A kind of high-efficiency propagation method of Rhus verniciflua

Abstract

The invention discloses a method for efficiently propagating lacquer trees, and belongs to the technical field of lacquer tree propagation. The method comprises the following steps: collecting lacquer tree roots for root burying and seedling cultivation to obtain root sprout seedlings, then using the root sprout seedlings as explant materials for tissue culture, and improving the tissue culture process, thereby greatly improving the propagation coefficient of the lacquer tree, and the obtained tissue culture seedlings can maintain the excellent characteristics of the mother tree. The method for propagating lacquer trees of the invention has less harm to the mother tree, and can realize large-scale factory seedling cultivation.

Description

Technical Field

[0001] The invention relates to the technical field of Rhus verniciflua propagation, and in particular to a method for efficiently propagating Rhus verniciflua. Background Art

[0002] Toxicodendron vernicifluum is a deciduous tree of the Anacardiaceae family, mainly distributed in the central and western regions of my country, and is an important economic tree species. The main product of the lacquer tree is raw lacquer, which is an excellent natural coating with anti-corrosion and moisture-resistant effects. It is widely used in defense, machinery, petrochemical and other industries; the seeds of the lacquer tree can be used to extract lacquer oil, which is used to make ink oil, soap, etc.; the lacquer tree is also of high value in ecological restoration and is an excellent tree species for vegetation restoration. The economic and ecological benefits of the lacquer tree are both very high, and it has extremely high cultivation value. However, the distribution of lacquer trees is relatively scattered, and they have been damaged by humans, and many excellent genotypes are left with only single plants scattered around.

[0003] At present, the main propagation methods of lacquer trees are sowing and root burying. The sowing method is easy to collect seeds, convenient to transport, and low in cost, but the outer skin of lacquer tree seeds is hard and has lacquer resin, which makes it difficult for water to enter, resulting in a low germination rate of seeds. In addition, after seed propagation, the seedlings often have separated traits, and the excellent traits of the mother tree cannot be fixed. The root burying method is relatively simple to operate, genetically stable, and not easy to degenerate, but the seedling rate is low, the demand for raw materials for root burying seedlings is large, it is highly harmful to the mother tree, and it is impossible to achieve large-scale factory seedling cultivation.

[0004] Therefore, there is a need to find a new and efficient method for the propagation of lacquer trees that can reduce the impact on mother trees while maintaining excellent genotypes and realizing large-scale factory seedling cultivation, so as to achieve sustainable development in the protection, reproduction and application of lacquer tree populations. Summary of the invention

[0005] In view of this, the purpose of the present invention is to provide a method for efficiently propagating Rhus verniciflua, using the stem segments of the root sprout seedlings obtained by burying the roots as explants for tissue culture, and improving the tissue culture method, thereby increasing the efficiency of tissue culture propagation, reducing the damage to the mother tree, and realizing large-scale factory seedling cultivation.

[0006] The present invention solves the above technical problems by the following technical means:

[0007] A method for efficiently propagating Toxicodendron vernicifluum, the method comprising:

[0008] (1) Explant preparation: Collect root segments of excellent Rhus verniciflua for root burying and seedling cultivation, collect young stem segments of root sprouts from the seedling cultivation, and rinse them with running water for 1-2 hours to obtain explants;

[0009] (2) Explant disinfection: disinfecting the explant to obtain a disinfected explant;

[0010] (3) Induction culture: Use sterile paper to absorb the moisture on the surface of the sterilized explants, then inoculate them into the induction medium, first culture them in the dark at 20-23°C for 15-20 days, and then culture them under alternating light and dark conditions for 15-25 days to grow adventitious buds;

[0011] (4) Proliferation culture: Cut the adventitious buds with a height of 2-4 cm and inoculate them into a proliferation medium for proliferation culture. Differentiated seedlings are obtained in 30-60 days.

[0012] (5) Rooting culture: Cut the differentiated seedlings with a height of 3-4 cm and inoculate them into rooting medium for rooting culture. After 30-50 days, the Rhus verniciflua tissue culture seedlings are obtained;

[0013] (6) Hardening and transplanting: Move the Rhus tree tissue culture seedlings to the greenhouse for hardening and then transplant them.

[0014] Further, the specific operation of explant disinfection is as follows:

[0015] The explants were cut into 1-3 cm segments, immersed in 70% ethanol solution for 40-70 seconds, taken out, rinsed with sterile water for 1-3 times, immersed in 0.1% mercuric chloride solution for 6-10 minutes, then taken out and rinsed with sterile water for 3-7 times to obtain sterilized explants.

[0016] Furthermore, the components of the induction medium, proliferation medium and rooting medium are as follows:

[0017] Induction medium: MS+0.2mg / L TDZ+0.1mg / L 6-BA+0.2mg / L NAA+0.2mg / LIBA+0.1mg / L2,4-D+3-5g / L agar+0.1-0.2g / L sodium thioglycolate+0.1-0.2g / L acetic anhydride+0.5-1g / L modified activated carbon;

[0018] Proliferation medium: 1 / 2MS + 1mg / L 6-BA + 0.5mg / L NAA + 0.3mg / L 2,4-D + 4-6g / L agar + 0.05-0.1g / L sodium thioglycolate + 0.05-0.1g / L acetic anhydride + 0.5-1g / L modified activated carbon;

[0019] Rooting medium: 1 / 2MS + 0.5mg / L NAA + 0.1mg / L 2,4-D + 4-5g / L agar + 0.05-0.1g / L jasmone + 0.05-0.1g / L 2-hydroxychalcone + 0.5-1g / L modified activated carbon.

[0020] Furthermore, the preparation method of modified activated carbon in the induction medium, proliferation medium and rooting medium is as follows:

[0021] Add p-tert-butyl cyclohexyl acetate to a 60wt% ethanol solution and stir to dissolve, then put it into 60-70℃ hot water and heat it in a water bath for 20-30min, then add isobutyramide and activated carbon and continue to heat it in a water bath for 30-60min. After heating in the water bath, put it into an ultrasonic oscillator and perform ultrasonic treatment at 20-40KHz and 40-50℃ for 30-60min, then filter out the solid, and dry it in a 45℃ oven to constant weight to obtain modified activated carbon.

[0022] The urushiol and other phenolic substances are contained in the rhubarb tree, which are oxidized to form quinone substances under the action of oxygen and polyphenol oxidase during tissue culture, causing poisoning to explants and causing explant death. Therefore, while preventing the oxidation of phenolic substances as much as possible, it is also necessary to reduce the accumulation of quinone substances. The present invention adds sodium thioglycolate, acetic anhydride and modified activated carbon to the culture medium, and the sodium thioglycolate and acetic anhydride work together to reduce the activity of polyphenol oxidase and phenolic substances, inhibit the occurrence of oxidative browning reaction of phenolic substances, and the activated carbon is used to adsorb the browning product quinone substances. The sodium thioglycolate, acetic anhydride and activated carbon work together to control the whole process of the browning reaction, reduce the damage of quinone substances produced by the browning of phenolic substances to explants, and improve the survival rate of tissue culture seedlings. However, during the tissue culture process, activated carbon added to the culture medium is prone to adsorb nutrients in the culture medium and hinder the growth of tissue culture seedlings. Therefore, acetic acid is used to react tert-butyl cyclohexyl ester, isobutyramide and activated carbon to change the distribution of functional groups on the surface of activated carbon. The modified activated carbon and quinone substances form hydrogen bonds through association reaction to fix the quinone substances inside the activated carbon, reducing the damage of quinone substances to tissue culture seedlings. At the same time, the hydrophilicity of the activated carbon surface is improved, and hydrophobic nutrients such as hormones in the culture medium are inhibited from entering the activated carbon, reducing the adsorption of nutrients and ensuring the growth of tissue culture seedlings.

[0023] In addition, the sodium ions in sodium thioglycolate accumulate continuously in the tissue culture seedlings, which can easily cause the destruction of chlorophyll and inhibit the growth of tissue culture seedlings. Therefore, jasmone and 2-hydroxychalcone are added to the rooting medium to regulate the enzyme activity in the sumac seedlings to promote the lignification of tissue culture seedlings, increase the lignin content and thus promote the synthesis of chloroplasts, so that the tissue culture seedlings can grow healthily and improve the survival rate of seedling transplanting.

[0024] Furthermore, in the preparation process of the modified activated carbon, the mass ratio of tert-butyl cyclohexyl acetate, isobutylamide, and activated carbon is (0.1-0.3): (0.2-0.4): (6-12).

[0025] Furthermore, the pH of the induction medium, proliferation medium and rooting medium is 5.5-6.0.

[0026] Furthermore, the light-dark alternating culture conditions in the induction culture are: 12h light / 12h dark, the light intensity is 1300-2000lx, the light culture temperature is 23-27°C, and the dark culture temperature is 18-22°C.

[0027] Furthermore, the proliferation culture and rooting culture conditions are: 16h light / 8h dark, light intensity of 2000-3000lx, light culture temperature of 23-27°C, and dark culture temperature of 18-22°C.

[0028] Further, the seedling hardening operation is specifically as follows:

[0029] Move the sumac tissue culture seedlings to the greenhouse, harden them under natural light for 2-5 days, then transfer them to a shaded and ventilated place, open the lid to harden them for 2-5 days, then take out the sumac seedlings and wash the root culture medium, transfer them to the substrate after disinfection and culture them for 20-25 days before transplanting.

[0030] Furthermore, the matrix is ​​prepared from peat soil, leaf mold and perlite in a mass ratio of 3:2:1, the pH of the matrix is ​​6.5-7.5, and the water content is 60-70%.

[0031] Beneficial effects:

[0032] 1. The present invention collects root segments of excellent lacquer trees for root-burying seedling cultivation, and then uses the young stem segments of the root sprouts obtained from the seedling cultivation as explants for tissue culture. With the same amount of lacquer tree root segments collected, the reproduction coefficient of the lacquer trees is greatly improved, the damage to the mother trees is reduced, and the cultivated seedlings can maintain the excellent characteristics of the mother trees.

[0033] 2. The present invention improves the tissue culture process, adopts a specific culture medium for tissue culture, inhibits the browning incidence of tissue culture seedlings, promotes the robust growth of tissue culture seedlings, and increases the survival rate of tissue culture seedlings. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 Embodiment 1: Growth condition picture after the Lacquer Tree seedlings are transplanted. DETAILED DESCRIPTION

[0035] The present invention will be described in detail below with reference to specific embodiments and accompanying drawings:

[0036] The invention discloses a method for efficiently propagating Rhus verniciflua. Before propagating Rhus verniciflua, modified activated carbon is first prepared, as shown below:

[0037] Take 0.2g of p-tert-butyl cyclohexyl acetate, add it to 200g of 60wt% ethanol solution, stir and dissolve, then put it into 65℃ hot water and heat it in a water bath for 25min, then add 0.3g of isobutyramide and 10g of activated carbon, continue heating in a water bath for 40min, put it into an ultrasonic oscillator after heating in the water bath, ultrasonically treat it at 25KHz and 45℃ for 50min, then filter out the solid, and dry it in a 45℃ oven to constant weight to obtain modified activated carbon.

[0038] Embodiment 1: Toxicodendron vernicifluum efficient propagation method

[0039] The components of the induction medium, proliferation medium, and rooting medium of this example are as follows:

[0040] Induction medium: MS+0.2mg / L TDZ+0.1mg / L 6-BA+0.2mg / L NAA+0.2mg / LIBA+0.1mg / L2,4-D+4g / L agar+0.15g / L sodium thioglycolate+0.15g / L acetic anhydride+0.8g / L modified activated carbon;

[0041] Proliferation medium: 1 / 2MS+1mg / L 6-BA+0.5mg / L NAA+0.3mg / L 2,4-D+5g / L agar+0.07g / L sodium thioglycolate+0.07g / L acetic anhydride+0.7g / L modified activated carbon;

[0042] The rooting medium composition is: 1 / 2MS + 0.5mg / L NAA + 0.1mg / L 2,4-D + 4g / L agar + 0.07g / L jasmone + 0.07g / L 2-hydroxychalcone + 0.6g / L modified activated carbon;

[0043] (1) Explant preparation: dig out root segments of excellent lacquer trees that are healthy, free of pests and diseases, have thick and soft bark, and produce a large amount of lacquer, and the length is 15 cm to obtain lacquer roots; arrange the lacquer roots in order of large and small ends and bundle them into small bundles of 25 roots / bundle; dig a trench about 20 cm deep and 25 cm wide, and place the bundled lacquer roots in the trench with the big end facing up and the small end facing down, slightly tilted to the south, with a distance of about 5 cm between bundles. After placing them, cover them with soil and then cover them with plastic film for accelerator. Bud treatment: after the lacquer root sprouts, dig a trench about 20 cm deep and 30 cm wide, with a trench spacing of about 40 cm; add rooting powder to clean water and dilute it to 1000 times to obtain a rooting powder solution, put the sprouted lacquer root into the rooting powder solution and soak it for 15 seconds, then put the young shoots upward with the top slightly below the ground into the trench, and then cover the lacquer root with soil; after the lacquer root grows out seedlings, collect the young stem segments of the root sprout seedlings, rinse them with running water for 1.5 hours, and obtain explants;

[0044] (2) Explant disinfection: Cut the explant into small pieces of about 2 cm, soak them in 70% ethanol solution for 50 seconds, take them out and rinse them twice with sterile water, then soak them in 0.1% mercuric chloride solution for 7 minutes, then take them out and rinse them 5 times with sterile water to obtain sterilized explants;

[0045] (3) Induction culture: Use sterile paper to absorb the moisture on the surface of the sterilized explants, then inoculate them into the induction medium, first culture them in the dark at 21°C for 18 days, then culture them under 12 h light / 12 h dark conditions, with a light intensity of 1500 lx, a light culture temperature of 25°C, and a dark culture temperature of 20°C. Adventitious buds will grow after 20 days;

[0046] (4) Proliferation culture: Cut adventitious buds with a height of about 3 cm, inoculate them into a proliferation medium, and culture them under 16 h light / 8 h dark conditions, with a light intensity of 2500 lx, a light culture temperature of 25°C, and a dark culture temperature of 20°C. After 40 days, differentiated seedlings are obtained;

[0047] (5) Rooting culture: Cut the differentiated seedlings with a height of about 3 cm, inoculate them into rooting medium, and culture them under 16 h light / 8 h dark conditions, with a light intensity of 2600 lx, a light culture temperature of 25°C, and a dark culture temperature of 20°C. After 40 days, the Rhus verniciflua tissue culture seedlings were obtained;

[0048] (6) Hardening and transplanting: Move the tissue culture seedlings of the sumac tree to a greenhouse, harden them under natural light for 4 days, then transfer them to a shaded and ventilated place, and harden them for 4 days with the cover uncovered; mix peat soil, leaf mold, and perlite in a mass ratio of 3:2:1, place them in a high-pressure steam sterilizer, sterilize them at 121°C for 25 minutes, and obtain a matrix after cooling to room temperature. Adjust the pH of the matrix to 7 and the water content to 65%; take out the sumac tree seedlings and wash the root culture medium, then immerse the roots in a 0.1% carbendazim solution for 7 minutes for disinfection. After disinfection, take them out, wash them with clean water, and plant them in the matrix after culturing for 23 days before transplanting.

[0049] Embodiment 2: Toxicodendron vernicifluum efficient propagation method

[0050] In this embodiment, the induction medium, proliferation medium, and rooting medium are composed of:

[0051] Induction medium: MS+0.2mg / L TDZ+0.1mg / L 6-BA+0.2mg / L NAA+0.2mg / LIBA+0.1mg / L2,4-D+3g / L agar+0.1g / L sodium thioglycolate+0.1g / L acetic anhydride+0.6g / L modified activated carbon;

[0052] Proliferation medium: 1 / 2MS+1mg / L 6-BA+0.5mg / L NAA+0.3mg / L 2,4-D+4g / L agar+0.05g / L sodium thioglycolate+0.05g / L acetic anhydride+0.5g / L modified activated carbon;

[0053] Rooting medium: 1 / 2MS + 0.5mg / L NAA + 0.1mg / L 2,4-D + 4g / L agar + 0.05g / L jasmone + 0.05g / L 2-hydroxychalcone + 0.5g / L modified activated carbon;

[0054] (1) Explant preparation: dig out root segments of excellent lacquer trees that are healthy, free of pests and diseases, have thick and soft bark, and produce a large amount of lacquer, and the length is 10 cm to obtain lacquer roots; arrange the lacquer roots in order of large and small ends and bundle them into small bundles of 25 roots / bundle; dig a trench about 20 cm deep and 25 cm wide, and place the bundled lacquer roots in the trench with the big end facing up and the small end facing down, slightly tilted to the south, with a distance of about 5 cm between bundles. After placing them, cover them with soil and then cover them with plastic film. Germination treatment: after the lacquer root sprouts, dig a trench about 20 cm deep and 30 cm wide, with a trench spacing of about 40 cm, add rooting powder to clean water and dilute it to 1000 times to obtain a rooting powder solution, put the sprouted lacquer root into the rooting powder solution and soak it for 15 seconds, then put the young shoots upward with the top slightly below the ground into the trench, and then cover the lacquer root with soil; after the lacquer root grows out seedlings, collect the young stem segments of the root sprout seedlings, rinse them with running water for 1 hour, and obtain explants;

[0055] (2) Explant disinfection: Cut the explant into small pieces of about 2 cm, soak them in 70% ethanol solution for 40 seconds, take them out and rinse them twice with sterile water, then soak them in 0.1% mercuric chloride solution for 6 minutes, then take them out and rinse them 4 times with sterile water to obtain sterilized explants;

[0056] (3) Induction culture: Use sterile paper to absorb the moisture on the surface of the sterilized explants, then inoculate them into the induction medium, first culture them in the dark at 20°C for 15 days, then culture them under 12 h light / 12 h dark conditions, with a light intensity of 1300 lx, a light culture temperature of 23°C, and a dark culture temperature of 18°C. After 25 days, adventitious buds will grow;

[0057] (4) Proliferation culture: Cut the adventitious buds with a height of about 4 cm, inoculate them into the proliferation medium, and culture them under the conditions of 16 h light / 8 h dark, with a light intensity of 2300 lx, a light culture temperature of 23°C, and a dark culture temperature of 18°C. After 55 days, differentiated seedlings were obtained;

[0058] (5) Rooting culture: Cut the differentiated seedlings with a height of about 3 cm, inoculate them into rooting medium, and culture them under 16 h light / 8 h dark conditions, with a light intensity of 2300 lx, a light culture temperature of 23°C, and a dark culture temperature of 18°C. After 40 days, the Rhus verniciflua tissue culture seedlings were obtained;

[0059] (6) Hardening and transplanting: Move the tissue culture seedlings of the sumac tree to a greenhouse, harden them under natural light for 3 days, then transfer them to a shaded and ventilated place, and harden them for 3 days with the cover opened; mix peat soil, leaf mold, and perlite in a mass ratio of 3:2:1, place them in a high-pressure steam sterilizer, sterilize them at 121°C for 25 minutes, cool them to room temperature to obtain a matrix, adjust the pH of the matrix to 6.5, and adjust the water content to 60%; take out the sumac tree seedlings and wash the root culture medium, then immerse the roots in 0.1% carbendazim solution for 7 minutes for disinfection, take them out after disinfection, wash them with clean water, load them into the matrix and culture them for 20 days before transplanting.

[0060] Embodiment 3: Rhus verniciflua efficient propagation method

[0061] In this embodiment, the induction medium, proliferation medium, and rooting medium are composed of:

[0062] Induction medium: MS+0.2mg / L TDZ+0.1mg / L 6-BA+0.2mg / L NAA+0.2mg / LIBA+0.1mg / L2,4-D+5g / L agar+0.2g / L sodium thioglycolate+0.2g / L acetic anhydride+1g / L modified activated carbon;

[0063] Proliferation medium: 1 / 2MS+1mg / L 6-BA+0.5mg / L NAA+0.3mg / L 2,4-D+6g / L agar+0.1g / L sodium thioglycolate+0.1g / L acetic anhydride+0.8g / L modified activated carbon;

[0064] Rooting medium: 1 / 2MS + 0.5mg / L NAA + 0.1mg / L 2,4-D + 5g / L agar 0.1g / L jasmone + 0.1g / L 2-hydroxychalcone + 0.8g / L modified activated carbon;

[0065] (1) Explant preparation: dig out root segments of excellent lacquer trees that are healthy, free of pests and diseases, have thick and soft bark, and produce a large amount of lacquer, up to a length of 20 cm to obtain lacquer roots; arrange the lacquer roots in order of large and small ends and bundle them into small bundles of 25 roots / bundle; dig a trench about 20 cm deep and 25 cm wide, and place the bundled lacquer roots in the trench with the big end facing up and the small end facing down, slightly tilted to the south, with a distance of about 5 cm between bundles. After placing them, cover them with soil and then cover them with plastic film. Germination treatment: after the lacquer root sprouts, dig a trench about 20 cm deep and 30 cm wide, with a trench spacing of about 40 cm, add rooting powder to clean water and dilute it to 1000 times to obtain a rooting powder solution, put the sprouted lacquer root into the rooting powder solution and soak it for 15 seconds, then put the young shoots upward with the top slightly below the ground into the trench, and then cover the lacquer root with soil; after the lacquer root grows out seedlings, collect the young stem segments of the root sprout seedlings, rinse them with running water for 2 hours, and obtain explants;

[0066] (2) Explant disinfection: Cut the explant into small pieces of about 2 cm, soak them in 70% ethanol solution for 70 seconds, take them out and rinse them three times with sterile water, then soak them in 0.1% mercuric chloride solution for 10 minutes, then take them out and rinse them seven times with sterile water to obtain sterilized explants;

[0067] (3) Induction culture: Use sterile paper to absorb the moisture on the surface of the sterilized explants, then inoculate them into the induction medium, first culture them in the dark at 23°C for 16 days, then culture them under 12 h light / 12 h dark conditions, with a light intensity of 1600 lx, a light culture temperature of 27°C, and a dark culture temperature of 22°C. After 18 days, adventitious buds will grow;

[0068] (4) Proliferation culture: Cut the adventitious buds with a height of about 3 cm, inoculate them into the proliferation medium, and culture them under the conditions of 16 h light / 8 h dark, with a light intensity of 2600 lx, a light culture temperature of 27°C, and a dark culture temperature of 22°C. After 35 days, differentiated seedlings are obtained;

[0069] (5) Rooting culture: Cut the differentiated seedlings with a height of about 3.5 cm, inoculate them into rooting medium, and culture them under 16 h light / 8 h dark conditions, with a light intensity of 2700 lx, a light culture temperature of 25°C, and a dark culture temperature of 22°C. After 32 days, the Rhus verniciflua tissue culture seedlings were obtained;

[0070] (6) Hardening and transplanting: Move the tissue culture seedlings of the sumac tree to a greenhouse, harden them under natural light for 5 days, then transfer them to a shaded and ventilated place, and harden them for 5 days with the cover uncovered; mix peat soil, leaf mold, and perlite in a mass ratio of 3:2:1, place them in a high-pressure steam sterilizer, sterilize them at 121°C for 25 minutes, and obtain a matrix after cooling to room temperature. Adjust the pH of the matrix to 7.5 and the water content to 70%; take out the sumac tree seedlings and wash the root culture medium, then immerse the roots in a 0.1% carbendazim solution for 7 minutes for disinfection. After disinfection, take them out, wash them with clean water, and place them in the matrix for 22 days before transplanting.

[0071] Comparative Example 1: Highly efficient propagation method of Rhus verniciflua

[0072] In contrast to Example 1, the only difference is that sodium thioglycolate was not added to the induction medium and the proliferation medium when the Toxicodendron vernicifluum was efficiently propagated in Comparative Example 1.

[0073] Comparative Example 2: Highly Efficient Propagation Method of Rhus verniciflua

[0074] In contrast to Example 1, the only difference is that in Comparative Example 2, acetic anhydride was not added to the induction medium and the proliferation medium during the efficient propagation of Toxicodendron vernicifluum.

[0075] Comparative Example 3: Highly Efficient Propagation Method of Rhus verniciflua

[0076] In contrast to Example 1, the only difference is that in Comparative Example 3, no jasmone was added to the rooting medium during the efficient propagation of Toxicodendron vernicifluum.

[0077] Comparative Example 4: Highly Efficient Propagation Method of Rhus verniciflua

[0078] In contrast to Example 1, the only difference is that in Comparative Example 4, 2-hydroxychalcone was not added to the rooting medium during the efficient propagation of Toxicodendron vernicifluum.

[0079] Comparative Example 5: Highly efficient propagation method of Rhus verniciflua

[0080] In contrast to Example 1, the only difference is that in Comparative Example 5, no modified activated carbon was added to the culture medium during the efficient propagation of Toxicodendron vernicifluum.

[0081] Comparative Example 6: Highly Efficient Propagation Method of Rhus verniciflua

[0082] In contrast to Example 1, the only difference is that the preparation method of the modified activated carbon added to the culture medium during the efficient propagation of Rhus verniciflua in Comparative Example 6 is different. In this comparative example, no tert-butyl cyclohexyl acetate is added during the preparation of the modified activated carbon. The preparation method of the modified activated carbon is as follows:

[0083] Take 0.3g of isobutyramide and 10g of activated carbon, add them to 200g of 60wt% ethanol solution, stir and mix evenly, then put it into 65℃ hot water and heat it in a water bath for 40min. After the water bath heating is completed, put it into an ultrasonic oscillator and ultrasonically treat it at 25KHz and 45℃ for 50min. Then filter out the solid and dry it in a 45℃ oven to constant weight to obtain modified activated carbon.

[0084] The rest of the raw materials and steps are the same as those in Example 1.

[0085] Comparative Example 7: Highly Efficient Propagation Method of Rhus verniciflua

[0086] In contrast to Example 1, the only difference is that the modified activated carbon preparation method used in the culture medium for efficient propagation of Rhus verniciflua in Comparative Example 7 is different. In this comparative example, no isobutyramide is added during the preparation of the modified activated carbon. The modified activated carbon preparation method is as follows:

[0087] Take 0.2g of p-tert-butyl cyclohexyl acetate and add it to 200g of 60wt% ethanol solution, stir and dissolve it, then put it into 65℃ hot water and heat it in a water bath for 25min, then add 10g of activated carbon and continue to heat it in a water bath for 40min, after the water bath heating is completed, put it into an ultrasonic oscillator, and ultrasonically treat it at 25KHz and 45℃ for 50min, then filter out the solid, and dry it in a 45℃ oven to constant weight to obtain modified activated carbon;

[0088] The rest of the raw materials and steps are the same as those in Example 1.

[0089] Comparative Example 8: Highly Efficient Propagation Method of Rhus verniciflua

[0090] In contrast to Example 1, the only difference is that in Comparative Example 8, conventional activated carbon is used as the activated carbon added to the culture medium during the efficient propagation of Toxicodendron vernicifluum.

[0091] Experiment: Experiment on efficient propagation of Rhus verniciflua

[0092] Observe and record the browning rate and rooting rate of each group of explants in Example 1 and Comparative Examples 1-8. After hardening the sumac tissue culture seedlings obtained from each group of propagation, they were cultured in the matrix for 23 days before transplanting. The transplanting site was selected in Tianma Village, Xiema Street, Beibei District, Chongqing City. The climate there is a subtropical humid climate controlled by the Southeast Asian monsoon circulation, with warm winters and early springs, abundant rainfall, an average annual temperature of 18.3°C, an average annual rainfall of 1105.4mm, and a frost-free period of 334d. The transplanting operation and later management and maintenance are the same. Record the transplant survival rate of each group of seedlings. The data on the browning rate, rooting rate and transplant survival rate of the explants in Example 1 and Comparative Examples 1-8 are shown in Table 1:

[0093] Table 1

[0094] Explant browning rate (%) Rooting rate (%) Survival rate (%) Example 1 1.4％ 91.3％ 94.5％ Comparative Example 1 19.7％ 81.9％ 90.3％ Comparative Example 2 18.2％ 82.4％ 91.1％ Comparative Example 3 2.1％ 84.5％ 85.4％ Comparative Example 4 1.9％ 85.6％ 85.7％ Comparative Example 5 22.7％ 77.7％ 83.6％ Comparative Example 6 18.8％ 82.9％ 91.8％ Comparative Example 7 18.6％ 82.5％ 92.2％ Comparative Example 8 14.5％ 80.2％ 87.5％

[0095] According to the data analysis in Table 1:

[0096] (1) In implementation 1, the browning rate of the Rhus verniciflua explants was the lowest, and the rooting rate and survival rate were higher than those of comparative examples 1-8. In comparative example 1, sodium thioglycolate was not added to the induction medium and the proliferation medium during the Rhus verniciflua tissue culture, which failed to reduce the activity of polyphenol oxidase, resulting in the generation of more quinone substances and disordered plant tissue metabolism, thereby causing an increase in the browning rate of the explants. In comparative example 2, acetic anhydride was not added to the induction medium and the proliferation medium, and the activity of urushiol was relatively high. After contacting with polyphenol oxidase, quinone substances were generated, which affected the metabolism of plant tissues and caused an increase in the browning rate of the explants.

[0097] (2) In Comparative Examples 3 and 4, no jasmone or 2-hydroxychalcone was added to the rooting medium, respectively, and the lignification degree of the tissue culture seedlings was poor, the quality of the obtained tissue culture seedlings was relatively poor, and the transplant survival rate was reduced;

[0098] (3) In comparative example 5, activated carbon was not added to the culture medium, and quinone substances could not be adsorbed, resulting in disordered plant tissue metabolism, increased browning rate of explants, and reduced rooting rate and transplant survival rate;

[0099] (4) In comparative examples 6 and 7, the modified activated carbons were prepared without adding acetic acid to tert-butyl cyclohexyl ester or isobutylamide to treat the activated carbons. The activated carbons reduced their binding rate to quinones and adsorbed nutrients, which increased the browning rate and reduced the rooting rate and transplant survival rate.

[0100] (5) In Comparative Example 8, the culture medium uses conventional activated carbon, which can adsorb quinone substances and reduce the browning rate. However, it also adsorbs nutrients in the culture medium, hindering the growth of tissue culture seedlings. The rooting rate and transplanting survival rate both decrease to varying degrees. This indicates that the improved tissue culture method of the present invention can be used to culture the tissue of Rhus verniciflua, which can effectively reduce the browning rate of explants, improve the rooting rate and transplanting survival rate; and the root sprout seedlings obtained by burying the roots of Rhus verniciflua as explants can greatly improve the breeding efficiency and realize the factory-based seedling cultivation of Rhus verniciflua.

[0101] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Although the present invention is described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the purpose and scope of the technical solutions of the present invention, which should be included in the scope of the claims of the present invention. The techniques, shapes, and structural parts not described in detail in the present invention are all known technologies.

Claims

1. A method for efficiently propagating Rhus verniciflua, characterized in that: The method is as follows: (1) Explant preparation: Collect the young stem segments of the root sprouting seedlings obtained by buried root seedling cultivation, and rinse them with running water to obtain explants; (2) Explant disinfection: disinfecting the explant to obtain a disinfected explant; (3) Induction culture: remove the water from the surface of the sterilized explants, then inoculate them into the induction culture medium, culture them in darkness for 15-20 days, and then culture them under alternating light and dark to obtain adventitious buds; (4) Proliferation culture: Cut the adventitious buds and inoculate them into the proliferation medium for proliferation culture to obtain differentiated seedlings; (5) Rooting culture: cutting the differentiated seedlings and inoculating them into rooting medium for rooting culture to obtain Rhus verniciflua tissue culture seedlings; (6) Hardening and transplanting: Move the Rhus serrata tissue culture seedlings to the greenhouse for hardening and then transplanting; The components of the induction medium, proliferation medium and rooting medium are as follows: Induction medium: MS+0.2mg / L TDZ+0.1mg / L 6-BA+0.2mg / L NAA+0.2mg / L IBA+0.1mg / L 2,4-D+3-5g / L agar+0.1-0.2g / L sodium thioglycolate+0.1-0.2g / L acetic anhydride+0.5-1g / L modified activated carbon; Proliferation medium: 1 / 2MS + 1mg / L 6-BA + 0.5mg / L NAA + 0.3mg / L 2,4-D + 4-6g / L agar + 0.05-0.1g / L sodium thioglycolate + 0.05-0.1g / L acetic anhydride + 0.5-1g / L modified activated carbon; Rooting medium: 1 / 2MS + 0.5mg / L NAA + 0.1mg / L 2,4-D + 4-5g / L agar + 0.05-0.1g / L jasmone + 0.05-0.1g / L 2-hydroxychalcone + 0.5-1g / L modified activated carbon; The preparation method of modified activated carbon in the induction medium, proliferation medium and rooting medium is as follows: Add tert-butyl cyclohexyl acetate to the ethanol solution and stir to dissolve, then add isobutylamide and activated carbon and continue to heat in a water bath for 30-60 minutes, perform ultrasonic treatment after the water bath heating is completed, filter out the solid, and dry to obtain modified activated carbon; In the preparation process of the modified activated carbon, the mass ratio of tert-butyl cyclohexyl acetate, isobutylamide and activated carbon is (0.1-0.3): (0.2-0.4): (6-12).

2. A method for efficiently propagating Toxicodendron vernicifluum according to claim 1, characterized in that: The explant disinfection operation is specifically as follows: The explants were cut into 1-3 cm segments, immersed in 70% ethanol solution for 40-70 seconds, taken out, rinsed with sterile water for 1-3 times, immersed in 0.1% mercuric chloride solution for 6-10 minutes, and then taken out and rinsed with sterile water for 3-7 times to obtain sterilized explants.

3. A method for efficiently propagating Toxicodendron vernicifluum according to claim 2, characterized in that: The pH of the induction medium, proliferation medium and rooting medium is 5.5-6.

0.

4. A method for efficiently propagating Toxicodendron vernicifluum according to claim 1, characterized in that: The light-dark alternating culture conditions in the induction culture are: 12h light / 12h dark, the light intensity is 1300-2000lx, the light culture temperature is 23-27°C, and the dark culture temperature is 18-22°C.

5. A method for efficiently propagating Toxicodendron vernicifluum according to claim 4, characterized in that: The proliferation culture and rooting culture conditions are: 16h light / 8h dark, light intensity of 2000-3000lx, light culture temperature of 23-27°C, and dark culture temperature of 18-22°C.

6. A method for efficiently propagating Toxicodendron vernicifluum according to claim 5, characterized in that: The seedling hardening operation is as follows: Move the sumac tissue culture seedlings to the greenhouse, harden them under natural light for 2-5 days, then transfer them to a shaded and ventilated place, open the lid to harden them for 2-5 days, then take out the sumac seedlings and wash the root culture medium, transfer them to the substrate after disinfection and culture them for 20-25 days before transplanting.

7. A method for efficiently propagating Toxicodendron vernicifluum according to claim 6, characterized in that: The matrix is ​​prepared by mixing peat soil, leaf mold and perlite in a mass ratio of 3:2:1, and the pH value of the matrix is ​​6.5-7.5, and the water content is 60-70%.

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