CN105104187B - 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, and in particular to a kind of strong ball root media of Stigma Croci tissue culture bulb and
Tissue culture method.
Background technology
Stigma Croci (Crocus sativus L.) another name Stigma Croci, Stigma Croci, are the perennial plants of Iridaceae crocus
Thing, is also a kind of common spice.It is mainly distributed on the ground such as Europe, Mediterranean and the Central Asia, BeiJing, China, Shandong, Zhejiang and four
There is cultivation on the ground such as river.Stigma Croci is a kind of rare Chinese medicine, with gynoecium top there are three red stigmas to be used as medicine, and stigma contains west
Saffloside, Stigma Croci aldehyde and the Carotenoids derivant of Stigma Croci bitter principle three, it is various with anticancer, antioxidation and blood fat reducing etc.
Effect, also with good enhancing immunity and anti-tumor activity.
Stigma Croci adopts " two-period form " cultural method China more, i.e., then May by bulb from digging in ground, carry out room
It is interior cultivation and pick flowers, November will pick flowers after bulb plant in ground.Stigma Croci is bred by bulb separation, and breeding coefficient is low,
1.5 left and right.Crocus bulb typically just can bloom in more than 8g, and bulb is bigger, bloom more, and bulb is less, bloom fewer, very
To not blooming, but bottom set plants year by year the big bulb that can be allowed to become to bloom by planting technology.According to statistics below 1g is little
Year by year plantation can be only achieved 6g and plant bulb standard again for bulb Jing 3 years；The bottom set of 1g weights takes 2 years；3-5 weights take 1 year up to this
Standard.In addition, only 5000 mu or so of China plantation Stigma Croci area, account for 20% of market demand or so, it is most of or according to
Bad import.Therefore, Crocus bulb being bred by the way of tissue-culturing rapid propagation, and then produces high-quality Stigma Croci filigree can solve state
The insufficient problem of interior market demand.
Crocus bulb is bred by tissue-culturing rapid propagation and can be combined with detoxification technology, solve virus in Stigma Croci planting process
Yield and quality decline problem that accumulation is caused.Part is reported for the domestic and international research with regard to Stigma Croci tissue-culturing rapid propagation, but
The strong ball root culture of tissue culture bulb always restricts the key of tissue culture bulb quality and transplanting survival rate.Existing Stigma Croci group
Training rapid propagation system by bud induction, subculture, strong sprout, bottom set induce, root culture system acquisition tissue culture bulb diameter often
Less than 0.5cm；In addition, the tissue culture bulb root system for obtaining easily sticks up root phenomenon, Root Absorption culture medium nutrient is affected.These feelings
Condition causes that tissue culture bulb transplanting survival rate is low, and the seedling exercising phase is long, and the grown in field cycle is long.
Application publication number discloses a kind of side of oriental hybrid lily tissue cultured seedling strong sprout for the patent documentation of the A of CN 103733994
Method, including：The bulb of oriental hybrid lily aseptic seedling is taken, scale is placed in inducing culture carries out bud inducement cultivation；Induction is sprouted
Afterwards, bud is connected in strong ball culture medium of taking root and is cultivated, obtain tissue cultured seedling；Wherein, the composition of the strong ball culture medium of taking root
For：MS powder, 4-6g/L；Sucrose, 60-70g/L；Agar, 6-8g/L；Paclobutrazol, 5 × 10-4-5×10-3mmol/L.The invented party
Method can promote expanding or/and increasing weight for tissue cultured seedling bulb, so as to obtain the tissue cultured seedling of stalwartness, shorten the production cycle, save
Cost.
In Stigma Croci tissue culture factorial praluction, how by optimization culture based formulas, the bulb for expanding is cultivated, so as to improve
Tissue culture bulb quality, raising transplanting survival rate, shortening tissue culture bulb, in the grown in field cycle, are technical problems urgently to be resolved hurrily.
The content of the invention
The present invention provides a kind of Stigma Croci tissue culture bulb strong ball root media, can promote Stigma Croci tissue culture expansion of corms and
Take root, improve the quality of tissue culture bulb, improve transplanting survival rate, meet market demand.
To solve above-mentioned technical problem, the invention provides a kind of strong ball root media of Stigma Croci tissue culture bulb, described
Culture medium culture medium based on LS culture medium, contains in every liter of culture medium：Carrageenan 6-8g, white sugar 40-60g, activated carbon
0.1-0.5g, banana meat 20-40g, mashed potato 10-30g, naphthalene acetic acid 0.1-1mg, indolebutyric acid 1-5mg, paclobutrazol 1-5mg,
PH value 5.8-6.0.
The strong ball root media of the Stigma Croci tissue culture bulb of the present invention adopts LS (Linsmaier＆Skoog) culture medium conduct
Most common culture medium MS (Murshige＆Skoog) culture medium basis phase used in basal medium, with tissue culture production
Than LS culture medium eliminates glycine, pyridoxine hydrochloride and nicotinic acid, is suitable for herbal tissue culture.
The preparation method of the banana meat is its sarcocarp of Fructus Musae peeling and taking, to add water and stir into pasty state with blender.
The preparation method of the mashed potato is that Rhizoma Solani tuber osi is cleaned to remove the peel, in mass ratio 1：5 ratio adds water, after boiling
Pasty state is stirred into blender.
Naphthalene acetic acid (1-Naphthaleneacetic acid, NAA) is a kind of colorless solid for being soluble in organic solvent, is
A kind of auximone parahormone, in being usually used in the root of hair powder of commercialization or rooting agent.
Indolebutyric acid (3-Indolybutyric acid, IBA) is a kind of white crystalline solid, is soluble in ethanol etc. organic
Solvent, 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 biological red mould in vivo
The synthesis of element, can be used to cultivate healthy and strong tissue cultured seedling.Have been found to that ball can be promoted in the tissue culture of Bulbus Lilii, Zantedeschia aethiopica Spreng. and gladioluss
Stem expands.
Activated carbon (AC) has very strong adsorptivity to phenols and its oxide, and plant tissue culture has been widely used in
In, the dark situation taken root can be provided in plant tissue culture rooting process, brown stain is prevented, improve the internal soluble protein of culture
With the content of total sugar.
Carrageenan (Carrageenan) is the parent extracting from the red algae zostera marina such as Eucheuma muricatum (Gmel.) Web.Van Bos., Eucheuma gelatinosum, Pelvetia siliquosa Tseng et C. F. Chang
Aqueous colloidal, is white or light brown granule or powder, acts on identical with agar powder (Agar), is commonly used for plant tissue culture solid
As holder in culture medium.
White sugar can support the vigorous growth of most Vitro Plant cultures, therefore by the mark as plant tissue culture
Quasi- carbon source and extensively apply, can to a certain extent reduce the pollution of microorganism.
Preferred scheme, contains in every liter of culture medium：Naphthalene acetic acid 0.4-0.6mg, indolebutyric acid 2-3mg, paclobutrazol 2-
3mg。
The scheme being more highly preferred to, contains in every liter of culture medium：Carrageenan 7g, white sugar 50g, activated carbon 0.2g, banana meat
30g, mashed potato 20g, naphthalene acetic acid 0.5mg, indolebutyric acid 3mg, paclobutrazol 2mg, pH value is 5.8.
The invention provides a kind of tissue culture method of Stigma Croci, comprises the following steps：
(1) Stigma Croci Multiple Buds are taken, in being seeded to inducing culture, inducing culture obtains bulb；
(2) bulb is seeded in described culture medium, cultivates to bulb diameter and be not less than 10mm.
Inducing culture culture medium based on LS culture medium, every liter of inducing culture contains：Naphthalene acetic acid 0.1-
1mg, white sugar 40-60g, agar powder 6-8g, pH value is 5.8.
Preferred scheme, every liter of inducing culture contains：Naphthalene acetic acid 0.5mg, white sugar 50g, agar powder 7g.
In step (1), the condition of inducing culture is：20 ± 2 DEG C of temperature, daily 12h illumination, intensity of illumination 1500lux.
In step (2), condition of culture is：20 ± 2 DEG C of temperature, daily 12h illumination, intensity of illumination 1500lux.
In step (1), inducing culture to bulb is not more than 5mm.
Compared with prior art, beneficial effects of the present invention are：
(1) banana meat that hestening rooting is with the addition of in culture medium prescription of the present invention and the Rhizoma Solani tuber osi for promoting strong sprout, acquisition
Tissue culture bulb rooting efficiency is good, bulb robust growth, and well developed root system, is difficult to stick up root, improves transplanting survival rate.
(2) paclobutrazol for promoting expansion of corms is with the addition of in culture medium of the present invention, the tissue culture bulb diameter of acquisition is reachable
10.5mm, does not about use 2.2 times of strong ball culture medium, improves the quality of tissue culture bulb, shortens the grown in field of bulb
Cycle.
(3) in culture medium of the present invention using carrageenan as solid medium holder, compared with conventional agar powder, no
Only rooting efficiency is good, and low price.
Specific embodiment
The present invention is further explained with reference to specific embodiment.
Embodiment 1
(1) with culture medium：
Every liter of bottom set inducing culture is composed of the following components：LS+ white sugar 50g+NAA 0.5mg+ agar powder 7g, pH=
5.8。
Per liter of strong ball root media is composed of the following components：LS+ white sugar 50g+ banana meat 30g+ mashed potato 20g+ are more
Effect azoles 2mg+IBA 3mg+NAA 0.5mg+ carrageenan 7g+ activated carbon 0.2g, pH=5.8.
(2) the Stigma Croci Multiple Buds of robust growth are chosen, single bud is cut into, bottom set inducing culture is inoculated into, is cultivated
20 ± 2 DEG C of temperature, (12h/d, intensity of illumination 1500lux) culture, induces diameter 5mm or so in 50 days or so under illumination condition
Bottom set.
(3) bottom set for inducing is inoculated into into strong ball root media, 20 ± 2 DEG C of cultivation temperature, under illumination condition
(12h/d, intensity of illumination 1500lux) is cultivated, and bottom set bulb diameter increases to 10mm or so within 30 days or so.
Embodiment 2
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 40g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 2mg+IBA 1mg+NAA 1mg+ carrageenan 7g+ activated carbons
0.2g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 3
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 60g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 3mg+IBA 2mg+NAA 0.8mg+ carrageenans 7g+ are active
Charcoal 0.1g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 4
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 50g+ banana meat 20g+ mashed potato 20g+ paclobutrazol 2mg+IBA 1mg+NAA 1mg+ carrageenan 6g+ activated carbons
0.3g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 5
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 50g+ banana meat 40g+ mashed potato 20g+ paclobutrazol 4mg+IBA 4mg+NAA 0.3mg+ carrageenans 6g+ are active
Charcoal 0.4g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 6
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 50g+ banana meat 30g+ mashed potato 10g+ paclobutrazol 2mg+IBA 5mg+NAA 0.2mg+ carrageenans 8g+ are active
Charcoal 0.5g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 7
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 50g+ banana meat 30g+ mashed potato 30g+ paclobutrazol 5mg+IBA 1mg+NAA 1mg+ carrageenan 8g+ activated carbons
0.2g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 8
Bottom set inducing culture strengthens ball prescription of rooting medium as follows using the culture medium prescription in embodiment 1：With 1L
Meter, LS+ white sugar 40g+ banana meat 30g+ mashed potato 20g+ paclobutrazol 1mg+IBA 1mg+NAA 1mg+ carrageenan 7g+ activated carbons
0.2g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Embodiment 9
If a matched group, bottom set inducing culture using the culture medium prescription in embodiment 1, adopt by strong ball root media
With the culture medium prescription of document report.Strong ball prescription of rooting medium is as follows：In terms of 1L, 1/2MS+ sucrose 30g+6-BA 1.0mg
+ NAA 0.2mg+ agar 7g, pH=5.8.
Bottom set is induced and condition of culture is with embodiment 1.
Result of implementation：
By the data (table 1) for comparing each embodiment, show using culture medium prescription of the present invention in bulb diameter, number of taking root
And three aspects of root length can obtain preferable effect, with embodiment 1 in strong ball prescription of rooting medium it is optimum.By comparing
Embodiment 1 and matched group embodiment 9, show to add in culture medium of the present invention banana meat, Rhizoma Solani tuber osi and paclobutrazol can promote it is western red
Flower tissue culture bulb is taken root and is expanded.
The different embodiments culture of table 1 obtains Stigma Croci tissue culture bulb situation
| Incubation time | Bulb diameter (mm) | Take root and count (bar) | Root length (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 (3)
1. a kind of Stigma Croci tissue culture bulb strengthens ball root media, it is characterised in that the culture medium is based on LS culture medium
Culture medium, contains in every liter of culture medium：Carrageenan 7g, white sugar 50g, activated carbon 0.2g, banana meat 30g, mashed potato 20g, naphthalene
Acetic acid 0.5mg, indolebutyric acid 3mg, paclobutrazol 2mg, pH value 5.8-6.0.
2. a kind of tissue culture method of Stigma Croci, comprises the following steps：
(1) Stigma Croci Multiple Buds are taken, in being seeded to inducing culture, inducing culture obtains bulb；
(2) bulb is seeded in the culture medium described in claim 1, cultivates to bulb diameter and be not less than 10mm；
Inducing culture culture medium based on LS culture medium, every liter of inducing culture contains：It is naphthalene acetic acid 0.1-1mg, white
Sugared 40-60g, agar powder 6-8g, pH value is 5.8；
In step (1), the condition of inducing culture is：20 ± 2 DEG C of temperature, daily 12h illumination, intensity of illumination 1500lux；
In step (2), condition of culture is：20 ± 2 DEG C of temperature, daily 12h illumination, intensity of illumination 1500lux；
In step (1), inducing culture to bulb is not more than 5mm.
3. the tissue culture method of Stigma Croci as claimed in claim 2, it is characterised in that every liter of inducing culture contains：Naphthalene acetic acid
0.5mg, white sugar 50g, agar powder 7g.
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| Application Number | Priority Date | Filing Date | Title |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201510443466.3A CN105104187B (en) | 2015-07-24 | 2015-07-24 | Crocus sativus L. tissue cultured corm strengthening and rooting medium and tissue culture method |
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| CN105104187B true CN105104187B (en) | 2017-04-12 |
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