JPH01247083A - Issue-culture of asparagus - Google Patents

Abstract

PURPOSE:To proliferate seedlings of asparagus in high efficiency, by culturing a globular embryo in a medium containing >=0.2M of sucrose or >=0.1M of sucrose and >=0.05M of glucose to obtain a banana-type embryo. CONSTITUTION:An indefinite embryo of globular embryo stage originated from an embryo, shoot apex, phyllodium, subterranean stem, other tissues or cells of asparagus or a callus produced from the above tissue is subjected to tissue culture in a medium containing >=0.2M of sucrose (in the case of the single use of sucrose) or >=0.1M of sucrose and >=0.05M of glucose (in the case of combined use of sucrose and glucose). The medium used in the above procedure is preferably produced by adding a plant hormone such as naphthaleneacetic acid, vitamins such as biotin and amino acids such as glycine to a medium such as Nitsch-Nitsch medium, Linsmeier-skoog medium, etc. The indefinite embryo can be grown in high efficiently and converted to a banana-type embryo. A large amount of high-quality seedlings can be proliferated from a tissue of asparagus in high efficiency by this process.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to asparagus tissue culture, and particularly to a tissue culture method suitable for promoting the growth of asparagus somatic embryos.

[Conventional technology]

Asparagus has traditionally been propagated by seeding or division. Asparagus is dioecious,
Male plants are said to be better in terms of quality and cultivation management. Furthermore, since it is a crop with large genetic variations, propagation by sowing leads to the contamination of female plants and also causes differences in yield and quality among plants, which poses a serious problem for growers. In addition, propagation by division requires a lot of manpower and a long period of time, is extremely inefficient, and has a very high risk of transmitting viral diseases.

In order to improve the above-mentioned problems, mass propagation of superior strain clones by tissue culture has recently attracted attention. Usually, for mass propagation of asparagus, IJI tissue is cultivated,
After propagating a large number of shoots, each shoot is transplanted into a rooting medium and becomes a seedling through differentiation of adventitious roots. However, since the rooting rate is generally low, there is a problem that the propagation efficiency is low. In addition, attempts have been made to culture tissue fragments to form callus, and then use somatic embryos produced from the callus to mass multiply (for example,
1st Year Horticultural Society Autumn Conference Research Presentation Abstracts P210-211
, November 23-25, 1988, Ryukyu University, 1988
Annual Horticultural Society Research Presentation Abstracts P254-255, 1988
．． 10.7-9 at Kyushu University). Although growing somatic embryos solves the problem of low rooting rate, the problem is that most isolated somatic embryos develop callus or malformation during culture.

In view of the above, the development of a method to efficiently promote the normal growth of somatic embryos is awaited.

[Problem to be solved by the invention]

The present inventors recognized that the conventional asparagus tissue culture method has the above-mentioned problems, and attempted to efficiently propagate asparagus seeds by culturing tissue using a new method different from the conventional method. We considered the method.

[Means to solve the problem]

As a result, the present inventors discovered a substance that acts on somatic embryos of asparagus to promote growth, and based on these findings, discovered a method for efficiently propagating asparagus seeds and seedlings. That is, according to the present invention, a banana-shaped embryo is obtained by culturing a spherical embryo obtained by culturing asparagus tissue or cells using a medium containing 0.2 M or more of sucrose. Banana-shaped embryos are obtained by culturing asparagus tissue culture methods and spherical embryos obtained by culturing asparagus tissues or cells in a medium containing shakerose 0.1 M or more and glucose 0.05 M or more. A method for culturing asparagus tissue is provided.

Plants to which the method of the present invention can be applied include asparagus.

The tissue culture method of the present invention is carried out using asparagus somatic embryos at the globular stage. Specifically, somatic embryos at the spherical embryo stage derived from embryos, shoot tips, aggregates, rhizomes, other tissues, cells, or calluses derived therefrom can be exemplified.

In the tissue culture method according to the present invention, tissue culture is performed after adding a specific concentration of sucrose or specific concentrations of sucrose and glucose to the culture medium.

The tissue culture method according to the present invention will be described in detail below.

In the tissue culture method of the present invention, sucrose alone or sucrose and glucose are added to a culture medium at specific concentrations, and tissue culture is performed.

The concentration of the compound added to the medium is 0.2M or more for sucrose alone, 0.1M or more for sucrose and 0.05M or more for glucose when used in combination with glucose, and at this concentration, the growth of the somatic embryo is In good condition.

The medium used in the present invention is a medium containing inorganic components and a carbon source as essential components, to which are added plant hormones and vitamins, and further added with amino acids as necessary.

Inorganic components of the medium include inorganic salts containing elements such as nitrogen, phosphorus, potassium, sodium, calcium, magnesium, sulfur, iron, manganese, zinc, boron, molybdenum, chlorine, iodine, cobalt, etc. Specifically, potassium nitrate, sodium nitrate, ammonium nitrate, potassium chloride, calcium chloride, potassium hydrogen phosphate, sodium dihydrogen phosphate, magnesium sulfate,
Examples include compounds such as magnesium chloride, sodium sulfate, ferrous sulfate, ferric sulfate, manganese sulfate, copper sulfate, sodium molybdate, molybdenum trioxide, potassium iodide, zinc sulfate, boric acid, and cobalt chloride.

In addition to the above-mentioned sucrose and glucose, other carbohydrates and their derivatives, organic acids such as fatty acids, and primary alcohols such as ethanol may be added as carbon sources for the medium.

Examples of plant hormones in the medium include naphthalene acetic acid (NAA), indole acetic acid (IAA), p-chlorophenoxyacetic acid, and 2,4-dichlorophenoxyacetic acid (2.
, 4-D), indolebutyric acid (IBM), and auxins such as derivatives thereof and benzyladenine (BA)
Examples include cytokinins such as , kinetin, and zeatin.

The vitamins in the medium include biotin, thiamine (vitamin B), pyridoxine (vitamin B6), pyridoxal, pyridoxamine, calcium pantothenate,
Ascorbic acid (vitamin C), inositol, nicotinic acid, nicotinamide and riboflavin (vitamin B
, ), etc.

Examples of amino acids in the medium include glycine, alanine, glutamine, cysteine, phenylalanine, and lysine.

The medium of the present invention usually contains about 0.1 of the inorganic components.
μH to about 100mM, the plant hormones to about 0
．． 01■/l or about 150mg/j! and the above amino acids at 0 to about 1000 ■/I! , it is desirable to use it together.

Specifically, the medium used for tissue culture according to the present invention may be a conventionally known culture medium used for plant tissue culture, such as Murashige Skoog ('62
) [Murashige & Skoog] medium,
Linsmeyer Skoog (RM-1965) [Li
nsmaier & Skoog] medium, white (
'63) [White] medium, Gamborg [Gam
borg]'s 8-5 medium, Sankata-9 medium, -1-7
An example of a medium prepared by adding the carbon source and plant hormones described above to a medium such as Chi. Among them, especially Ecchi Ecchi, Linsmeyer Skoog, Murashige
A medium prepared using Skoog's medium is preferred. Regarding the composition of the conventionally known culture medium mentioned above, for example, "New Plant Tissue Culture J" by Yukinai, Nakagome, and Furuya, p.386-p.3
91, Asakura Shoten, 1979. The medium that can be used in the present invention is a liquid medium or a solid medium containing usually 0.1 to 2% of a gelling agent such as agar or Gelrite.

According to the present invention, somatic embryos of asparagus can be grown efficiently and can be transitioned from a globular embryo stage to a banana-shaped embryo. Furthermore, when the plants obtained according to the present invention are cultivated normally, they can grow into healthy plants with constant properties, so asparagus seeds and seedlings can be efficiently propagated.

〔Example〕

Examples 1 to 2 Somatic embryos corresponding to spherical embryos were collected from Asparagus callus under a stereoscopic microscope, and the resulting somatic embryos were grown in MS medium containing 0.2 or 0.2 sucrose as the base of the nitrogen source. 3
M, biotin 0.05■/2, folic acid 0.5■/l,
Contains glutamine 2mM, agar 7g/l, pH 5,
Prepare a sterile Murashigeskoog solid medium from step 6, place 20 asparagus somatic embryos on it, and incubate at 25°.
When cultured for 2 weeks under C2 darkness, the results shown in Table 1 were obtained as the percentage of growth from spherical embryos to banana-shaped embryos (sody embryo growth rate).

The growth rate of somatic embryos in both mediums supplemented with 0.2 or 0.3 M of sucrose increased compared to Comparative Example 1.

Example 3 Asparagus somatic embryos were cultured in the same manner as in Example 1 except that 0.1 M of sucrose and 0.1 M of glucose were added, and the results shown in Table 1 were obtained. At this time, the growth rate of somatic embryos increased compared to Comparative Example 1.

Example 4 Asparagus somatic embryos were cultured in the same manner as in Example 1 except that 0.2 M of sucrose and 0.1 M of glucose were added, and the results shown in Table 1 were obtained. At this time, the growth rate of somatic embryos increased compared to Comparative Example 1.

Example 5 Asparagus somatic embryos were cultured in the same manner as in Example 1 except that 0.1 M of sucrose and 0.2 M of glucose were added, and the results shown in Table 1 were obtained. At this time, the growth rate of somatic embryos increased compared to Comparative Example 1.

Comparative Example 1 Asparagus somatic embryos were cultured in the same manner as in Example 1, except that 0.1M shake rose was added.

(Left space on wooden page) [Effect of the invention] By using the asparagus tissue culture method of the present invention, normal development of asparagus somatic embryos into banana-shaped embryos without callus formation or malformation can be significantly improved. This promotes the propagation of seeds and seedlings in large quantities through this process.

Therefore, according to the method of the present invention, high-quality plants can be cultured in large quantities from asparagus tissues more efficiently than in conventional methods, and seeds and seedlings can be propagated in large quantities.

Applicant: Mitsui Petrochemical Industries, Ltd. Agent: Patent attorney Yusuke Hiraki

Claims (2)

[Claims] (1) Asparagus tissue culture, characterized in that a spherical embryo obtained by culturing asparagus tissues or cells is cultured using a medium containing 0.2 M or more of sucrose to obtain a banana-shaped embryo. Method. (2) A banana-shaped embryo is obtained by culturing a spherical embryo obtained by culturing asparagus tissues or cells using a medium containing 0.1 M or more of sucrose and 0.05 M or more of glucose. Asparagus tissue culture method.