CN111084104B - Method for rapidly propagating orange-heart ginger flowers - Google Patents
Method for rapidly propagating orange-heart ginger flowers Download PDFInfo
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Abstract
The invention discloses a rapid propagation method of orange-heart ginger flower, which adopts red light: blue light: carrying out adventitious bud multiplication subculture by using white light according to the illumination ratio of 7-8: 1:1, and adopting red light in the rooting stage: blue light: the white light illumination ratio is 4-5: 3-5: 1, and then 800-1000ppm CO is used 2 The method for rooting culture promotes the growth of tissue culture seedlings and adventitious roots, improves the quality of ginger seedlings, shortens the culture time of 20-25 days, and implements large-scale production. Provides a favorable way for tissue culture growth and vegetative growth of the orange heart ginger flower.
Description
Technical Field
The invention particularly relates to a rapid propagation method of orange-heart ginger flower.
Background
'orange heart ginger blossom' (Hedychium, also known as Chengxin) is a perennial herb of the genus Hedychium of the family Zingiberaceae, and is a new species formed by hybridization with white ginger blossom (Hedychium coronarium) as a female parent and pink ginger blossom (Hedychium, also known as Fen) as a male parent, and the variety approval serial numbers are: yue Ju Hua: 20170005.
the height of the orange-heart ginger flower is 1.3-1.8 m. The leaves are needle-shaped, the front surface is smooth, and the back surface is densely covered with short white hairs. The spike-shaped inflorescences grow at the tops, grow for 19-21 cm, and each inflorescence blossoms 4-6 flowers. The corolla is yellowish, the lobe is yellowish, the lip is white, the base part has a yellowish to orange plaque, the lateral degenerated stamen is white, the base part has an orange plaque, the filament is orange red, and the anther is orange red. The fragrance is elegant and the gardenia is fragrant. In Guangzhou, the natural flowering phase of 'orange-heart ginger flower' is 5-11 months. The orange-heart ginger flower inherits the characteristics of bright and bright color and long flowering period of a male parent and also inherits the characteristics of ginger blast resistance and cold tolerance of a female parent. Meanwhile, the shape and the size ratio of each part of the orange-heart-shaped ginger flower are more coordinated than those of parents, the base part of the lip valve is provided with heart-shaped yellowish patches, the ornamental value is better, the fragrance is pleasant, and the orange-heart-shaped ginger flower is an excellent cutting flower material and is also a good landscaping plant.
With the improvement of the overall aesthetic level of society, people also put higher requirements on flowering flowers and garden plants. Flower plants with single flower color, short flowering period and lack of fragrance can easily cause aesthetic fatigue, and new product injection with rich color, unique flower type and pleasant fragrance is urgently needed. The 'orange heart ginger flower' has bright and colorful color and pleasant faint scent, and is very suitable for being used as a fresh cut flower or landscaping. But the propagation coefficient is low and limited by seasons, so that the large-scale production cannot be carried out, and the market popularization is influenced.
According to the existing problems, improvement of the culture technology of the orange heart ginger flower is a problem to be solved urgently.
Disclosure of Invention
The invention aims to adopt an improved tissue culture technology to carry out large-scale production on the orange heart ginger flower, improve the propagation coefficient of the orange heart ginger flower, shorten the tissue culture period of the orange heart ginger flower, reduce the sensitivity of the orange heart ginger flower on the influence of seasons and improve the seedling quality.
The technical scheme adopted by the invention is as follows:
the invention provides a method for rapidly propagating orange-heart ginger flowers in a first aspect. According to an embodiment of the invention, the method comprises the steps of:
1) accelerating germination of underground tubers of orange-heart ginger flowers;
2) taking buds as explants, inoculating the buds to an adventitious bud induction culture medium after disinfection, and inducing the adventitious buds;
3) culturing the adventitious bud in a cluster bud culture medium to form a cluster bud;
4) transferring the cluster buds to a rooting culture medium, and performing rooting culture to obtain grouped cultured seedlings;
5) transplanting the tissue culture seedlings into the mixed matrix for culture after the tissue culture seedlings grow into seedlings, and obtaining the 'orange heart ginger flower' plants through conventional culture.
According to an embodiment of the present invention, the step 1) germination accelerating method comprises the following steps: soaking tubers in 6-benzyladenine solution, and culturing in dark.
According to an embodiment of the present invention, the adventitious bud induction medium of step 2) includes the following components: 0.5-1.0 mg of thidiazuron, 6-8 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and adjusting the pH value to 5.8-6.0.
According to an embodiment of the invention, the adventitious bud induction conditions of step 2) are: the culture temperature is 25-28 ℃, and the white light radiation is 30-60 mu mol/m 2 And/s, the illumination time is 12-14 hours/day, and the culture time is 7-10 days.
According to an embodiment of the present invention, the culture medium for multiple shoots in step 3) comprises the following components: 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 25-30 g of sucrose and 5-7 g of agar, and adjusting the pH to 5.8-6.0.
According to an embodiment of the present invention, the adventitious bud culture conditions in step 3) are: the temperature is 25-28 ℃, the number ratio of red light bulbs to blue light bulbs to white light bulbs is 7-8: 1, and the effective light radiation is 60-90 mu mol/m 2 And/s, the illumination time is 14-16 hours/day.
According to an embodiment of the invention, the rooting medium of step 4) comprises the following components: 0.5-1.5 g of active carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 25-30 g of cane sugar and 5-7 g of agar, and the pH is adjusted to 5.8-6.0.
According to the embodiment of the invention, the rooting culture conditions in the step 4) are as follows: carbon dioxide concentration during culture is 800ppm up to1000ppm, and the culture temperature is 25-28 ℃; the number ratio of red light bulbs, blue light bulbs and white light bulbs is 4-5: 3-5: 1 during culture, and the effective light radiation is 60-90 mu mol/m 2 And the illumination time is 14-16 hours/day.
According to the embodiment of the invention, the mixed matrix in the step 5) comprises peat soil and perlite; the mass ratio of the peat soil to the perlite is 4-6: 1.
According to the embodiment of the invention, the wavelength of the red light is 650 nm-700 nm, the wavelength of the blue light is 400 nm-450 nm, and the white light is composite light.
When the tissue culture seedling takes root for culture, CO is filled in 2 Can increase carbon source, and is favorable for tissue culture seedling leaf to absorb CO 2 Carrying out photosynthesis to generate organic matters for the root growth of the tissue culture seedlings; reducing red light and enhancing blue light are beneficial to inducing the generation and growth of adventitious roots.
The invention can improve red light to promote the growth of plant stem; the low proportion of blue light promotes the photosynthetic effect, is beneficial to the accumulation of organic matters, but the excessive blue light can inhibit the growth; white light is to supplement other light qualities, which are beneficial for plant growth at low ratios.
The method adopts red light: blue light: and (5) carrying out adventitious bud multiplication subculture under the illumination condition of 7-8: 1:1 of white light. Can promote the growth of tissue culture seedling, shorten culture time and improve the quality of tissue culture seedling.
Red light is adopted in the rooting stage: blue light: illumination conditions of 4-5: 3-5: 1 for white light, and charging of 800-1000ppm CO 2 And (5) performing rooting culture. Can promote photosynthesis, promote generation of adventitious root, shorten culture time, and improve quality.
The invention has the beneficial effects that:
the invention successfully utilizes the plant tissue culture technology to realize the tissue culture propagation of the orange heart ginger flower, and compared with the prior art, the invention utilizes the specific proportion of red light and blue light and adds CO 2 Gas-promoted growth of tissue culture seedlings has shortened the tissue culture period. Moreover, compared with the traditional tissue culture mode, the rapid propagation of the seedlings can be realized, the whole process is shortened by 20-25 days, the investment is less, and the yield is high; group ofThe method has the advantages of strong seedling culture, developed root system, fast seedling emergence, high stability, strong seedling, good seedling quality, strong resistance and good growth, and provides an effective way for marketization of the orange-heart ginger flowers.
Drawings
FIG. 1 shows germination acceleration by sand storage.
FIG. 2 is explant inoculation.
FIG. 3 shows the generation of adventitious buds.
FIG. 4 shows single bud transfer.
FIG. 5 is the formation of multiple shoots.
FIG. 6 shows rooting culture.
FIG. 7 shows the growth of the root seedling.
FIG. 8 shows the transplanting of the grown seedlings.
Detailed Description
Reagents and materials:
chlorothalonil is purchased from chemical industry Co., Ltd, and the registration number of the pesticide is PD 85159-6; 6-Benzylanine was purchased from Guangzhou Dingguo Biotechnology Inc., CAS number: 1214-39-7, order number: DH 038-2; mercury mercurial is available from sigma aldrich (shanghai) trade, inc, CAS number: 7487-94-7, cat #: M1136-25G; thidiazuron was purchased from guang nations biotechnology limited, CAS No.: 51707-55-2, order number: DH 428-1; naphthylacetic acid was purchased from guangzhou dingguo biotechnology limited, CAS No.: 86-87-3, order number: DH 211-2.
The invention is further described below with reference to examples and comparative examples:
example 1
(1) Sterilizing and germinating underground tuber of orange-heart ginger flower
Digging underground tubers of 'orange-heart ginger flowers' from the field, removing roots, overground stems and leaves, washing with tap water, brushing and cleaning with a soft brush, soaking in chlorothalonil solution with the mass fraction of 0.3% for 20 minutes, taking out, and wiping with sterile paper. Then, underground tubers were soaked in 1 mg/L6-benzyladenine (6-BA) solution for 3 minutes, taken out and embedded in sterilized wet sand, and cultured at 28 ℃ in the dark.
FIG. 1 shows germination acceleration by sand storage.
(2) Selecting explant, inducing adventitious bud
Cutting off leaves of buds, keeping buds with the length of 1cm as explants, soaking the buds in 75% alcohol for 30 seconds, disinfecting the buds for 8 minutes by using 0.1mg/L mercuric chloride solution, washing the buds for 5 times by using sterile water, inoculating the explants to an adventitious bud induction culture medium (each liter of the culture medium comprises 0.5-1.0 mg of thidiazuron, 6-benzyladenine 6-8 mg, 0.1-0.3 mg of naphthylacetic acid, 30g of sucrose and 5g of agar, and the rest components are components of an MS culture medium, and adjusting the pH to 5.8-6.0) for culture, wherein the culture temperature is 28 ℃, the LED white light illumination is carried out, and the effective light radiation is 60 mu mol/m 2 And/s, the illumination time is 12 hours/day.
FIG. 2 is explant inoculation and FIG. 3 produces adventitious buds.
(3) Multiplication and subculture of adventitious buds
Cutting the adventitious bud into single pieces, culturing in cluster bud induction culture medium (each liter of culture medium comprises 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, and the rest components are components of MS culture medium, and adjusting pH to 5.8-6.0) to breed cluster bud, culturing at 28 deg.C, illuminating with LED, illuminating with red light, blue light and white light at a ratio of 8:1, and irradiating with composite effective light radiation of 60 μmol/m 2 And/s, the illumination time is 16 hours/day for multiplication and subculture of the adventitious bud.
FIG. 4 is a single shoot transfer, and FIG. 5 is a cluster shoot.
(4) Rooting culture of tissue culture seedling
Cutting buds growing to 2-3 cm high, transferring the buds to a rooting culture medium (each liter of the culture medium comprises 0.5-1.5 g of activated carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and the rest components are components of 1/2MS culture medium, the pH value is 5.8-6.0), carrying out rooting culture to form tissue culture seedlings, controlling the concentration of carbon dioxide in a culture chamber to be 1000ppm, carrying out culture at 28 ℃, carrying out illumination by adopting the ratio of red light, blue light and white light to be 5: 4: 1, and carrying out effective optical radiation to be 60 mu mol/m 2 And/s, illumination time 16 hours/day.
FIG. 6 rooting culture, FIG. 7 rooting seedling.
(5) Transplanting of grown seedlings
Taking out the seedlings, washing off the culture medium at the roots, planting the seedlings in a mixed matrix of peat soil and perlite in a mass ratio of 6: 1, and keeping the humidity at 60% and shading at 40%. The cultivation can be carried out in open field after 7 days, and the tissue culture seedlings can bloom after 8-10 months of cultivation. FIG. 8 transplanting the grown seedlings.
As a result: the adventitious bud is illuminated at the ratio of red light, blue light and white light of 8:1, 1 successive transfer period can be completed in 20 days, and 3-4 buds can be formed by multiplying 1 bud in each 1 successive transfer period.
In the process of cultivating the cluster buds into group seedlings, the concentration of carbon dioxide is controlled to be 1000ppm, and the ratio of red light, blue light and white light is 5: 4: 1 for illumination, so that the result shows that the buds grow to 7cm in 20 days, more than 5 adventitious roots can grow, the root length reaches more than 6.0cm, and the rooting rate reaches 100%.
Example 1 took 74 days from sprouting to flowering of tubers.
Example 2
(1) Sterilizing and accelerating germination of underground tubers of orange-heart ginger flowers
Digging underground tubers of 'orange-heart ginger flowers' from the field, removing roots, overground stems and leaves, washing with tap water, brushing and cleaning with a soft brush, soaking in chlorothalonil solution with the mass fraction of 0.3% for 20 minutes, taking out, and wiping with sterile paper. Then, underground tubers were soaked in 1 mg/L6-benzyladenine (6-BA) solution for 3 minutes, taken out and embedded in sterilized wet sand, and cultured at 28 ℃ in the dark.
FIG. 1 shows germination acceleration by sand storage.
(2) Selecting explant, inducing adventitious bud
Cutting off leaves of buds, reserving buds with the length of 1cm as explants, soaking the buds in 75% alcohol for 30 seconds, then disinfecting the buds for 8 minutes by 0.1mg/L mercuric chloride solution, washing the buds for 5 times by using sterile water, inoculating the explants after washing to an adventitious bud induction culture medium (each liter of the culture medium comprises 0.5-1.0 mg of thidiazuron, 6-benzyl adenine 6-8 mg, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, and the balance of MS culture medium components, adjusting the pH to 5.8-6.0, culturing at the temperature of 28 ℃, irradiating by using LED white light, and irradiating by using effective light of 60 mu mol/m 2 And/s, the illumination time is 12 hours/day.
FIG. 2 is explant inoculation and FIG. 3 produces adventitious buds.
(3) Multiplication and subculture of adventitious buds
Cutting the adventitious bud into single pieces, culturing in cluster bud induction culture medium (each liter of culture medium comprises 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, and the rest components are components of MS culture medium, and adjusting pH to 5.8-6.0) to breed cluster bud, culturing at 28 deg.C, illuminating with LED, and illuminating with red light, blue light and white light at a ratio of 7: 1, and composite effective light radiation of 60 μmol/m 2 And/s, the illumination time is 16 hours/day for multiplication and subculture of the adventitious bud.
FIG. 4 is a single shoot transfer, and FIG. 5 is a cluster shoot.
(4) Rooting culture of tissue culture seedling
Cutting buds growing to 2-3 cm high, transferring the buds to a rooting culture medium (each liter of the culture medium comprises 0.5-1.5 g of activated carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and the rest components are components of 1/2MS culture medium, the pH value is 5.8-6.0), carrying out rooting culture to form tissue culture seedlings, controlling the carbon dioxide concentration in a culture chamber to be 800ppm, carrying out culture at 28 ℃, carrying out illumination by adopting the ratio of red light, blue light and white light to be 5: 4: 1, and carrying out effective optical radiation to be 60 mu mol/m 2 And/s, illumination time 16 hours/day.
FIG. 6 rooting culture, FIG. 7 rooting seedling.
(5) Transplanting of grown seedlings
Taking out the seedlings, washing off the culture medium at the roots, planting the seedlings in a mixed matrix of peat soil and perlite in a mass ratio of 6: 1, and keeping the humidity at 60% and shading at 40%. The cultivation can be carried out in open field after 7 days, and the tissue culture seedlings can bloom after being cultivated for 8-10 months. FIG. 8 transplanting the grown seedlings.
As a result: example 2 took 75 days from tuber sprouting to flowering.
Example 3
(1) Sterilizing and accelerating germination of underground tubers of orange-heart ginger flowers
Digging underground tubers of 'orange-heart ginger flowers' from the field, removing roots, overground stems and leaves, washing with tap water, brushing and cleaning with a soft brush, soaking in chlorothalonil solution with the mass fraction of 0.3% for 20 minutes, taking out, and wiping with sterile paper. Then, underground tubers were soaked in 1 mg/L6-benzyladenine (6-BA) solution for 3 minutes, taken out and embedded in sterilized wet sand, and cultured at 28 ℃ in the dark.
FIG. 1 shows germination acceleration by sand storage.
(2) Selecting explant, inducing adventitious bud
Cutting off leaves of buds, reserving buds with the length of 1cm as explants, soaking the buds in 75% alcohol for 30 seconds, then disinfecting the buds for 8 minutes by 0.1mg/L mercuric chloride solution, washing the buds for 5 times by using sterile water, inoculating the explants after washing to an adventitious bud induction culture medium (each liter of the culture medium comprises 0.5-1.0 mg of thidiazuron, 6-benzyl adenine 6-8 mg, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, and the balance of MS culture medium components, adjusting the pH to 5.8-6.0, culturing at the temperature of 28 ℃, irradiating by using LED white light, and effectively irradiating by using 60 mu mol/m light radiation 2 And/s, the illumination time is 12 hours/day.
FIG. 2 is explant inoculation and FIG. 3 produces adventitious buds.
(3) Multiplication and subculture of adventitious buds
Cutting the adventitious bud into single pieces, culturing in cluster bud induction culture medium (each liter of culture medium comprises 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, and the rest components are components of MS culture medium, and adjusting pH to 5.8-6.0) to breed cluster bud, culturing at 28 deg.C, illuminating with LED, and illuminating with red light, blue light and white light at a ratio of 7: 1, and composite effective light radiation of 60 μmol/m 2 And/s, the illumination time is 16 hours/day for multiplication subculture of adventitious buds.
FIG. 4 is a single shoot transfer, and FIG. 5 is a cluster shoot.
(4) Rooting culture of tissue culture seedling
Cutting buds growing to 2-3 cm high, transferring the buds to a rooting culture medium (each liter of the culture medium comprises 0.5-1.5 g of activated carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and the balance of components of 1/2MS culture medium, and the pH value is 5.8-6.0) to form tissue culture seedling, controlling the concentration of carbon dioxide in the culture chamber to be 1000ppm, controlling the culture temperature to be 28 ℃, and adopting the ratio of red light, blue light and white light to be 6: 3: 1 to carry out illumination, wherein the effective light radiation is 60 mu mol/m 2 And/s, illumination time 16 hours/day.
FIG. 6 rooting culture, FIG. 7 rooting seedling.
(5) Transplanting of grown seedlings
Taking out the seedlings, washing off the culture medium at the roots, planting the seedlings in a mixed matrix of peat soil and perlite in a mass ratio of 6: 1, and keeping the humidity at 60% and shading at 40%. The cultivation can be carried out in open field after 7 days, and the tissue culture seedlings can bloom after 8-10 months of cultivation. FIG. 8 transplanting the grown seedlings.
As a result: example 3 took 80 days from tuber sprouting to flowering.
Next, the light conditions at the adventitious bud growth stage were examined.
(1) Adventitious bud proliferation stage
Experimental group setup: CK (all white light); t1 (Red: blue: white: 8: 1); t2 (red: blue: white: 8: 2: 1); t3 (red: blue: white: 8: 3: 1); t4 (Red: blue: white: 6: 1); t5 (red: blue: white: 4: 1), light intensity set at 60 μmol/m 2 /s。
TABLE 1 growth of adventitious buds cultured under different light conditions for 20 days
| Experimental group | Multiplication factor of single bud | Bud length (cm) | Number of blades |
| CK | 3.3 | 2.3 | 2 |
| T1 | 4.1 | 3.2 | 4 |
| T2 | 4.2 | 3.1 | 4 |
| T3 | 3.5 | 2.4 | 3 |
| T4 | 3.8 | 2.8 | 4 |
| T5 | 3.3 | 2.2 | 2 |
As can be seen from Table 1, the T1 and T2 treated groups have the highest single bud multiplication times, the longest bud length and the highest average leaf number, which indicates that the orange-heart ginger flowers have better multiplication subculture effect under the conditions. The multiplication times are reduced along with the increase of the blue light proportion, the growth period is prolonged, and the number of leaves is reduced; a similar situation occurs as the proportion of red light decreases.
In conclusion, the orange-heart ginger flower has better proliferation and subculture effects under the conditions that the ratio of red to blue to white is 8 to 1.
(2) Rooting stage
Experimental group setup: CK (all white light); t1 (red: blue: white: 5: 4: 1); t2 (red: blue: white: 5: 3: 1); t3 (red: blue: white: 3: 4: 1); t4 (red: blue: white: 7: 4: 1); t5 (Red: blue: white: 5: 6: 1) with an intensity of 60. mu. mol/m 2 /s。
TABLE 2 growth of 20-day tissue culture seedlings cultured under different illumination conditions
As can be seen from Table 2, in the rooting stage, the ratio of red light to blue light is close to each other within a certain range, and the higher ratio of red light is helpful for rooting of the tissue culture seedling.
In sum, under the condition that the ratio of red, blue and white is 5: 4: 1, the orange heart ginger flower has better rooting effect.
TABLE 3 different CO 2 The growth condition of tissue culture seedlings cultured for 20 days under the concentration
| Experimental group | Average number of roots | Root length (cm) | Plant height (cm) | Number of blades |
| CK | 3.5 | 4.2 | 5.5 | 6 |
| T1 | 5.4 | 6.3 | 6.8 | 7 |
| T1+400ppmCO 2 | 5.4 | 6.4 | 7.2 | 7 |
| T1+600ppmCO 2 | 5.4 | 6.4 | 7.4 | 7 |
| T1+800ppmCO 2 | 5.6 | 6.9 | 7.8 | 7 |
| T1+1000ppmCO 2 | 6.1 | 7.1 | 7.7 | 7 |
| T1+1200ppmCO 2 | 5.8 | 6.7 | 7.5 | 7 |
As can be seen from Table 3, in the rooting stage, a certain amount of CO was added 2 Can improve the parameters of the number of roots, the root length, the plant height and the like, and multiple indexes follow CO in a certain range 2 Increase in concentration as CO increases 2 The concentration is 800-1000ppm, and the better effect can be obtained. When CO is present 2 When the concentration exceeds 1000ppm, various indexes are not increased but are decreased to some extent, which indicates that the CO is too high 2 The concentration is not favorable for the growth of tissue culture seedlings.
In conclusion, CO 2 The orange heart ginger flower has better rooting effect when the concentration is 800-1000 ppm.
Comparative example
(1) Sterilizing and accelerating germination of underground tubers of orange-heart ginger flowers
Digging underground tuber of 'orange-heart ginger flower' from field, removing root, aboveground stem and leaf, washing with tap water, brushing with soft brush, soaking in chlorothalonil solution with mass fraction of 0.3% for 20 min, taking out, wiping with sterile paper, soaking underground tuber in 6-benzyl adenine solution of 1mg/L for 3 min, taking out, embedding in sterilized wet sand, and culturing at 28 deg.C in dark
(2) Selecting explant, inducing adventitious bud
Cutting off leaves of buds, reserving buds with the length of 1cm as explants, soaking the buds in 75 percent alcohol for 30 seconds, disinfecting the buds for 8 minutes by 0.1mg/L mercuric chloride solution, washing the buds for 5 times by using sterile water, inoculating the explants to an adventitious bud induction culture medium (containing 0.5 to 1.0mg of thidiazuron, 6 to 8mg of 6-benzyladenine, 0.1 to 0.3mg of naphthylacetic acid, 30g of sucrose and 5g of agar per liter, adjusting the pH to 5.8 to 6.0 and the balance of MS culture medium) for culture under the conditions of the temperature of 28 ℃, LED white light illumination and effective light illuminationThe injection rate is 60 mu mol/m 2 And/s, the illumination time is 12 hours/day.
(3) Multiplication and subculture of adventitious buds
Cutting adventitious buds into single pieces, culturing in cluster bud induction culture medium (each liter contains 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of sucrose, 5g of agar, pH is adjusted to 5.8-6.0, and the rest is the components of MS culture medium), culturing at 28 deg.C under white light irradiation with composite effective light radiation of 60 μmol/m 2 And/s, the illumination time is 16 hours/day for multiplication and subculture of the adventitious bud.
(4) Rooting culture of tissue culture seedling
Cutting buds growing to 2-3 cm high, transferring the buds to a rooting culture medium (each liter contains 0.5-1.5 g of active carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 30g of cane sugar, 5g of agar, pH 5.8-6.0 and the balance of components of 1/2MS culture medium) for rooting culture to form tissue culture seedlings, wherein carbon dioxide is not added, the culture condition is 28 ℃, the temperature is LED under white light illumination, and the effective light radiation is 60 mu mol/m 2 And/s, illumination time 16 hours/day.
(5) Transplanting of grown seedlings
Taking out the seedlings, washing off the culture medium at the roots, planting the seedlings in a mixed matrix of peat soil and perlite in a mass ratio of 6: 1, and keeping the humidity at 60% and shading at 40%. The cultivation can be carried out in open field after 7 days, and the tissue culture seedlings can bloom after being cultivated for 8-10 months.
As a result:
the adventitious buds can complete 1 subculture period in 30 days only by adopting white light illumination, and 2-3 buds are formed by multiplying 1 bud in each 1 subculture period. In the process of cultivating the cluster buds into the group culture seedlings, carbon dioxide is not added, only the LED white light is used for illumination, and the result shows that the buds grow to 7cm in 35 days, 4-5 adventitious roots can grow, the root length reaches 6cm, and the rooting rate reaches 100%. Sprout from tuber to flower, the comparative example lasts for 99 days.
In conclusion, the method can improve the propagation coefficient of the orange-heart ginger flower, shorten the tissue culture period and realize large-scale production.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (1)
1. A method for rapid propagation of orange-heart ginger flower is characterized by comprising the following steps:
1) accelerating germination of underground tubers of orange-heart ginger flowers;
2) taking buds as explants, inoculating the buds to an adventitious bud induction culture medium after disinfection, and inducing the adventitious buds;
3) culturing the adventitious bud in a cluster bud culture medium to form a cluster bud;
4) transferring the cluster buds to a rooting culture medium, and performing rooting culture to obtain grouped cultured seedlings;
5) transplanting the tissue culture seedlings into a mixed matrix for culture after the tissue culture seedlings become seedlings, and obtaining 'orange heart ginger flower' plants through conventional culture; step 1) the pregermination method comprises the following steps: soaking tubers in 1 mg/L6-benzyladenine solution, and culturing in the dark;
step 2) each liter of the adventitious bud induction culture medium comprises 0.5-1.0 mg of thidiazuron, 6-8 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and the rest components are components of an MS culture medium, and the pH value is adjusted to 5.8-6.0;
step 2) the adventitious bud induction conditions are as follows: the culture temperature is 25-28 ℃, and the white light radiation is 30-60 mu mol/m 2 The illumination time is 12-14 hours/day, and the culture is carried out for 7-10 days;
the multiple bud culture medium in the step 3) comprises 8-10 mg of 6-benzyladenine, 0.1-0.3 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar per liter, and the rest components are components of an MS culture medium, and the pH is adjusted to 5.8-6.0;
step 3) the culture conditions of the adventitious buds are as follows: the temperature is 25-28 ℃, the number ratio of red light bulbs to blue light bulbs to white light bulbs is (7-8) to 1:1, and the effective light radiation is 60-90 mu mol/m 2 The illumination time is 14-16 hours/day;
step 4), each liter of the rooting culture medium comprises 0.5-1.5 g of activated carbon, 0.1-0.5 mg of 6-benzyladenine, 0.4-0.8 mg of naphthylacetic acid, 30g of cane sugar and 5g of agar, and the balance of the components are 1/2MS culture medium components, and the pH value is 5.8-6.0;
the rooting culture conditions in the step 4) are as follows: the concentration of carbon dioxide is 800 ppm-1000 ppm during culture, and the culture temperature is 25-28 ℃; the number ratio of red light bulbs, blue light bulbs and white light bulbs is 4-5: 3-5: 1 during culture, and the effective light radiation is 60-90 mu mol/m 2 The illumination time is 14-16 hours/day;
the mixed matrix in the step 5) comprises peat soil and perlite; the mass ratio of the peat soil to the perlite is (4-6) to 1;
the red light wavelength is 650 nm-700 nm, the blue light wavelength is 400 nm-450 nm, and the white light is composite light.
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| CN102187810A (en) * | 2010-03-16 | 2011-09-21 | 中国科学院华南植物园 | Tissue culture propagation method for curcuma soloensis |
| CN107667860A (en) * | 2017-10-18 | 2018-02-09 | 广州普邦园林股份有限公司 | Tissue culture propagation method of alpinia japonica |
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