A method for rapid tissue propagation of Dalbergia odorifera

Abstract

The invention discloses a tissue rapid propagation method of Dalbergia odorifera, which comprises inserting a Dalbergia odorifera stem segment into a Dalbergia odorifera sprouting and rooting medium for sprouting and rooting culture, obtaining a Dalbergia odorifera plant having clustered buds and roots, wherein the Dalbergia odorifera sprouting and rooting medium comprises an MS basal medium, 4-8g / L agar, 25-35g / L sucrose, 1.0-2.0mg / L activated carbon, 15-25ml / L coconut milk, 0.12-0.18mg / L 6-BA and 0.08-0.12mg / L KT, a pH of 5.5-6.0, a culture temperature of 20-28°C, a culture illumination time of 14-18 hours per day, a culture illumination intensity of 1500-3000lx, and a culture time of 15-20 days. The method can efficiently obtain the regenerated seedling of Dalbergia odorifera, which can be used for forestry production.

Description

Technical Field

[0001] The invention belongs to the field of plant tissue culture and relates to a tissue rapid propagation method of Dalbergia odorifera. Background Art

[0002] Dalbergia odorifera (scientific name: Dalbergia odorifera), also known as Hainan Dalbergia, Hainan Huanghuali, and Huali, is a plant of the genus Dalbergia. It is a tall tree native to Hainan, my country, and is well-known as the most precious type of redwood. The wood properties of Dalbergia odorifera are extremely stable, and it does not deform, crack, or bend. It is a high-quality furniture wood. It is now a national second-level protected plant.

[0003] The reproduction and growth of Dalbergia odorifera grown under natural conditions are very slow, which is difficult to meet the large demand. The survival rate of ordinary cuttings is low and is restricted by the season. Therefore, artificial tissue propagation of Dalbergia odorifera has a good application prospect. Summary of the invention

[0004] In order to solve the problems existing in the prior art, the present invention provides a tissue rapid propagation method of Dalbergia odorifera. The method comprises the following steps: inserting a Dalbergia odorifera stem segment into a Dalbergia odorifera sprouting and rooting culture medium for sprouting and rooting culture to obtain a Dalbergia odorifera plant having clustered sprouts and roots, wherein the Dalbergia odorifera sprouting and rooting culture medium comprises an MS basal culture medium, 4-8 g / L agar, 25-35 g / L sucrose, 15-25 ml / L coconut milk, 0.12-0.18 mg / L 6-BA and 0.08-0.12 mg / L KT.

[0005] In some embodiments, the Dalbergia odorifera sprouting and rooting culture medium further includes 1.0-2.0 mg / L activated carbon.

[0006] In some embodiments, the Dalbergia odorifera sprouting and rooting medium includes MS basal medium, 5.5 g / L agar, 30 g / L sucrose, 20 ml / L coconut milk, 1.5 mg / L activated carbon, 0.15 mg / L 6-BA and 0.1 mg / L KT.

[0007] In some embodiments, the pH of the Dalbergia odorifera sprouting and rooting medium is 5.5-6.0.

[0008] In some embodiments, the Dalbergia odorifera stem segment contains 1, 2 or 3 axillary buds.

[0009] In some embodiments, the Dalbergia odorifera stem segment is from Dalbergia odorifera twigs.

[0010] In some embodiments, the Dalbergia odorifera stem segment is a sterilized Dalbergia odorifera stem segment.

[0011] In some embodiments, the disinfection steps of the Dalbergia odorifera stem segment are: rinse with clean water for 20-60 minutes, disinfect in 70-80v / v% alcohol for 15-25s, wash with deionized water 1-3 times, disinfect in 1.8-2.2w / v% sodium hypochlorite aqueous solution for 4-6 minutes, and wash with deionized water 3-5 times.

[0012] In some embodiments, the culture conditions for the sprout and rooting culture are: culture temperature 20-28° C., culture light time 14-18 hours per day, culture light intensity 1500-3000 lx, and culture time 15-20 days.

[0013] In some embodiments, the method further comprises transplanting the Dalbergia odorifera plants having clustered buds and roots into soil to obtain Dalbergia odorifera plants growing in soil.

[0014] In some embodiments, a culture solution is irrigated into the soil to maintain the soil at 60-80% humidity for 7-10 days, wherein the culture solution includes MS basal medium, 25-35 g / L sucrose, 15-25 ml / L coconut water, 1.0-2.0 mg / L activated carbon, 0.12-0.18 mg / L 6-BA and 0.08-0.12 mg / L KT.

[0015] In some embodiments, the MS basal medium includes 600 mg / L ammonium nitrate, 500 mg / L calcium nitrate tetrahydrate, 1700 mg / L potassium nitrate, 370 mg / L magnesium sulfate heptahydrate, 170 mg / L potassium dihydrogen phosphate, 0.025 mg / L copper sulfate pentahydrate, 6.2 mg / L boric acid, 18.9 mg / L manganese sulfate monohydrate, 0.25 mg / L sodium molybdate dihydrate, 8.6 mg / L zinc sulfate heptahydrate, 0.025 mg / L cobalt chloride hexahydrate, 0.83 mg / L potassium iodide, 27.8 mg / L ferrous sulfate heptahydrate, 37.26 mg / L Na2-EDTA, 2.0 mg / L glycine, 100 mg / L inositol, 0.50 mg / L niacin, 0.1 mg / L vitamin B1 and 0.5 mg / L vitamin B6. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 This is a photo of the rapid propagation plant of Dalbergia odorifera.

[0017] Figure 2 This is a photo of the rapid propagation plant of Dalbergia odorifera. DETAILED DESCRIPTION

[0018] In order to make the objectives, technical solutions and advantages of the present invention more clear, the embodiments of the present invention will be further described in detail below with reference to the accompanying drawings.

[0019] Example 1: Rapid tissue propagation of Dalbergia odorifera

[0020] 1. Preparation of culture medium

[0021] (1) Basic culture medium

[0022] The basal culture medium is MS culture medium, which specifically includes macroelements: 600 mg / L ammonium nitrate, 500 mg / L calcium nitrate tetrahydrate, 1700 mg / L potassium nitrate, 370 mg / L magnesium sulfate heptahydrate, 170 mg / L potassium dihydrogen phosphate; trace elements: 0.025 mg / L copper sulfate pentahydrate, 6.2 mg / L boric acid, 18.9 mg / L manganese sulfate monohydrate, 0.25 mg / L sodium molybdate dihydrate, 8.6 mg / L zinc sulfate heptahydrate, 0.025 mg / L cobalt chloride hexahydrate, 0.83 mg / L potassium iodide; iron salt: 27.8 mg / L ferrous sulfate heptahydrate, 37.26 mg / L Na2-EDTA; organic components: 2.0 mg / L glycine, 100 mg / L inositol, 0.50 mg / L niacin, 0.1 mg / L vitamin B1, 0.5 mg / L vitamin B6.

[0023] (2) Dalbergia odorifera sprout and rooting medium

[0024] MS medium was used as the basic medium, and the formula also included 5.5 g / L agar, 30 g / L sucrose, 20 ml / L coconut milk, and 1.5 mg / L activated carbon. The hormone components and contents in the medium are shown in Table 1. The pH was 5.75.

[0025] Coconut juice was purchased from Blue Ridge Beverage Company, Inc.

[0026] 2. Obtaining and sterilizing explants

[0027] (I) Selection and treatment of explants

[0028] Take the young branches of Dalbergia odorifera grown in the field, remove the leaves, clean the dust, soil and other impurities on the surface of the branches, and rinse them under tap water for 30-40 minutes.

[0029] (II) Disinfection of explants

[0030] 1. Prepare 100 ml of 75 v / v% alcohol, add 1-2 drops of Tween-20, shake well and set aside; prepare 100 ml of 2 w / v% (2 g / 100 mL) sodium hypochlorite aqueous solution, add 1-2 drops of Tween-20 and set aside;

[0031] 2. Cut the branches into segments of moderate length, ensuring that each branch contains an axillary bud, and the axillary bud is at least 0.5 cm away from the lower end of the stem segment.

[0032] 3. Place the cut stem segments into the 75v / v% alcohol prepared in advance for disinfection for 20 seconds, and shake slowly.

[0033] 4. Pour out the alcohol and add sterilized deionized water to rinse twice.

[0034] 5. Add the prepared sodium hypochlorite solution and disinfect for 5 minutes, shaking slowly to achieve better disinfection effect.

[0035] 6. Pour out the sodium hypochlorite solution and wash with sterile deionized water 4-5 times until the smell of sodium hypochlorite is gone. Place the stem segments on sterilized filter paper to absorb excess water and set aside.

[0036] 3. Cluster bud and root induction

[0037] For each culture dish of Dalbergia odorifera sprouting and rooting medium corresponding to the hormone in Table 1, one sterilized stem segment was inoculated for induction culture. Culture conditions: temperature 24°C, photoperiod 16 hours light / 8 hours dark, light intensity 1500-3000lx, culture time 15-20d. The culture results are shown in the last two columns of Table 1, and 10 culture dishes were cultured in each culture medium.

[0038] Table 1

[0039] serial number 6-BA(mg / L) KT(mg / L) Cluster bud situation Rooting 1 0.05 0.06 Slow growth Slow rooting 2 0.08 0.1 Slow growth Slow rooting time 3 0.15 0.1 Faster rooting Short rooting time, many adventitious roots 4 0.20 0.3 Slow growth Slow rooting

[0040] It can be seen that the culture medium No. 3 in Table 1 is the best and is called the optimal culture medium. Figure 1 and Figure 2 The figures show the Dalbergia odorifera plants grown with clustered shoots and roots obtained by culture in medium No. 3.

[0041] 4. Transplantation

[0042] The optimal medium minus the agar was used as the culture medium for transplanting.

[0043] Transplant the Dalbergia odorifera plants in the culture dish corresponding to the optimal culture medium into the soil, the soil composition is: vermiculite: nutrient soil: moss (1:1:1 weight ratio), add the above culture solution into the soil, and add the amount to keep the soil humidity at 70%. Maintain the temperature at 20-28℃, the light and dark cycle is 16h light / 8h dark, the light intensity is 1500-3000lx, stop irrigating the culture solution after 7-10 days of culture, use ordinary clean water for irrigation, stop light and temperature and humidity control, and enter routine management.

[0044] Therefore, the method of the present invention can achieve the rapid propagation of Dalbergia odorifera.

[0045] It is known from common technical knowledge that the present invention can be implemented by other embodiments that do not deviate from its spirit or essential features. Therefore, the above disclosed embodiments are only illustrative in all respects and are not exclusive. All changes within the scope of the present invention or within the scope equivalent to the present invention are included in the present invention.

Claims

1. A tissue rapid propagation method for Dalbergia odorifera, the method comprising: inserting a Dalbergia odorifera stem segment into a Dalbergia odorifera sprouting and rooting medium for sprouting and rooting culture to obtain a Dalbergia odorifera plant having clustered buds and roots, the Dalbergia odorifera sprouting and rooting medium comprising MS basal medium, 4-8 g / L agar, 25-35 g / L sucrose, 15-25 ml / L coconut milk, 0.12-0.18 mg / L 6-BA and 0.08-0.12 mg / L KT; the culture conditions for the sprouting and rooting culture are: a culture temperature of 20-28° C., a culture illumination time of 14-18 hours per day, a culture illumination intensity of 1500-3000 lx, and a culture time of 15-20 days; The method further comprises transplanting the Dalbergia odorifera plant having clustered buds and roots into soil to obtain the Dalbergia odorifera plant growing in the soil; Watering the soil with a culture solution to maintain the soil at a humidity of 60-80% for 7-10 days, wherein the culture solution consists of MS basal medium, 25-35 g / L sucrose, 15-25 ml / L coconut milk, 1.0-2.0 mg / L activated carbon, 0.12-0.18 mg / L 6-BA and 0.08-0.12 mg / L KT; The MS basal medium consists of 600 mg / L ammonium nitrate, 500 mg / L calcium nitrate tetrahydrate, 1700 mg / L potassium nitrate, 370 mg / L magnesium sulfate heptahydrate, 170 mg / L potassium dihydrogen phosphate, 0.025 mg / L copper sulfate pentahydrate, 6.2 mg / L boric acid, 18.9 mg / L manganese sulfate monohydrate, 0.25 mg / L sodium molybdate dihydrate, 8.6 mg / L zinc sulfate heptahydrate, 0.025 mg / L cobalt chloride hexahydrate, 0.83 mg / L potassium iodide, 27.8 mg / L ferrous sulfate heptahydrate, 37.26 mg / L Na2-EDTA, 2.0 mg / L glycine, 100 mg / L inositol, 0.50 mg / L nicotinic acid, 0.1 mg / L vitamin B1 and 0.5 mg / L vitamin B6.

2. The method according to claim 1, characterized in that The Dalbergia odorifera sprouting and rooting culture medium also includes 1.0-2.0 mg / L activated carbon.

3. The method according to claim 2, characterized in that The Dalbergia odorifera sprouting and rooting culture medium consists of MS basic culture medium, 5.5 g / L agar, 30 g / L sucrose, 20 ml / L coconut milk, 1.5 mg / L activated carbon, 0.15 mg / L 6-BA and 0.1 mg / L KT.

4. The method according to claim 1, characterized in that The pH of the Dalbergia odorifera sprouting and rooting culture medium is 5.5-6.

0.

5. The method according to claim 1, characterized in that The Dalbergia odorifera stem segment contains 1, 2 or 3 axillary buds.

6. The method according to claim 1, characterized in that The Dalbergia odorifera stem segments are from Dalbergia odorifera young branches.

7. The method according to claim 1, characterized in that The Dalbergia odorifera stem segment is a sterilized Dalbergia odorifera stem segment.

8. The method according to claim 7, characterized in that The disinfection steps of Dalbergia odorifera stem segments are as follows: rinse with clean water for 20-60 minutes, disinfect in 70-80 v / v % alcohol for 15-25 seconds, wash with deionized water 1-3 times, disinfect in 1.8-2.2 w / v % sodium hypochlorite aqueous solution for 4-6 minutes, and wash with deionized water 3-5 times.

Images