CN112931219A - Tissue culture and rapid propagation method for cymbidium - Google Patents
Tissue culture and rapid propagation method for cymbidium Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
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Abstract
The invention provides a method for tissue culture and rapid propagation of cymbidium sinense, which comprises the following steps of (1) taking side buds of 1-2 months old on a main stem of a mother plant of the cymbidium sinense as explants; (2) sterilizing, inoculating to an induction culture medium, performing dark culture, performing illumination culture, and performing synchronous induction and multiplication culture; the induction culture medium contains 0.5-0.6mg/L brassin; (3) when 1-1.5 cm of buds are differentiated, taking the buds, inoculating a new induction culture medium with the same formula for subculture; changing the induction culture medium every 5-8 days of culture, and increasing the brassin by 0.1-0.2 mg/L every time of change; (4) inoculating to a rooting culture medium for rooting culture to obtain a cymbidium maculatum seedling; (5) hardening, cleaning and transplanting the seedlings for cultivation; the method can obviously improve the inductivity of the tissue culture and rapid propagation of the cymbidium sinense and effectively shorten the induced rooting period, and the seedling has high growth speed and good root activity, thereby realizing the rapid propagation of the tissue culture seedling of the cymbidium sinense.
Description
Technical Field
The invention relates to the technical field of tissue culture, in particular to a tissue culture and rapid propagation method of spotted orchid.
Background
The variegated orchid is a plant spice, is named as a perfume bag (Pandanus amarylifolius) and is called as a vanilla leaf, resists yin and is favorable for dampness, has few plant diseases and insect pests, and is a vanilla with strong vitality. It is rich in active ingredients such as antioxidant, and unique fragrance, can be applied in various cakes, beverages, and special meals, and has effects of refreshing, clearing pathogenic fire, tranquilizing, relaxing muscles and tendons, and activating collaterals.
In recent years, the planting of the spotted orchid is gradually popularized in the areas of Hainan, Yunnan and the like, but the planting scale of the spotted orchid in Hainan is only about 2 ten thousand mu at present, the spotted orchid seedlings mainly adopt stem cutting propagation as a main part, the spotted orchid seedling propagation has the problems of difficult scale production, uneven individuals, scattered plant shapes after planting, low yield and the like, so that the difficulty of the rapid propagation technology for breaking the spotted orchid seedling tissues is urgently needed, and important conditions are provided for realizing the Hainan spotted orchid planting industry to break through the scale of more than 100 ten thousand mu. However, the existing research on the tissue culture rapid propagation technology of the Spot orchid still has a large blank, and has the problems of low induction rate, low multiplication coefficient, poor root growth activity of tissue culture seedlings, long culture period and the like of the tissue culture rapid propagation of the Spot orchid seedlings, so that the difficulty of popularization of the tissue culture rapid propagation technology of the Spot orchid is limited.
Disclosure of Invention
In view of the above, the invention provides a tissue culture and rapid propagation method of spotted orchid.
The technical scheme of the invention is realized as follows:
the invention provides a tissue culture and rapid propagation method of spotted orchid, which comprises the following steps:
step 1: cutting new lateral buds on main stems of the cymbidium ensifolium stock plant to 1-2 cm to be used as explants;
step 2: sterilizing and disinfecting the explant, inoculating the explant into an induction culture medium, performing dark culture, then performing illumination culture, and performing synchronous induction and multiplication culture; the induction culture medium contains 0.5-0.6mg/L brassin;
and step 3: when the buds are differentiated from the explants, taking the buds, inoculating the buds to a new induction culture medium with the same formula as the step 2 for subculture; replacing a new induction culture medium every 5-8 days after culture, and increasing 0.1-0.2 mg/L of brassinolide in the induction culture medium replaced every time;
and 4, step 4: inoculating the cluster bud plants more than or equal to 2cm into a rooting culture medium for rooting culture until the spotted orchid seedlings with the height of 9-12 cm are obtained; returning the cluster bud plants less than 2cm to the step 3 for subculture;
and 5: and (4) hardening the seedling of the spotted orchid obtained in the step (4), and transplanting and cultivating the seedling.
Further explaining, in the step 2, the explant is sterilized, specifically, the explant is soaked in 0.05-0.1% mercuric chloride solution in which 3-6 drops of tween-20 are dripped by a dripper with the diameter of 1.0mm for 10-15 min, is washed by sterile water for 2-3 times, is soaked in 75% ethanol solution for 30-60 s, is washed by the sterile water for 3-4 times, and is drained to reduce the explant-induced pollution.
More preferably, the induction medium is MS, 0.03-0.07mg/L TDZ, 0.8-1.0 mg/L6-BA, 0.5-0.6mg/L brassin, 0.2-0.8mg/L kinetin, 0.1-0.5mg/LNAA, 1.0-1.5g/L peptone, 8-12g/L white sugar and 4.5-6.0g/L agar;
the rooting culture medium comprises: 1/3MS, 0.8-1.5 mg/L6-BA, 0.05-0.1mg/L NAA, 0.02-0.04mg/L brassin, 5-7g/L banana puree, 10-20g/L white sugar and 7.0-8.0g/L agar.
More preferably, the induction medium is MS, 0.05mg/L TDZ, 0.9 mg/L6-BA, 0.55mg/L brassin, 0.5mg/L kinetin, 0.3mg/L LNAA, 1.3g/L peptone, 10g/L white sugar and 5.2g/L agar;
the rooting culture medium comprises: 1/3MS, 1.2 mg/L6-BA, 0.08mg/L NAA, 0.03mg/L brassin, 6g/L banana puree, 15g/L white sugar and 7.5g/L agar.
Further explaining, in the step 2, after the explants are inoculated to an induction culture medium, dark culture is carried out for 5-7 days at 25 +/-2 ℃; then, the temperature is increased to 26 +/-2 ℃, the illumination intensity is 1000-1500 lx, and the cultivation is carried out for 10-12 d under the condition that the illumination time is 3-5 h/d; and transferring the culture medium into a new induction culture medium with the same formula, and performing synchronous induction and multiplication culture under the conditions of 26 +/-2 ℃, the illumination intensity of 1500-2000 lx and the illumination time of 6-8 h/d.
More preferably, in the step 3, the temperature of the subculture is 28 +/-2 ℃, the illumination intensity is 1500-2000 lx, the illumination time is 10-12 h/d, and the subculture time is 20-24 d; regulating and controlling the condition of subculture, promoting the differentiation and proliferation of the adventitious bud, improving the proliferation coefficient of the adventitious bud and shortening the culture period.
More preferably, in the step 4, the temperature of rooting culture is 28 +/-2 ℃, the illumination intensity is 1500-2500 lux, the illumination time is 12-14 h/d, the rooting culture condition is regulated and controlled, the growth and development of the root system are promoted, and the induced rooting period is shortened.
More preferably, in the step 5, before hardening off, the cymbidium sinense seedlings obtained in the step 4 are transferred to a solid MS culture medium and cultured for 5-7 days, so that the survival rate of transplanting the cymbidium sinense seedlings is further improved.
More preferably, the seedling hardening of the cymbidium seedlings is specifically that the cymbidium seedlings are transferred to the outdoor with bottles and hardened at normal temperature for 5-7 d, and after the seedlings are hardened for 3-5 d by uncovering, seedling transplanting cultivation is carried out.
More preferably, in the step 2-4, the humidity of the synchronous induction and proliferation culture is 55-65%, and the humidity of the rooting culture is 60-80%.
Compared with the prior art, the invention has the beneficial effects that: the invention aims to fully improve the induction rate of the tissue culture and rapid propagation of the spotted orchid, effectively shorten the induced rooting period, increase the growth speed of the seedling and improve the root activity of the tissue culture seedling, thereby realizing the method for rapidly establishing a spotted orchid plant regeneration system in a short period, realizing the rapid propagation of the spotted orchid tissue culture seedling and providing important conditions for the supply and the propagation of the spotted orchid seedling.
(1) According to the invention, the bud induction differentiation of the explant is directly promoted by effectively regulating the explant induction culture conditions, utilizing certain low-temperature dark culture and short-time illumination culture and combining with a corresponding induction culture medium formula, so that independent explant dedifferentiation is not required, the adventitious bud induction rate is improved, the adventitious bud character is ensured, synchronous bud induction differentiation and proliferation culture are realized, and the culture period is shortened.
(2) By selecting the buds with a certain differentiation degree and carrying out subculture, the concentration of brassinon in the induction culture medium is gradually increased, the rapid growth of cluster bud plants is further promoted, and the multiplication coefficient of adventitious buds is remarkably improved while the stable growth quality of seedlings is ensured.
(3) By optimizing the rooting culture medium and adding a certain amount of banana puree by adopting a low-concentration brassin combination, the rooting of the cymbidium maculatum seedlings is effectively promoted, the induced rooting is promoted, the root activity of the cymbidium maculatum tissue culture seedlings is improved, and the survival of the transplanting of the cymbidium maculatum seedlings is facilitated.
Detailed Description
In order to better understand the technical content of the invention, specific examples are provided below to further illustrate the invention.
The experimental methods used in the examples of the present invention are all conventional methods unless otherwise specified.
The materials, reagents and the like used in the examples of the present invention can be obtained commercially without specific description.
Example 1-tissue culture rapid propagation method of spotted orchid seedlings, comprising the following steps:
(1) cutting the newly-born 1-month-old lateral bud on the main stem of the mother plant of the cymbidium goeringii to 2cm to be used as an explant;
(2) sterilizing and disinfecting the explant, inoculating the explant into an induction culture medium, and performing dark culture at 25 +/-2 ℃ for 5 days; culturing for 10 days at 26 + -2 deg.C and illumination intensity of 1000lx and illumination time of 3 h/d; transferring to a new induction culture medium with the same formula, and performing synchronous induction and proliferation culture under the conditions of 26 +/-2 ℃, illumination intensity of 1500lx and illumination time of 6h/d, wherein the humidity is 55%;
the induction culture medium is MS, 0.03mg/L TDZ, 0.8 mg/L6-BA, 0.5mg/L brassin, 0.2mg/L kinetin, 0.1mg/LNAA, 1.0g/L peptone, 8g/L white sugar and 4.5-6.0g/L agar;
(3) when 1cm of buds are differentiated from the explant, the buds are taken and inoculated into a new induction culture medium with the same formula as the step 2, and subcultured for 24d under the conditions that the temperature is 28 +/-2 ℃, the illumination intensity is 1500lx and the illumination time is 10h/d, and the humidity is 55%; changing a new induction culture medium every 8 days of culture, and increasing the brassin in the induction culture medium changed every time by 0.2 mg/L;
(4) inoculating the cluster bud plants more than or equal to 2cm into a rooting culture medium, and carrying out rooting culture under the conditions that the temperature is 28 +/-2 ℃, the illumination intensity is 1500lux, and the illumination time is 12h/d, wherein the humidity is 60 percent until the spotted orchid seedlings with the height of 8cm are obtained; returning the cluster bud plants less than 2cm to the step 3 for subculture;
the rooting culture medium comprises: 1/3MS, 0.8 mg/L6-BA, 0.05mg/L NAA, 0.02mg/L brassin, 5g/L banana puree, 10g/L white sugar and 7.0g/L agar;
(5) and (4) transferring the spotted orchid seedlings obtained in the step (4) to the outdoor for hardening at normal temperature for 5d with a bottle, opening the cover for hardening for 3d, taking out and cleaning, and transplanting and cultivating the seedlings.
Wherein, the sterilization and disinfection of the explant are as follows: soaking the explant in 0.08% mercuric chloride solution dropwise added with 5 drops of Tween-20 for 13min, washing with sterile water for 3 times, soaking in 75% ethanol solution for 50s, washing with sterile water for 4 times, and draining.
Embodiment 2-tissue culture rapid propagation method of spotted orchid seedling, comprising the following steps:
(1) cutting a side bud of 2 months old newly grown on a main stem of a mother plant of the herba Zeylanicae maculatae to 1cm to be used as an explant;
(2) sterilizing and disinfecting the explant, inoculating the explant into an induction culture medium, and performing dark culture at 25 +/-2 ℃ for 7 days; rotating to 26 +/-2 ℃, and culturing for 12 days under the conditions of illumination intensity of 1500lx and illumination time of 5 h/d; transferring to a new induction culture medium with the same formula, and performing synchronous induction and proliferation culture under the conditions of 26 +/-2 ℃, illumination intensity of 2000lx and illumination time of 8h/d, wherein the humidity is 65%;
the induction culture medium is MS, 0.07mg/L TDZ, 1.0 mg/L6-BA, 0.6mg/L brassin, 0.8mg/L kinetin, 0.5mg/LNAA, 1.5g/L peptone, 12g/L white sugar and 6.0g/L agar;
(3) when 1.5cm of buds are differentiated from the explant, inoculating the buds into a new induction culture medium with the same formula as the step 2, and carrying out subculture for 20d under the conditions of the temperature of 28 +/-2 ℃, the illumination intensity of 2000lx and the illumination time of 12h/d, wherein the humidity is 65%; changing a new induction culture medium every 5 days of culture, and increasing the brassin in the induction culture medium changed every time by 0.1 mg/L;
(4) inoculating the cluster bud plants more than or equal to 2cm into a rooting culture medium, and carrying out rooting culture under the conditions that the temperature is 28 +/-2 ℃, the illumination intensity is 2500lux and the illumination time is 14h/d, and the humidity is 80% until the spotted orchid seedlings with the height of 10cm are obtained; returning the cluster bud plants less than 2cm to the step 3 for subculture;
the rooting culture medium comprises: 1/3MS, 1.5 mg/L6-BA, 0.1mg/L NAA, 0.04mg/L brassin, 7g/L banana puree, 20g/L white sugar and 8.0g/L agar;
(5) and (4) transferring the spotted orchid seedlings obtained in the step (4) to an outdoor room temperature with a bottle, hardening the seedlings for 7d, opening the cover, hardening the seedlings for 5d, taking out, cleaning, and transplanting and cultivating the seedlings.
Embodiment 3-tissue culture rapid propagation method of spotted orchid seedling, comprising the following steps:
(1) cutting the side bud of 2 months old newly grown on the main stem of the mother plant of the herba Zeylanicae maculatae to 2cm to be used as an explant;
(2) sterilizing and disinfecting the explant, inoculating the explant into an induction culture medium, and performing dark culture at 25 +/-2 ℃ for 6 days; rotating to 26 +/-2 ℃, and culturing for 11 days under the conditions of illumination intensity of 1500lx and illumination time of 4 h/d; transferring to a new induction culture medium with the same formula, and performing synchronous induction and proliferation culture under the conditions of 26 +/-2 ℃, illumination intensity of 2000lx and illumination time of 7h/d, wherein the humidity is 60%;
the induction culture medium is MS, 0.05mg/L TDZ, 0.9 mg/L6-BA, 0.55mg/L brassin, 0.5mg/L kinetin, 0.3mg/LNAA, 1.3g/L peptone, 10g/L white sugar and 5.2g/L agar;
(3) when 1.3cm of buds are differentiated from the explant, inoculating the buds into a new induction culture medium with the same formula as the step 2, and carrying out subculture for 23d under the conditions that the temperature is 28 +/-2 ℃, the illumination intensity is 2000lx and the illumination time is 11h/d, and the humidity is 60%; changing a new induction culture medium every 6 days of culture, and increasing the brassin in the induction culture medium changed every time by 0.15 mg/L;
(4) inoculating the cluster bud plants more than or equal to 2cm into a rooting culture medium, and carrying out rooting culture under the conditions that the temperature is 28 +/-2 ℃, the illumination intensity is 2500lux and the illumination time is 13h/d, and the humidity is 70 percent until the spotted orchid seedlings with the height of 9cm are obtained; returning the cluster bud plants less than 2cm to the step 3 for subculture;
the rooting culture medium comprises: 1/3MS, 1.2 mg/L6-BA, 0.08mg/L NAA, 0.03mg/L brassin, 6g/L banana puree, 15g/L white sugar and 7.5g/L agar;
(5) and (4) transferring the spotted orchid seedlings obtained in the step (4) to the outdoor for hardening at normal temperature for 6d with a bottle, opening the cover for hardening for 4d, taking out and cleaning, and transplanting and cultivating the seedlings.
Example 4-this example differs from example 3 in that: before hardening off seedlings, the obtained herba Epilobii Himalayan seedlings are transferred to a solid MS culture medium, cultured for 6d and then transferred to the outdoor to be hardened off at normal temperature for 6 d.
Comparative example 1-this comparative example differs from example 3 in that: in the step 2, after the explant is inoculated to an induction culture medium, the explant is directly transferred to a new induction culture medium with the same formula after being cultured in dark at the temperature of 28 +/-2 ℃ for 17d, and the explant is synchronously induced and cultured in a proliferation way under the conditions of 26 +/-2 ℃, the illumination intensity of 2000lx and the illumination time of 7 h/d.
Comparative example 2-this comparative example differs from example 3 in that: the formula of the induction culture medium is adjusted as follows: 0.1mg/L, and the content of the brassin is not changed in the substrate culture process.
Comparative example 3-this comparative example differs from example 3 in that: the formula of the induction culture medium is adjusted as follows: 0.9mg/L, and in the process of subculture, the content of the brassinolide is increased by 0.1mg/L after replacing the induction culture medium every 6 days.
Comparative example 4-this comparative example differs from example 3 in that: the brassin content in the rooting medium formula is adjusted to be 0.06 mg/L.
Comparative example 5-this comparative example differs from example 3 in that: the formulation of the rooting culture medium is adjusted without adding brassin, and the content of the banana puree is increased to 10 g/L.
And (3) determining the effect of tissue culture of the spotted orchid seedlings according to the tissue culture and rapid propagation method of the spotted orchid seedlings.
The experimental method comprises the following steps: taking the comparative examples in examples 1-3 and comparative examples 1-3 as treatment groups, each treatment group is repeated for 3 times, the blank control is culture by taking MS as a basic culture medium, the bud induction rate, the proliferation multiple, the root induction period and the root growth condition of the herba Zealand tissue culture seedlings of each experimental group are respectively counted, and the results are shown in the following table:
the calculation method comprises the following steps: adventitious bud induction rate (%) × 100% (number of induced bud explants/number of inoculations); the multiplication times of the adventitious buds are equal to the multiplication adventitious buds number/inoculation buds number;
as can be seen from the above table, the tissue culture and rapid propagation method of the cymbidium maculatum seedlings can realize the adventitious bud induction rate of more than 90%, meanwhile, the multiplication times of the adventitious buds are obviously increased, while in comparative examples 1-3, the induction rate of the adventitious buds is reduced, the multiplication times are reduced, and meanwhile, the period from induction to rooting is obviously prolonged; the invention shows that the induction culture conditions of the explants are effectively controlled and the formula of the induction culture medium is optimized, so that the adventitious bud induction rate is effectively improved, the multiplication coefficient is obviously improved, and the culture period of the tissue culture seedlings of the cymbidium maculatum is shortened; in comparative examples 4-5, the induced rooting period is prolonged, the length of the adventitious roots is shortened, and the root system activity is reduced, which shows that the method can promote induced rooting and improve the root system activity of the cymbidium maculatum tissue culture seedling by optimizing the rooting culture medium, adopting low-concentration brassin and adding a certain amount of banana puree.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (10)
1. A tissue culture and rapid propagation method of spotted orchid is characterized in that: the method comprises the following steps:
step 1: cutting new lateral buds on main stems of the cymbidium ensifolium stock plant to 1-2 cm to be used as explants;
step 2: sterilizing and disinfecting the explant, inoculating the explant into an induction culture medium, performing dark culture, then performing illumination culture, and performing synchronous induction and multiplication culture; the induction culture medium contains 0.5-0.6mg/L brassin;
and step 3: when the buds are differentiated from the explants, taking the buds, inoculating the buds to a new induction culture medium with the same formula as the step 2 for subculture; replacing a new induction culture medium every 5-8 days after culture, and increasing 0.1-0.2 mg/L of brassinolide in the induction culture medium replaced every time;
and 4, step 4: inoculating the cluster bud plants more than or equal to 2cm into a rooting culture medium for rooting culture until the spotted orchid seedlings with the height of 9-12 cm are obtained; returning the cluster bud plants less than 2cm to the step 3 for subculture;
and 5: and (4) hardening the seedling of the spotted orchid obtained in the step (4), and transplanting and cultivating the seedling.
2. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: and 2, sterilizing and disinfecting the explant, specifically, dipping the explant in 0.05-0.1% mercuric chloride solution dropwise added with 3-6 drops of Tween-20 for 10-15 min, washing with sterile water for 2-3 times, dipping in 75% ethanol solution for 30-60 s, washing with sterile water for 3-4 times, and draining.
3. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: the induction culture medium is MS, 0.03-0.07mg/L TDZ, 0.8-1.0 mg/L6-BA, 0.5-0.6mg/L brassin, 0.2-0.8mg/L kinetin, 0.1-0.5mg/LNAA, 1.0-1.5g/L peptone, 8-12g/L white sugar and 4.5-6.0g/L agar; the rooting culture medium comprises: 1/3MS, 0.8-1.5 mg/L6-BA, 0.05-0.1mg/L NAA, 0.02-0.04mg/L brassin, 5-7g/L banana puree, 10-20g/L white sugar and 7.0-8.0g/L agar.
4. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: the induction culture medium is MS, 0.05mg/L TDZ, 0.9 mg/L6-BA, 0.55mg/L brassin, 0.5mg/L kinetin, 0.3mg/LNAA, 1.3g/L peptone, 10g/L white sugar and 5.2g/L agar; the rooting culture medium comprises: 1/3MS, 1.2 mg/L6-BA, 0.08mg/L NAA, 0.03mg/L brassin, 6g/L banana puree, 15g/L white sugar and 7.5g/L agar.
5. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: in the step 2, after the explant is inoculated to an induction culture medium, dark culture is carried out for 5-7 days at 25 +/-2 ℃; then, the temperature is increased to 26 +/-2 ℃, the illumination intensity is 1000-1500 lx, and the cultivation is carried out for 10-12 d under the condition that the illumination time is 3-5 h/d; and transferring the culture medium into a new induction culture medium with the same formula, and performing synchronous induction and multiplication culture under the conditions of 26 +/-2 ℃, the illumination intensity of 1500-2000 lx and the illumination time of 6-8 h/d.
6. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: in the step 3, the temperature of the subculture is 28 +/-2 ℃, the illumination intensity is 1500-2000 lx, the illumination time is 10-12 h/d, and the subculture time is 20-24 d.
7. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: in the step 4, the temperature of rooting culture is 28 +/-2 ℃, the illumination intensity is 1500-2500 lux, and the illumination time is 12-14 h/d.
8. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: before hardening, the cymbidium seedlings obtained in the step 4 are transferred to a solid MS culture medium and cultured for 5-7 days.
9. The tissue culture and rapid propagation method of the spotted orchid according to claim 8, which is characterized in that: and the seedling hardening of the cymbidium seedlings is specifically realized by turning the cymbidium seedlings with bottles to outdoor normal temperature for hardening for 5-7 d, uncovering and hardening for 3-5 d, and transplanting and cultivating seedlings.
10. The tissue culture and rapid propagation method of the spotted orchid according to claim 1, which is characterized in that: in the step 2-4, the humidity of synchronous induction and proliferation culture is 55-65%, and the humidity of rooting culture is 60-80%.
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| CN115119747A (en) * | 2022-04-30 | 2022-09-30 | 中国热带农业科学院海口实验站 | Leaf tissue culture medium of cymbidium maculatum and tissue culture rapid propagation method |
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