CN105104187A - Crocus sativus L. tissue cultured corm strengthening and rooting medium and tissue culture method - Google Patents
Crocus sativus L. tissue cultured corm strengthening and rooting medium and tissue culture method Download PDFInfo
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a Crocus sativus L. tissue cultured corm strengthening and rooting medium and a tissue culture method. The provided medium employs an LS medium as a basic medium, 1L of the medium comprises 6-8g of carrageenan, 40-60 parts of white sugar, 0.1-0.5 part of active carbon, 20-40 parts of banana flesh, 10-30g of mashed potato, 0.1-1mg of naphthylacetic acid, 1-5mg of indolebutyric acid and 1-5mg of paclobutrazol, and the pH value is 5.8-6.0. The provided medium can promote strengthening and rooting of Crocus sativus L. tissue cultured corm, root warping is not easy, and the transplanting survival rate is raised. The tissue culture method comprises the following steps: firstly, Crocus sativus L. multiple shoots are taken and inoculated into an induction medium and a corm is obtained after induction culture; secondly, the corm is inoculated in the provided medium, and the corm is cultured until the diameter is not less than 10mm. The diameter of the Crocus sativus L. tissue cultured corm cultured through the method is 2.2 times of a diameter of the Crocus sativus L. corm cultured without the strengthening medium, the tissue cultured corm quality is raised, and the field growth cycle of corms is shortened.
Description
Technical field
The invention belongs to field of plant tissue culture, be specifically related to a kind of west safflower group training bulb and strengthen ball root media and tissue culture method.
Background technology
West safflower (CrocussativusL.) calls safflower, Crocus sativus, is the perennial plant of Iridaceae crocus, is also a kind of common spices.Mainly be distributed in the ground such as Europe, Mediterranean and the Central Asia, there is cultivation on the ground such as BeiJing, China, Shandong, Zhejiang and Sichuan.West safflower is a kind of rare traditional Chinese medicine, with gynoecium top, there are three red column caps to be used as medicine, column cap contains crocin, west safflower aldehyde and west safflower bitter principle three Carotenoids derivative, there is the multiple efficacies such as anticancer, anti-oxidant and reducing blood lipid, also there is good develop immunitypty and antitumor activity.
West safflower at China's many employings " two-period form " cultivation method, namely then May by kind of ball from digging in ground, carry out indoor culture and pick flowers, the kind ball after picking flowers is planted in ground by November.West safflower is bred by bulb separation, and reproduction coefficient is low, about 1.5.Crocus bulb generally just can be bloomed at more than 8g, and bulb is larger, and bloom more, bulb is less, blooms fewer, does not even bloom, but bottom set is planted year by year by planting technology and can be made it to become the large bulb that can bloom.Below 1g bottom set was planted year by year through 3 years and just can be reached 6g and heavily plant bulb standard according to statistics; The bottom set that 1g is heavy takes 2 years; What 3-5 was heavy take 1 year reaches this standard.In addition, China's plantation safflower area only about 5000 mu, accounts for about 20% of market demand, major part or dependence on import.Therefore, adopt the mode of tissue-culturing rapid propagation to breed Crocus bulb, and then production high-quality west safflower filigree can solve the under-supply problem of domestic market demand amount.
In conjunction with detoxification technology, virus in west safflower planting process can also be solved and accumulates the Yield and qualities decline problem caused by tissue-culturing rapid propagation breeding Crocus bulb.Both at home and abroad about the existing part report of research of west safflower tissue-culturing rapid propagation, but the strong ball culture of rootage of group training bulb is the key of restriction group training bulb quality and transplanting survival rate always.The group that existing west safflower tissue-culturing rapid propagation system is obtained by bud inducement, subculture, strong sprout, bottom set induction, culture of rootage system trains bulb diameter often less than 0.5cm; In addition, the group training bulb root system of acquisition easily sticks up root phenomenon, affects Root Absorption medium nutrient.These situations cause group training bulb transplanting survival rate low, and the hardening phase is long, and the grown in field cycle is long.
Application publication number is the method that patent document discloses a kind of oriental hybrid lily plantlet in vitro strong sprout of CN103733994A, comprising: the bulb getting oriental hybrid lily aseptic seedling, scale is placed in inducing culture and carries out bud inducement cultivation; Bud grafting is cultivated after sprouting by induction to strong ball medium of taking root, and obtains plantlet in vitro; Wherein, to take root described in the consisting of of strong ball medium: MS powder, 4-6g/L; Sucrose, 60-70g/L; Agar, 6-8g/L; Paclobutrazol, 5 × 10
-4-5 × 10
-3mmol/L.This inventive method can promote expanding of plantlet in vitro bulb or/and increase weight, thus can obtain healthy and strong plantlet in vitro, shortens the production cycle, cost-saving.
In west safflower group training factorial praluction, how by optimization culture based formulas, cultivate the bulb expanded, thus improvement group training bulb quality, raising transplanting survival rate, shortening group training bulb are in the grown in field cycle, are technical problems urgently to be resolved hurrily.
Summary of the invention
The invention provides a kind of west safflower group training bulb and strengthen ball root media, can promote that west safflower group is trained expansion of corms and takes root, the quality of improvement group training bulb, improves transplanting survival rate, meets market demand.
For solving the problems of the technologies described above, the invention provides a kind of west safflower group training bulb and strengthen ball root media, described medium is medium based on LS medium, contain in often liter of medium: carragheen 6-8g, white sugar 40-60g, active carbon 0.1-0.5g, banana meat 20-40g, mashed potato 10-30g, methyl α-naphthyl acetate 0.1-1mg, indolebutyric acid 1-5mg, paclobutrazol 1-5mg, pH value 5.8-6.0.
West safflower group training bulb of the present invention is strengthened ball root media and is adopted medium based on LS (Linsmaier & Skoog) medium, with group train produce in use compared with the most general medium MS (Murshige & Skoog) medium basis, LS medium eliminates glycine, puridoxine hydrochloride and nicotinic acid, is suitable for herbal tissue cultures.
The preparation method of described banana meat is its pulp of banana peeling and taking, adds water to stir into pasty state with agitator.
The preparation method of described mashed potato cleans peeling potato, and the ratio of 1:5 adds water in mass ratio, boils rear agitator and stirs into pasty state.
Methyl α-naphthyl acetate (1-Naphthaleneaceticacid, NAA) is a kind of colorless solid being soluble in organic solvent, is a kind of auximone parahormone, is usually used in commercial root of hair powder or rooting agent.
Indolebutyric acid (3-Indolybutyricacid, IBA) is a kind of white crystalline solid, is soluble in the organic solvents such as ethanol, is a kind of auximone parahormone, promotes plant root system development.
Paclobutrazol (Paclobutrazol) is a kind of plant growth retardent of high-efficiency low-toxicity, suppresses the synthesis of organism inner gibberellin, can be used for cultivating healthy and strong plantlet in vitro.Be proved and can have promoted expansion of corms in the group training of lily, common calla and sword lily.
Active carbon (AC) has very strong adsorptivity to phenols and oxide thereof, be widely used in Plant Tissue Breeding, the dark situation of taking root can be provided in Plant Tissue Breeding rooting process, prevent brown stain, improve the content cultivating soluble protein and total reducing sugar in body.
Carragheen (Carrageenan) is the hydrophilic colloid extract from the red algae sea grass such as Eucheuma, agar, pelvetia silquosa; for white or light brown particle or powder; act on identical with agar powder (Agar), be often used in plant tissue culture solid culture medium as holder.
White sugar can support the vigorous growth of most Vitro Plant culture, and therefore the extensive use by the standard carbon source as Plant Tissue Breeding, can reduce the pollution of microorganism to a certain extent.
Preferred scheme, contains in often liter of medium: methyl α-naphthyl acetate 0.4-0.6mg, indolebutyric acid 2-3mg, paclobutrazol 2-3mg.
More preferred scheme, contains in often liter of medium: carragheen 7g, white sugar 50g, active carbon 0.2g, banana meat 30g, mashed potato 20g, methyl α-naphthyl acetate 0.5mg, indolebutyric acid 3mg, paclobutrazol 2mg, pH value is 5.8.
The invention provides the tissue culture method of a kind of west safflower, comprise the following steps:
(1) get west safflower Multiple Buds, be seeded in inducing culture, Fiber differentiation obtains bulb;
(2) described bulb is seeded in described medium, is cultured to bulb diameter and is not less than 10mm.
Described inducing culture is medium based on LS medium, and often liter of inducing culture contains: methyl α-naphthyl acetate 0.1-1mg, white sugar 40-60g, agar powder 6-8g, and pH value is 5.8.
Preferred scheme, often liter of inducing culture contains: methyl α-naphthyl acetate 0.5mg, white sugar 50g, agar powder 7g.
In step (1), the condition of Fiber differentiation is: temperature 20 ± 2 DEG C, 12h illumination every day, intensity of illumination 1500lux.
In step (2), condition of culture is: temperature 20 ± 2 DEG C, 12h illumination every day, intensity of illumination 1500lux.
In step (1), Fiber differentiation is not more than 5mm to bulb.
Compared with prior art, beneficial effect of the present invention is:
(1) with the addition of the banana meat of hestening rooting in culture medium prescription of the present invention and promote the potato in strong sprout, the group training bulb rooting efficiency of acquisition is good, bulb robust growth, and well developed root system, not easily sticks up root, improves transplanting survival rate.
(2) with the addition of the paclobutrazol promoting expansion of corms in medium of the present invention, the group training bulb diameter of acquisition can reach 10.5mm, is about 2.2 times that do not use strong ball medium, improves the quality of group training bulb, shorten the grown in field cycle of bulb.
(3) adopt carragheen as solid culture medium holder in medium of the present invention, compared with conventional agar powder, not only rooting efficiency is good, and low price.
Embodiment
The present invention is explained further below in conjunction with specific embodiment.
Embodiment 1
(1) medium is joined:
Often liter of bottom set inducing culture is composed of the following components: LS+ white sugar 50g+NAA0.5mg+ agar powder 7g, pH=5.8.
Often liter of strong ball root media is composed of the following components: LS+ white sugar 50g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 2mg+IBA3mg+NAA0.5mg+ carragheen 7g+ active carbon 0.2g, pH=5.8.
(2) choose the west safflower Multiple Buds of robust growth, be cut into single bud, be inoculated into bottom set inducing culture, cultivation temperature 20 ± 2 DEG C, under illumination condition, (12h/d, intensity of illumination 1500lux) cultivates, and within about 50 days, induces the bottom set of diameter about 5mm.
(3) bottom set induced is inoculated into strong ball root media, cultivation temperature 20 ± 2 DEG C, under illumination condition, (12h/d, intensity of illumination 1500lux) cultivates, and within about 30 days, bottom set bulb diameter increases to about 10mm.
Embodiment 2
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, and strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 40g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 2mg+IBA1mg+NAA1mg+ carragheen 7g+ active carbon 0.2g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 3
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 60g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 3mg+IBA2mg+NAA0.8mg+ carragheen 7g+ active carbon 0.1g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 4
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, and strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 50g+ banana meat 20g+ mashed potato 20g+ paclobutrazol 2mg+IBA1mg+NAA1mg+ carragheen 6g+ active carbon 0.3g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 5
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 50g+ banana meat 40g+ mashed potato 20g+ paclobutrazol 4mg+IBA4mg+NAA0.3mg+ carragheen 6g+ active carbon 0.4g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 6
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 50g+ banana meat 30g+ mashed potato 10g+ paclobutrazol 2mg+IBA5mg+NAA0.2mg+ carragheen 8g+ active carbon 0.5g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 7
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, and strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 50g+ banana meat 30g+ mashed potato 30g+ paclobutrazol 5mg+IBA1mg+NAA1mg+ carragheen 8g+ active carbon 0.2g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 8
Bottom set inducing culture adopts the culture medium prescription in embodiment 1, and strong ball prescription of rooting medium is as follows: in 1L, LS+ white sugar 40g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 1mg+IBA1mg+NAA1mg+ carragheen 7g+ active carbon 0.2g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Embodiment 9
If a control group, bottom set inducing culture adopts the culture medium prescription in embodiment 1, and strong ball root media adopts the culture medium prescription of bibliographical information.Strong ball prescription of rooting medium is as follows: in 1L, 1/2MS+ sucrose 30g+6-BA1.0mg+NAA0.2mg+ agar 7g, pH=5.8.
Bottom set induction and condition of culture are with embodiment 1.
Result of implementation:
By the data (table 1) of each embodiment of comparison, show that application culture medium prescription of the present invention can obtain good effect in bulb diameter, take root number and long three of root, optimum with the strong ball prescription of rooting medium in embodiment 1.By comparing embodiment 1 and control group embodiment 9, show that adding banana meat, potato and paclobutrazol in medium of the present invention can promote that west safflower group trains taking root and expanding of bulb.
The different embodiment of table 1 is cultivated and is obtained west safflower group training bulb situation
| Incubation time | Bulb diameter (mm) | Take root and count (bar) | Root long (cm) | |
| Example 1 | 80d | 10.5 | 18.5 | 2.1 |
| Example 2 | 80d | 8.1 | 15.3 | 1.6 |
| Example 3 | 80d | 8.3 | 14.4 | 1.8 |
| Example 4 | 80d | 10.2 | 16.5 | 2.0 |
| Example 5 | 80d | 8.8 | 15.2 | 1.5 |
| Example 6 | 80d | 9.5 | 17.1 | 1.7 |
| Example 7 | 80d | 10.1 | 17.2 | 1.6 |
| Example 8 | 80d | 10.3 | 18.2 | 1.9 |
| Example 9 | 80d | 7.3 | 12.4 | 1.2 |
Claims (9)
1. a west safflower group training bulb strengthens ball root media, it is characterized in that, described medium is medium based on LS medium, contain in often liter of medium: carragheen 6-8g, white sugar 40-60g, active carbon 0.1-0.5g, banana meat 20-40g, mashed potato 10-30g, methyl α-naphthyl acetate 0.1-1mg, indolebutyric acid 1-5mg, paclobutrazol 1-5mg, pH value 5.8-6.0.
2. medium as claimed in claim 1, is characterized in that, contain in often liter of medium: methyl α-naphthyl acetate 0.4-0.6mg, indolebutyric acid 2-3mg, paclobutrazol 2-3mg.
3. medium as claimed in claim 1, is characterized in that, contain in often liter of medium: carragheen 7g, white sugar 50g, active carbon 0.2g, banana meat 30g, mashed potato 20g, methyl α-naphthyl acetate 0.5mg, indolebutyric acid 3mg, paclobutrazol 2mg.
4. a tissue culture method for west safflower, comprises the following steps:
(1) get west safflower Multiple Buds, be seeded in inducing culture, Fiber differentiation obtains bulb;
(2) described bulb is seeded in the arbitrary described medium of claim 1-3, is cultured to bulb diameter and is not less than 10mm.
5. the tissue culture method of west safflower as claimed in claim 4, it is characterized in that, described inducing culture is medium based on LS medium, and often liter of inducing culture contains: methyl α-naphthyl acetate 0.1-1mg, white sugar 40-60g, agar powder 6-8g, and pH value is 5.8.
6. the tissue culture method of west safflower as claimed in claim 5, it is characterized in that, often liter of inducing culture contains: methyl α-naphthyl acetate 0.5mg, white sugar 50g, agar powder 7g.
7. the tissue culture method of west safflower as claimed in claim 4, it is characterized in that, in step (1), the condition of Fiber differentiation is: temperature 20 ± 2 DEG C, 12h illumination every day, intensity of illumination 1500lux.
8. the tissue culture method of west safflower as claimed in claim 4, it is characterized in that, in step (2), condition of culture is: temperature 20 ± 2 DEG C, 12h illumination every day, intensity of illumination 1500lux.
9. the tissue culture method of west safflower as claimed in claim 4, it is characterized in that, in step (1), Fiber differentiation is not more than 5mm to bulb.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108419679A (en) * | 2018-05-31 | 2018-08-21 | 浙江大学 | The tissue culture method of west safflower |
| CN109792925A (en) * | 2019-03-18 | 2019-05-24 | 杭州市农业科技教育总站 | West safflower bulbil breeds bulb method |
| CN109964655A (en) * | 2018-12-21 | 2019-07-05 | 杭州师范大学 | West safflower is quickly picked flowers method |
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| CN102239805A (en) * | 2011-06-28 | 2011-11-16 | 浙江省萧山棉麻研究所 | Tissue-culture quick breeding method for iris tectorum |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108419679A (en) * | 2018-05-31 | 2018-08-21 | 浙江大学 | The tissue culture method of west safflower |
| CN109964655A (en) * | 2018-12-21 | 2019-07-05 | 杭州师范大学 | West safflower is quickly picked flowers method |
| CN109964655B (en) * | 2018-12-21 | 2022-01-14 | 杭州师范大学 | Rapid flower picking method for crocus sativus |
| CN109792925A (en) * | 2019-03-18 | 2019-05-24 | 杭州市农业科技教育总站 | West safflower bulbil breeds bulb method |
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