a concentration of 150 mg / L and a spraying amount of 5 mL / plant, and shading treatment is carried out during the same period (pollination period). Pure water is used as the solvent for the hormone solutions.
[0048] Table 1 Compound treatment of multiple plant hormones and shading design
[0049]
[0050]
[0051] 1.3. Sample collection and measurement methods
[0052] According to the relevant requirements of "GB / T21138 - 2019", the relevant data on the quality of the seeds harvested from each experimental treatment are inspected and recorded.
[0053] Based on the experimental results obtained from the compound treatment of multiple plant hormones and shading design in 1.2.2, small - scale production tests are carried out, and the labor requirements, labor costs, average yield per mu, and output value per mu under two different experimental treatments of CK and T10 are counted to consider the difficulty of hiring workers and the labor costs.
[0054] 2. Experimental results and analysis
[0055] 2.1. Experimental results and analysis of inflorescence trimming methods
[0056] Table 2 Influence of different inflorescence trimming methods on the time span of flower organ maturity
[0057] Processing Time span (d) for the flower container to mature X1 15.3±4.2 X2 10.0±3.0 X3 6.0±2.1
[0058] As can be seen from Table 2, the minimum time span required for the flower organs of tobacco plants under treatment X3 to mature is 6 days, indicating that the tobacco plants treated by the method of the present invention can significantly reduce the time span required for different flower organs to mature.
[0059] 2.2. Experimental results and analysis of compound treatment of multiple plant hormones and shading
[0060] Table 3 Traits of tobacco flower organs under plant hormone stimulation
[0061]
[0062]
[0063] It can be found from Table 3 that when compared with the CK treatment, among the T1 to T9 treatments: for the T4 treatment, the average maturity rate at the first pollination was the highest, reaching 39.4%; for the T9 treatment, the average length of the stigma protruding from the corolla was the best, being -0.4 mm; for the T1, T2, T3, T7, T8, and T9 treatments, the pollination window period per batch was relatively consistent, all being 3 days; for the T8 treatment, the stigma exposure rate per plant was the highest, reaching 13.7%; for the T8 treatment, the fruit setting rate was the highest, reaching 92.3%.
[0064] When compared with the CK treatment, among the T10 and T11 treatments: for the T10 treatment, the average maturity rate at the first pollination was the highest, reaching 55.7%; for the T10 treatment, the average length of the corolla was the lowest, being 47.4%; for the T11 treatment, the average length of the style + ovary was the longest, being 51.2 mm, and for the T10 treatment, the average length of the stigma protruding from the corolla was the longest, being 2.5 mm; for the T10 treatment, the stigma exposure rate per plant was the highest, reaching 85.8%; for the T10 treatment, the average pollination window period was the longest, being 4 days; for the T10 treatment, the fruit setting rate was the highest, reaching 96%.
[0065] Table 4 Tobacco seed quality under different plant hormone combination treatments
[0066]
[0067]
[0068] The average number of seeds per fruit in the T10 treatment was the largest, being 3106.1 seeds (Table 4). The average weight of seeds per fruit in the T11 treatment was the heaviest, being 0.31 seeds. The 1000-seed weight in the T11 treatment was the heaviest, being 86.2 mg, and the 1000-seed weight in the T10 treatment was the second heaviest, being 85.8 mg. The seed germination rate in the T10 treatment was the highest, reaching 98.5%.
[0069] Table 5 Statistical results of relevant economic data
[0070]
[0071] As can be seen from Table 5, the number of pollinations in the T10 treatment was 2 times less than that in the CK treatment. The labor consumption per mu in the T10 treatment was 62 work units less than that in the CK treatment, saving 6200 yuan in labor cost per mu. The average yield per mu in the T10 treatment was 2.23 kg less than that in the CK treatment. The net income per mu in the T10 treatment increased by 2480 yuan compared with the CK treatment.
[0072] 3. Conclusions
[0073] The test results show that trimming the tobacco inflorescence during the budding stage of "Xiangyan No. 3", removing the apical inflorescence and retaining 8 - 10 flower branches can effectively reduce the time span required for the flower organs on the tobacco plants to develop and mature, which is beneficial for centralized pollination. After trimming the tobacco plant inflorescence, a chlormequat solution with a concentration of 500 mg / L is sprayed on the flower buds. On the third day, ethephon with a concentration of 50 mg / L is sprayed on the flower buds, and brassinolide with a concentration of 150 mg / L is sprayed at the initial flowering stage. The spraying amount of all hormones is 5 mL / plant. After spraying the brassinolide solution, shading is carried out simultaneously using a black sunshade net with a light transmittance of 65%, which can effectively extend the pollination window period, improve the seed quality, reduce the labor cost, and significantly improve the economic benefits.
[0074] The above is only the preferred specific embodiment of the present application, but the protection scope of the present application is not limited thereto. Any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present application should be covered by the protection scope of the present application. Therefore, the protection scope of the present application should be subject to the protection scope of the claims.
Claims
1. A method for improving the stigma receptivity and pollination efficiency of a tobacco stigma-exserted mutant line by using agronomic measures, characterized in that, Using the tobacco plants of the stigma-exposed mutant line as materials, they were treated by combining the inflorescence trimming method, the combined treatment with multiple plant hormones, and the shading treatment; Specifically, it includes the following steps: (1) At the budding stage of the tobacco plants, trim the inflorescence. After removing the apical inflorescence, retain 8-10 stronger branches; (2) Immediately spray the chlormequat solution after trimming the inflorescence, and spray the ethephon solution 3 days later; (3) Spray the brassinolide solution at the initial flowering stage of the tobacco plants, and conduct shading treatment during the same period; The concentration of the chlormequat solution is 500 mg / L, and the spraying amount is 5 mL / plant; The concentration of the ethephon solution is 50 mg / L, and the spraying amount is 5 mL / plant; The concentration of the brassinolide solution is 150 mg / L, and the spraying amount is 5 mL / plant; The shading treatment refers to shading with a black sunshade net with a light transmittance of 65%.