Microspore culture medium suitable for portable culture tank and application of microspore culture medium

Abstract

The invention discloses a microspore culture medium suitable for a portable culture tank and application of the microspore culture medium. The microspore culture medium is prepared from the following specific components: 15.0 to 18.0 mg / L of ammonium nitrate, 0 to 505.5 mg / L of potassium phosphate, 300.0 to 472.3 mg / L of calcium nitrate, 0.2 to 1.0 mg / L of boric acid, 400.0 to 550.0 mg / L of magnesium sulfate, 5.0 to 13.7 mg / L of potassium phosphate, 15.0 to 20.0 mg / L of NaFe. EDTA (Ethylene Diamine Tetraacetic Acid), 0.2 to 1.5 mg / L of manganese chloride, 0 to 1.44 mg / L of zinc sulfate, 0.01 to 0.30 mg / L of copper sulfate pentahydrate, 0.01 to 0.05 mg / L of sodium molybdate, 50.0 to 150.0 mg / L of sodium silicate and 0.1 to 10.5 mg / L of agar. The preservative comprises the following components: 0.01 to 3.0 mg / L of gelatin, 0.01 to 3.0 mg / L of xanthan gum, 10.0 to 30.0 mg / L of cane sugar, 0.01 to 0.2 g / L of potassium sorbate, 0.01 to 0.2 g / L of calcium lactate or / and calcium citrate, 0 to 0.1 g / L of zinc phosphate, 0 to 1.5 g / L of wormwood extract and the balance of sterile water. The culture medium is high in environment adaptability and convenient to move and transport, microspore culture can be achieved in a non-sterile environment, and science popularization of a microspore culture technology can be completed more completely.

Description

Technical Field

[0001] The present invention belongs to the technical field of microspore culture, and particularly relates to a microspore culture medium applicable to a portable culture tank and its application. Background Art

[0002] Generally, microspore culture is carried out in a sterile environment. Existing microspore culture media are not suitable for popular science experiments. The main problems are as follows:

[0003] 1. During the culture process of microspores, different culture media are required at different growth stages to ensure that microspores reach the optimal growth conditions and guarantee the accuracy of scientific research results. There are problems such as large volume of the culture medium, high requirements for culture technology, and high requirements for the environment.

[0004] 2. Microspore culture in a laboratory environment does not need to consider issues such as anti-breakage, weight, and volume of the colloid during transportation.

[0005] 3. During the scientific research process, it is usually small-scale experiments, and there is no need to consider energy conservation, weight loss, cost and environmental problems brought about by reducing fertilizer use.

[0006] 4. In the existing scientific research process, only specific growth stages of plants are studied, and the lack of harvest is too boring, which is not conducive to popular science promotion.

[0007] How to obtain a culture medium applicable to scenarios with bacteria and insects is of great significance for promoting the popularization of microspore culture. Summary of the Invention

[0008] The purpose of the present invention is to provide a microspore culture medium applicable to a portable culture tank and its application. This culture medium has strong environmental adaptability and is convenient for mobile transportation. It can achieve microspore culture in a non-sterile environment, and can more conveniently and completely complete the popular science publicity of microspore cultivation technology.

[0009] To achieve the above object, the technical solution of the present invention is as follows:

[0010] A microspore culture medium suitable for use in a portable culture tank, with specific components as follows: ammonium nitrate: 15.0 - 18.0 mg / L, potassium phosphate: 0 - 505.5 mg / L, calcium nitrate: 300.0 - 472.3 mg / L, boric acid: 0.2 - 1.0 mg / L, magnesium sulfate: 400.0 - 550.0 mg / L, potassium phosphate: 5.0 - 13.7 mg / L, NaFe·EDTA: 15.0 - 20.0 mg / L, manganese chloride: 0.2 - 1.5 mg / L, zinc sulfate: 0 - 1.44 mg / L, copper sulfate pentahydrate: 0.01 - 0.30 mg / L, sodium molybdate: 0.01 - 0.05 mg / L, sodium silicate: 50.0 - 150.0 mg / L, agar: 0.1 - 10.5 mg / L; gelatin: 0.01 - 3.0 mg / L, xanthan gum: 0.01 - 3.0 mg / L, sucrose: 10.0 - 30.0 mg / L, potassium sorbate: 0.01 - 0.2 g / L, calcium lactate or / and calcium citrate 0.01 - 0.2 g / L, zinc phosphate: 0 - 0.1 g / L, wormwood extract: 0 - 1.5 g / L, with the balance being sterile water. Preferably, the pH of the culture medium is 5.0 - 6.5.

[0011] The application of the microspore culture medium described in the present invention in microspore culture, and the culture environment is a non-sterile environment.

[0012] Preferably, during the culture process of microspores developing into seedlings, no seedling transplantation is required, and the obtained seedlings are transplanted into soil containing Hoagland's nutrient solution for cultivation. The soil components include: 10 - 40% coconut coir, 20 - 50% sand, and 10 - 30% perlite.

[0013] The use scenario of the culture medium described in the present invention is an environment with bacteria and insects such as at home. To enhance the survival rate of microspores in a complex environment, increase their shelf life, increase the popular science duration and effect, substances such as potassium sorbate, calcium citrate (or calcium lactate), and wormwood extract are added to the culture medium formula to achieve the effects of anti-corrosion and antibacterial, and insect repellent. At the same time, the additional preservatives, antibacterial agents, and natural insect repellent components contain nutrients such as nitrogen, phosphorus, potassium, calcium, and sugars, which have the effect of increasing fertility and can reduce the amount of fertilizer in the original culture medium formula.

[0014] According to the conventional operation, when microspores develop into cell clusters, plant tissues, young plants, seedlings and other stages, they need to be transplanted into special culture media corresponding to different stages to improve the survival rate and achieve the purpose of not affecting the experimental results. However, since the general public does not have the sterile environment and technical ability for transplanting seedlings during the planting process, therefore, the present invention meets the nutritional needs of the whole-stage development of microspores through nutrients such as nitrogen, phosphorus, potassium, calcium, and sugar added to the culture medium. All the nutrients required from the microspore stage to the seedling stage are already included in the culture medium. During the microspore to seedling stage, there is no need for later transplanting, fertilizing, or watering. Only after the seedlings have their own immune ability, they are transplanted into the soil to ensure their subsequent growth.

[0015] The present invention adds gelatin and xanthan gum to the culture medium, which enhances the structural strength of the culture medium and its ability to withstand transportation, preventing the colloid from breaking during transportation. And xanthan gum has a strong water retention effect, which can reduce the need for subsequent watering and reduce the pollution of the culture medium by exogenous bacteria.

[0016] The pH value of the culture medium described in the present invention is controlled at 5.0 - 6.5, which is different from the neutral pH value of ordinary culture media. The purpose is to reduce the intrusion of miscellaneous bacteria, improve the survival probability of microspores, and extend the popular science time.

[0017] After the microspores grow into seedlings, the seedlings already have a certain degree of self-immune resistance. The lid of the culture medium can be opened to transplant the seedlings into the soil with a pre-prepared stepped fertility. The seedlings grow in the soil from the seedling stage to the mature stage. The specific soil composition is: 10 - 40% coconut fiber, 20 - 50% sandy soil, 10 - 30% perlite, and fertilizers configured in stepped proportions according to experimental needs: 0%, 25%, 50%, 75%, 100% HoagLand's nutrient solution. Continue to observe the growth of the seedlings in the soil with different fertilities, which is convenient for observing the differences in the growth ability of seedlings obtained from microspore culture and seedlings obtained from seed culture and the growth of seedlings under different fertilization amounts, and increasing the content and interest of popular science from different perspectives of comparison.

[0018] Compared with the prior art, the beneficial effects of the present invention are:

[0019] The formula features and highlights of the culture medium of the present invention lie in adding potassium sorbate, calcium citrate (or calcium lactate), wormwood extract, etc. to the culture medium for anti-corrosion, antibacterial, and insect repellent purposes. At the same time, the additional preservatives, antibacterial agents, and natural insect repellent components contain nutrients such as nitrogen, phosphorus, potassium, calcium, and sugars, which have the effect of increasing fat content and can reduce the amount of fertilizer in the original culture medium formula. Through the new adjustment and compounding of nutrient components, the entire growth stage of microspores growing into seedlings can proceed smoothly in this culture medium, completing the observation and recording of the entire life cycle of asexually propagated plants, and can more conveniently and completely complete the popular science publicity of microspores, culture media, and microspore cultivation techniques.

[0020] Potassium sorbate, calcium citrate (or calcium lactate), wormwood extract, gelatin, xanthan gum, etc. are added to the culture medium of the present invention for anti-corrosion, antibacterial, insect repellent, increasing shelf life, increasing the duration of popular science, and enhancing transportation resistance. Generally, microspore culture is carried out in a sterile environment and is not easy to transport. This formula greatly solves these problems. Brief Description of the Drawings

[0021] Figure 1 It is a photograph taken after culturing the plant tissue of Example 1 of the present invention in the culture medium for one week;

[0022] Figure 2 It is a photograph taken after culturing the plant tissue of Example 1 of the present invention in the culture medium for two weeks;

[0023] Figure 3 It is a photograph taken after culturing the plant tissue of Example 1 of the present invention in the culture medium for three weeks;

[0024] Figure 4 It is a photograph taken after culturing the plant tissue of Example 1 of the present invention in the culture medium for four weeks;

[0025] Figure 5 Photographs taken after transporting the culture medium of Comparative Example 1;

[0026] Figure 6 Photographs taken after transporting the culture medium of Comparative Example 1;

[0027] Figure 7 Photographs taken after transporting the culture medium of Comparative Example 1;

[0028] Figure 8 Photographs taken after culturing the plant tissue of Comparative Example 2 in the culture medium for three weeks;

[0029] Figure 9 Photographs taken after culturing the plant tissue of Comparative Example 3 in the culture medium for four weeks. Detailed Description of the Invention

[0030] The present invention will be further explained and illustrated below in conjunction with specific embodiments and examples.

[0031] Example 1

[0032] The specific formula of the culture medium is as follows: ammonium nitrate 16.5 mg / L; potassium phosphate 105.5 mg / L; calcium nitrate 372.3 mg / L; boric acid 0.62 mg / L; magnesium sulfate 492.9 mg / L; potassium phosphate 5.7 mg / L; NaFe·EDTA 18.35 mg / L; manganese chloride 0.99 mg / L; zinc sulfate 0.44 mg / L; copper sulfate pentahydrate 0.12 mg / L; sodium molybdate 0.025 mg / L; sodium silicate 106.1 mg / L; agar 6.0 mg / L; gelatin 1.0 mg / L; xanthan gum 1.0 mg / L; sucrose 12.0 mg / L; potassium sorbate 0.1 g / L; calcium citrate 0.1 g / L; zinc phosphate 0.05 g / L; wormwood extract 0.5 g / L; sterile water 1 L.

[0033] Cut the tissue of barley anthers after reverse differentiation treatment and place it in the culture medium for cultivation. After one month of cultivation, it grows into seedlings, and then the seedlings are transplanted into the soil and continue to be planted in the soil until the mature stage.

[0034] The soil components are: 20% coconut coir, 50% sandy soil, 30% perlite, and fertilizers configured in a stepped ratio according to experimental needs: 0%, 25%, 50%, 75%, 100% HoagLand's nutrient solution.

[0035] During this period, continuous photos are taken for recording. Figures 1 to 4 Photos of the plant tissue in this example taken after one week, two weeks, three weeks, and four weeks of cultivation in the culture medium. It can be found from the photos that the tissue develops well in the culture medium and forms complete seedlings.

[0036] Example 2

[0037] The specific formula of the culture medium is as follows: ammonium nitrate 18.0 mg / L; potassium phosphate 405.5 mg / L; calcium nitrate 372.3 mg / L; boric acid 0.62 mg / L; magnesium sulfate 492.9 mg / L; potassium phosphate 7.7 mg / L; NaFe·EDTA 18.35 mg / L; manganese chloride 0.99 mg / L; zinc sulfate 1.14 mg / L; copper sulfate pentahydrate 0.12 mg / L; sodium molybdate 0.025 mg / L; sodium silicate 106.1 mg / L; agar 6.0 mg / L; gelatin 1.5 mg / L; xanthan gum 1.5 mg / L; sucrose 20.0 mg / L; potassium sorbate 0.1 g / L; calcium citrate 0.1 g / L; zinc phosphate 0.05 g / L; wormwood extract 1.0 g / L; sterile water 1 L.

[0038] The tissue of barley anthers after reverse differentiation treatment was excised and placed in a culture medium for cultivation. After one month of cultivation, it grew into seedlings, and then the seedlings were transplanted into the soil and continuously planted in the soil until the mature stage.

[0039] The soil components were: 40% coconut coir, 30% sandy soil, 30% perlite, and fertilizers configured in a stepped ratio according to experimental needs: 0%, 25%, 50%, 75%, 100% HoagLand's nutrient solution.

[0040] Example 3

[0041] The specific formula of the culture medium was as follows: ammonium nitrate 15.5 mg / L; potassium phosphate 405.5 mg / L; calcium nitrate 342.3 mg / L; boric acid 0.62 mg / L; magnesium sulfate 492.9 mg / L; potassium phosphate 10.7 mg / L; NaFe·EDTA 18.35 mg / L; manganese chloride 0.99 mg / L; zinc sulfate 1.04 mg / L; copper sulfate pentahydrate 0.12 mg / L; sodium molybdate 0.025 mg / L; sodium silicate 106.1 mg / L; agar 6.0 mg / L; gelatin 1.0 mg / L; xanthan gum 2.0 mg / L; sucrose 15.0 mg / L; potassium sorbate 0.15 g / L; calcium lactate 0.15 g / L; zinc phosphate 0.05 g / L; wormwood extract 1.05 g / L; sterile water 1 L.

[0042] The tissue of barley anthers after reverse differentiation treatment was excised and placed in a culture medium for cultivation. After one month of cultivation, it grew into seedlings, and then the seedlings were transplanted into the soil and continuously planted in the soil until the mature stage.

[0043] The soil components were: 30% coconut coir, 40% sandy soil, 30% perlite, and fertilizers configured in a stepped ratio according to experimental needs: 0%, 25%, 50%, 75%, 100% HoagLand's nutrient solution.

[0044] Comparative Example 1

[0045] Compared with Example 1 of the present invention, the difference in the culture medium components was that gelatin and xanthan gum were not added. The formed culture medium was placed in a container, and the changes in the culture medium after transportation were as Figure 5 、 Figure 6 、 Figure 7 shown. It could be seen from the figure that the culture medium was severely broken and could not be used for the cultivation of plant tissues.

[0046] It was explained that the gel components added to the culture medium were inaccurate and the strength was insufficient, which would cause the gel to break and the microspores could not be cultured. And the conventional method of adding carrageenan to increase the strength would make the culture medium too hard and was not conducive to the growth of plant roots.

[0047] Comparative Example 2

[0048] The difference from Example 1 of the present invention is that wormwood extract is not added to the culture medium.

[0049] Figure 8 This is a photo taken after the plant tissue was cultured in the culture medium for three weeks. From Figure 8 it can be seen that mold appeared during the tissue culture process.

[0050] Comparative Example 3

[0051] The difference from Example 1 of the present invention is that potassium sorbate and calcium citrate (or calcium lactate) are not added to the culture medium.

[0052] Figure 9 This is a photo taken after the plant tissue was cultured in the culture medium for four weeks. From Figure 9 it can be seen that mold appeared in the culture medium during the tissue culture process.

Claims

1. A microspore culture medium suitable for use in a portable culture tank, with the specific components being: ammonium nitrate: 15.0 - 18.0 mg / L, potassium phosphate: 0 - 505.5 mg / L, calcium nitrate: 300.0 - 472.3 mg / L, boric acid: 0.2 - 1.0 mg / L, magnesium sulfate: 400.0 - 550.0 mg / L, potassium phosphate: 5.0 - 13.7 mg / L, NaFe·EDTA: 15.0 - 20.0 mg / L, manganese chloride: 0.2 - 1.5 mg / L, zinc sulfate: 0 - 1.44 mg / L, copper sulfate pentahydrate: 0.01 - 0.30 mg / L, sodium molybdate: 0.01 - 0.05 mg / L, sodium silicate: 50.0 - 150.0 mg / L, agar: 0.1 - 10.5 mg / L; gelatin: 0.01 - 3.0 mg / L, xanthan gum: 0.01 - 3.0 mg / L, sucrose: 10.0 - 30.0 mg / L, potassium sorbate: 0.01 - 0.2 g / L, calcium lactate or / and calcium citrate 0.01 - 0.2 g / L, zinc phosphate: 0 - 0.1 g / L, wormwood extract: 0 - 1.5 g / L, and the balance being sterile water.

2. The microspore culture medium applicable to a portable culture tank as described in claim 1, wherein The pH of the culture medium is 5.0 - 6.

5.

3. Use of the microspore culture medium according to claim 1 or 2 in microspore culture, characterized in that, The culture environment is a non-sterile environment.

4. Use of the microspore culture medium according to claim 3 in microspore culture, characterized in that, During the culture process of microspores developing into seedlings, no seedling transplantation is required. The obtained seedlings are transplanted into soil containing HoagLand's nutrient solution for cultivation. The soil components include: 10 - 40% coconut fiber, 20 - 50% sand, and 10 - 30% perlite.

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