Efficient separation method for heterotypic seeds in soil seed bank of acorus comosus

Abstract

The invention discloses an efficient separation method for heterotypic seeds in a soil seed bank of acorus comosus, and belongs to the technical field of plant seed separation. The high-efficiency separation method of the heterotypic seeds in the soil seed bank of the heterotypic seeds comprises the following steps: collecting surface soil near a heterotypic plant to obtain a soil seed bank sample; separating and removing impurity components which can be identified by naked eyes and are not the seeds of the isaria heterophylla in the soil seed bank sample, then screening through a screen with the pore diameter of 400 meshes to obtain an unscreened part, and adding calcium sulfate to obtain a to-be-separated component; adding water into the to-be-separated component, stirring to obtain a turbid liquid, and then filtering through a screen with the pore diameter of 600 meshes to obtain a mixture containing the heterotypic seeds; and adding the mixture containing the heterotypic seeds into a 45wt% ethanol solution, and separating the heterotypic seeds by using different densities to complete the separation of the heterotypic seeds.

Description

Technical Field

[0001] The present invention relates to the technical field of plant seed separation, and particularly to an efficient separation method for dimorphic seeds in the soil seed bank of Suaeda aralocaspica. Background Art

[0002] In multiple fields such as ecological restoration, agricultural planting, and seed science research, efficiently and accurately separating seeds from complex soil seed banks is a crucial task. The soil seed bank contains rich seed resources, and these seeds can germinate and grow into new plants under suitable conditions, thereby participating in the construction and maintenance of the ecosystem. As an important basis for plant population renewal and vegetation restoration, the separation efficiency and purity of the seeds inside the soil seed bank will directly affect the effects of subsequent research and applications. However, in reality, the complexity of the soil environment poses a huge challenge to seed separation. Especially when the soil salinity is too high or the physical structure is poor, the separation efficiency and germination rate of seeds will be severely affected. Especially in saline-alkali soil, which often contains a large amount of sodium ions (Na + +), it will destroy the stability of soil colloids, cause soil particles to disperse and the ground to harden, and further deteriorate the air permeability and water permeability of the soil, which poses a great obstacle to seed germination and the growth of plant roots. How to efficiently and accurately separate seeds, especially dimorphic seeds, from complex soil seed banks is a technical problem currently faced.

[0003] Suaeda aralocaspica, as an annual herb with special ecological value, is mainly distributed in the saline soil environment of the Central Asian desert region and is only distributed in northern Xinjiang, China. Suaeda aralocaspica has two different morphological seeds: brown seeds (density of 0.45 g / ml) and black seeds (density of 0.38 g / ml). The brown seeds are round large fruits with a relatively high germination rate and can quickly form a temporary soil seed bank; while the black seeds are oval small fruits with a smooth surface, have non-deep physiological dormancy characteristics, and have a relatively low germination rate, but can be stored in the soil for a long time to form a persistent soil seed bank.

[0004] However, due to the differences in shape, size, texture, and germination characteristics between these two types of heteromorphic seeds, combined with the complexity of the soil environment, various factors such as various impurities, microorganisms, and soil particles of different properties will interfere. Existing traditional separation methods still have many problems when dealing with heteromorphic seeds in the soil seed bank of Suaeda heteroptera. For example, traditional methods (such as water washing and filtration method, manual selection method, etc.) are often inefficient and require a large amount of human and time costs. At the same time, the separation accuracy is insufficient, and it is difficult to accurately separate heteromorphic seeds from soil samples, resulting in low purity of the separated seeds, which will further affect the subsequent research and utilization of heteromorphic seeds of Suaeda heteroptera, such as inaccurate seed quantity statistics. In addition, when facing special soil environments such as salinization, the limitations of these methods are more prominent, and it is difficult to effectively cope with the adverse effects of high salt content and poor physical structure in the soil on seed separation. Therefore, there is an urgent need for a new, efficient, and accurate separation method to solve these problems. Summary of the Invention

[0005] The purpose of the present invention is to provide an efficient separation method for heteromorphic seeds in the soil seed bank of Suaeda heteroptera to solve the above problems in the background technology. The present invention uses calcium sulfate to improve the soil environment to efficiently separate heteromorphic seeds in the soil seed bank of Suaeda heteroptera, thereby solving the problems of low efficiency and insufficient separation accuracy existing in traditional separation methods, and improving the purity of seed separation and the accuracy of quantity statistics.

[0006] To achieve the above purpose, the present invention provides the following technical solutions:

[0007] One of the technical solutions of the present invention: Provide an efficient separation method for heteromorphic seeds in the soil seed bank of Suaeda heteroptera, including the following steps:

[0008] Collect the surface soil near the Suaeda heteroptera plants to obtain a soil seed bank sample;

[0009] Separate and remove the impurity components that can be visually recognized and are not Suaeda heteroptera seeds in the soil seed bank sample, and then screen through a sieve with a pore size of 400 mesh to obtain the unsieved part, add calcium sulfate to obtain the component to be separated;

[0010] Add water to the component to be separated and stir to obtain a suspension, and then filter through a sieve with a pore size of 600 mesh to obtain a mixture containing heteromorphic seeds;

[0011] Add the mixture containing heteromorphic seeds to the ethanol solution for density separation, and use different densities to separate heteromorphic seeds to complete the separation of heteromorphic seeds.

[0012] Further, the collection of the surface soil near the Suaeda heteroptera plants is to collect the surface soil within 1 m of the Suaeda heteroptera plants.

[0013] Furthermore, the surface soil is soil with a depth from the soil surface to 10-20 cm underground.

[0014] Furthermore, the added amount of calcium sulfate is 0.2wt% of the mass of the unscreened portion.

[0015] Furthermore, the stirring time is 5-10s.

[0016] Furthermore, the ratio of the volume of water added to the suspension to the mass of the components to be separated is 4L:1kg.

[0017] Furthermore, the concentration of the ethanol solution for density separation is 45wt%.

[0018] Furthermore, the efficient separation method is completed within 20 minutes; and the separation of heteromorphic seeds using different densities is completed within 5 minutes.

[0019] Furthermore, after separating the heteromorphic seeds by using different densities, the method further comprises the step of adding water to rinse the product system three times while avoiding damaging the seeds.

[0020] The beneficial technical effects of the present invention are as follows:

[0021] The present invention provides a highly efficient method for separating heteromorphic seeds in a soil seed bank of Phragmites australis, which can quickly and accurately separate heteromorphic seeds from the soil seed bank. The implementation of the method has significant technical advantages, and is particularly suitable for ecological, botanical and agricultural research that requires efficient separation and extraction of heteromorphic seeds of Phragmites australis, and provides strong support for the management and application of seed banks. The present invention significantly shortens the separation time, and the seeds separated by the separation method of the present invention have less mechanical damage and higher vitality, which is beneficial to subsequent seed germination experiments, and can be used to separate heteromorphic seeds in a soil seed bank of Phragmites australis.

[0022] The present invention uses calcium sulfate to improve the soil environment to efficiently separate the heteromorphic seeds in the soil seed bank of P. sphaerocephala, thereby solving the problems of low efficiency and insufficient separation accuracy in traditional separation methods, and improving the purity of seed separation and the accuracy of quantitative statistics. The present invention is expected to provide strong technical support and guarantee for the fields of ecological restoration, agricultural planting and seed science research, promote the process of plant population renewal and vegetation restoration, and promote the healthy development of the ecosystem. DETAILED DESCRIPTION

[0023] Now, various exemplary embodiments of the present invention are described in detail, and this detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present invention. 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.

[0024] In addition, 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. Intermediate values within any stated value or stated range, as well as each smaller range between any other stated value or intermediate values within the stated range, are also included in the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0025] 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. It should be noted that the aspects not described in detail in the present invention are all conventional operating means in the art and are not the focus of the present invention.

[0026] Regarding the use of "comprising", "including", "having", "containing", etc. in the present invention, they are all open-ended terms, meaning including but not limited to.

[0027] The soil seed bank refers to the seed storage repository naturally existing in the soil and is an important basis for plant population renewal and vegetation restoration. Usually, a large number of seeds exist at different depths in the soil, including normal seeds and heteromorphic seeds. Due to the particularity of heteromorphic seeds in terms of shape, size, texture, etc., many challenges are faced during the separation process. Traditional seed separation methods, such as the water washing and filtering method, the manual selection method, etc., expose many limitations when dealing with plants with heteromorphic seeds such as Suaeda heteroptera. For example, although the water washing and filtering method can remove some impurities, it is difficult to completely remove the soil particles tightly adhered to the heteromorphic seeds, and when the operation is improper, it may cause damage to some fragile seeds, thereby affecting the germination ability and subsequent utilization value of the seeds. The manual selection method is time-consuming, laborious, inefficient, and greatly affected by human factors, and it is difficult to guarantee the separation accuracy. In addition, the limitations of these methods are more prominent when facing special soil environments such as salinization, and it is difficult to effectively cope with the adverse effects of high salt content and poor physical structure in the soil on seed separation, and further improvement is still needed.

[0028] The present invention designs a method for improving the soil environment by using calcium sulfate to efficiently separate heteromorphic seeds from the soil seed bank of Suaeda heteroptera. Due to factors such as the special morphology of heteromorphic seeds in the soil seed bank of Suaeda heteroptera and the complex soil environment, traditional separation methods have problems such as low efficiency and insufficient separation accuracy. The present invention aims to solve these problems and provide a targeted and efficient separation scheme. The method involves obtaining soil samples with a core sampler, removing preliminary impurities, air-drying and sieving, adding water to the remaining soil to stir into a suspension, and adding calcium sulfate to adjust the soil salinity and improve the soil structure. Filtering is carried out with a nylon mesh bag of a specific pore size. During washing, 45wt% ethanol solution is used to separate the heteromorphic seeds of Suaeda heteroptera, and finally the seeds are obtained by drying. Compared with traditional methods, the present invention has significant advantages of high efficiency and precision, can effectively shorten the separation time, improve the purity of seed separation and the accuracy of quantity statistics, and at the same time change the chemical properties and physical characteristics of the soil, and has important application value in the research and utilization of heteromorphic seeds in the soil seed bank of Suaeda heteroptera.

[0029] The present invention discloses a method for efficiently separating heteromorphic seeds from the soil seed bank of Suaeda heteroptera, comprising the following steps:

[0030] Within a range of 1 m around the Suaeda heteroptera plants, soil samples with a depth from the soil surface to 10 - 20 cm underground are collected using a core sampler with a diameter of 20 cm and a height of 15 cm to obtain soil seed bank samples;

[0031] The large soil clods in the collected soil seed bank samples are crushed, and impurity components such as plant roots, fallen leaves, and stones that are visible to the naked eye and are not Suaeda heteroptera seeds are picked out, placed indoors for natural air-drying, and then sieved through a sieve with a mesh diameter of 600 meshes. The sieved components are discarded, and the unsieved part is retained; 2 g of calcium sulfate is added to each kilogram of the unsieved part to obtain the component to be separated;

[0032] The component to be separated is placed in a container, water is added so that the water surface is 0.5 - 1 cm higher than the soil surface, and then it is fully stirred for 5 - 10 s to obtain a suspension, and then filtered through a 600-mesh sieve to obtain a mixture containing heteromorphic seeds;

[0033] The mixture containing heteromorphic seeds is added to a 45wt% ethanol solution. Since the densities of the heteromorphic seeds of Suaeda heteroptera and other impurities in the 45wt% ethanol solution are different, the brown seeds will sink to the bottom and the black seeds will float on the surface. Therefore, different-shaped seeds are fished out and separated using a filter screen, and then slowly rinsed to avoid damaging the seeds, and dried at room temperature to obtain heteromorphic seed samples of different shapes.

[0034] Further, for areas where there may be more heteromorphic seeds (dense plant areas, complex terrain areas), the sampling density is appropriately increased (increasing the number of sampling points or collecting more soil samples at each sampling point).

[0035] Further, during the stirring process, control the stirring speed and time to prevent the seeds from overflowing with the bubbles or the seeds from germinating later due to excessive soaking time.

[0036] Further, the ratio of the volume of water added to the suspension to the mass of the component to be separated is 4 L:1 kg.

[0037] Further, the high-efficiency separation method is completed within 20 minutes.

[0038] In the present invention, "room temperature" is calculated as 10 - 30 °C unless otherwise specified.

[0039] All raw materials used in the following examples and comparative examples of the present invention are commercially available products.

[0040] Example 1

[0041] A high-efficiency separation method for heteromorphic seeds in the soil seed bank of Suaeda heteroptera, the specific steps are as follows:

[0042] I. In August 2023, collect the soil seed bank beside the Bayi Reservoir in Fukang City, Xinjiang Uygur Autonomous Region. Use a circular knife with a diameter of 20 cm to collect the soil with a depth from the soil surface to 10 cm underground within a range of 1 m around the Suaeda heteroptera plants, and the area is 314 cm 2 as a sample unit.

[0043] II. Crush the large soil blocks in the collected sample unit, pick out large plant roots, fallen leaves, stones and other impurity components that can be recognized by the naked eye and are not Suaeda heteroptera seeds. After natural air drying, sieve through a sieve with a mesh diameter of 400 meshes, retain the part that has not passed through the sieve, measure its pH value to be 8, and the salt content to be 5.3%. Then add 0.2 wt% of calcium sulfate equivalent to the raw material components to obtain the component to be separated.

[0044] III. Put the above-mentioned component to be separated into a large basin, add water so that the water surface is 0.6 cm higher than the soil surface, and then stir for 5 s to obtain a suspension (the ratio of the volume of water added to the suspension to the mass of the component to be separated is 4 L:1 kg). Filter the suspension through a 600-mesh sieve to obtain the mixture of the part that has not passed through the sieve.

[0045] IV. Add the mixture containing heteromorphic seeds obtained in step III to a 45 wt% ethanol solution. The black seeds float on the water surface, and the brown seeds sink to the bottom. Scoop them out separately, then rinse with clean water 3 times, and then use filter paper to absorb the water on the surface of the seeds to obtain the heteromorphic seed sample.

[0046] V. Verification results: The purity of the heteromorphic seeds separated by the method of this embodiment is high. The number and type of species identified are consistent with the actual situation. The separation time is only within 20 minutes, and the efficiency is much higher than the traditional method.

[0047] Example 2

[0048] The only difference from Example 1 is that step 1 is modified to:

[0049] In August 2023, soil seed banks were collected near the Bayi Reservoir in Fukang City, Xinjiang Uygur Autonomous Region. A circular knife was used to collect 5 areas with an area of ​​30 cm in depth from the soil surface to 8 cm underground within 1 m around the plants of P. 2 of soil units, mixed as sample units;

[0050] And modify the amount of water added in step three so that the water level is 0.8 cm above the soil surface.

[0051] Verification results: It is also able to efficiently and accurately separate heteromorphic seeds, and performs well in terms of separation efficiency and purity.

[0052] Example 3

[0053] The only difference from Example 1 is that step 1 is modified to:

[0054] Soil seed banks were collected in moderately to severely salinized soils. A 20-cm-diameter circular knife was used to collect seeds from the soil surface to 8 cm underground within 1 m around the plants of Pleurotus eryngii. The area was 314 cm 2 of soil as the sample unit.

[0055] Example 4

[0056] The only difference from Example 1 is that in step 2, after passing through a sieve with a mesh diameter of 100 meshes, the sieved part is removed, and the unsieved part is retained before subsequent operations are performed.

[0057] Example 5

[0058] The only difference from Example 1 is that the amount of water added in step 3 is modified to add water so that the water surface is 2 cm above the soil surface.

[0059] Example 6

[0060] The only difference from Example 1 is that step 3 is modified to:

[0061] Put the above-mentioned components to be separated into a large basin, add water so that the water surface is 0.6 cm higher than the soil surface, and then stir for 5 s to obtain a suspension (the ratio of the volume of water added to the mass of the components to be separated in the suspension is 4 L:1 kg). Then wait for the seeds to float to the water surface, and use a nylon mesh bag with a pore diameter of 0.2 mm to filter the upper-layer suspension to obtain the filtered mixture.

[0062] Example 7

[0063] The difference from Example 1 is only that drying the water on the seed surface with filter paper in Step 4 is modified to drying at room temperature with a hair dryer.

[0064] Example 8

[0065] The difference from Example 1 is only that Step 4 is modified to directly drying the filtered mixture at room temperature with a hair dryer, and then picking out the seeds.

[0066] Test Example 1

[0067] An efficient separation method for heteromorphic seeds in the soil seed bank of Suaeda heteroptera, the specific steps are as follows:

[0068] I. Collect the soil seed bank of Suaeda heteroptera in the Shawan area. Collect the soil with a depth from the soil surface to 15 cm underground and an area of 314 cm 2 around the Suaeda heteroptera plants within a range of 1 m as a sample unit.

[0069] II. Crush the large soil clods in the collected sample unit, pick out the large plant rhizomes, fallen leaves, stones and other impurities that can be recognized by the naked eye and are not Suaeda heteroptera seeds. After natural air drying, sieve through a sieve with a mesh diameter of 400 meshes, retain the part that has not passed through the sieve, measure its pH value to be 7.7 and the salt content to be 5.7%, and then add 0.2 wt% of calcium sulfate equivalent to the raw material components to obtain the components to be separated.

[0070] III. Put the above-mentioned components to be separated into a container, add water so that the water surface is 1 cm higher than the soil surface, and then stir until the soil sample in the container becomes a suspension (the ratio of the volume of water added to the mass of the components to be separated in the suspension is 4 L:1 kg), and filter with a 600-mesh sieve to obtain the filtered mixture.

[0071] IV. Put the filtered mixture obtained in Step III into a 45 wt% ethanol solution. After the seeds are separated, use a sieve to fish them out respectively, rinse with water 3 times, and dry at room temperature with a hair dryer to obtain the seed sample.

[0072] Using the separation method of this test example, the separation time is 15 min, including 15 black seeds and no brown seeds.

[0073] Comparative Example 1 (conventional method)

[0074] The only difference from Experimental Example 1 is that Step 3 and Step 4 are modified as follows:

[0075] 3. Filter the components to be separated obtained in step 2 through a 400-mesh sieve to obtain a filtered mixture.

[0076] 4. Manually pick out the seeds from the filtered mixture in step 3.

[0077] The separation method of Comparative Example 1 was used, and the separation time was 1 h, including 8 black seeds and no brown seeds.

[0078] Comparative Example 2 (conventional method)

[0079] The only difference from Experimental Example 1 is that after obtaining the sample unit, the following processing is performed:

[0080] Second, crush the large clods of soil in the collected sample units, pick out large plant rhizomes, fallen leaves, stones and other impurities that are identifiable to the naked eye and are not seeds of the Pleurotus eryngii, and then air-dry them naturally to obtain the components to be separated.

[0081] 3. Manually pick out the seeds from the components to be separated in step 2.

[0082] The separation method of Comparative Example 2 was used, and the separation time was 1.5 h, including 5 black seeds and no brown seeds.

[0083] From the above content, it can be seen that by adopting the separation method of the present invention, the separation time can be shortened to 15 minutes. Compared with 1 hour of the conventional sieving method (Comparative Example 1) and 1.5 hours of the manual selection method (Comparative Example 2), the present invention significantly shortens the separation time. In addition, the seeds separated by the separation method of the present invention have less mechanical damage and higher vitality, which is beneficial to the subsequent seed germination test and can be used to separate heteromorphic seeds in the soil seed bank of Pleurotus eryngii.

[0084] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. An efficient method for separating heteromorphic seeds in a soil seed bank of Phragmites australis, characterized in that: The following steps are involved: Collect the surface soil near the plants of Pleurotus eryngii to obtain soil seed bank samples; Separating and removing impurity components that are identifiable by naked eyes and are not seeds of Phragmites australis in the soil seed bank sample, and then sieving through a sieve with an aperture of 400 meshes to obtain an unsieved portion, adding calcium sulfate to obtain a component to be separated; Adding water to the components to be separated and stirring to obtain a suspension, and then filtering through a sieve with a pore size of 600 mesh to obtain a mixture containing heteromorphic seeds; The mixture containing the heteromorphic seeds is added into an ethanol solution for density separation, and the heteromorphic seeds are separated by using different densities to complete the separation of the heteromorphic seeds.

2. The efficient separation method according to claim 1, characterized in that: The surface soil near the collected spatholobus plants is the surface soil within 1 m of the collected spatholobus plants.

3. The efficient separation method according to claim 1, characterized in that: The surface soil is the soil with a depth from the soil surface to 10-20 cm underground.

4. The efficient separation method according to claim 1, characterized in that: The added amount of calcium sulfate is 0.2 wt% of the mass of the unscreened portion.

5. The efficient separation method according to claim 1, characterized in that: The stirring time is 5-10s.

6. The efficient separation method according to claim 1, characterized in that: The ratio of the volume of water added to the suspension to the mass of the components to be separated is 4L:1kg.

7. The efficient separation method according to claim 1, characterized in that: The concentration of the separation ethanol solution is 45 wt %.

8. The efficient separation method according to claim 1, characterized in that: The highly efficient separation method was completed within 20 min.

9. The efficient separation method according to claim 1, characterized in that: After separating the heteromorphic seeds by using different densities, the method further includes the step of adding water to rinse the product system three times without damaging the seeds.