JPH03187328A - Raising seedling of plant of genus dicentra - Google Patents

Abstract

PURPOSE:To produce young seedlings of a plant of genus Dicentra in large quantities in a short time at all seasons throughout the year the inducing a callus of a plant of genus Dicentra in a medium containing auxins, inducing indefinite bud from the callus and rooting the indefinite bud. CONSTITUTION:A part of the tissue (e.g. leaf) of a plant of genus Dicentra is cultured in a callus-inducing medium containing auxins as phytohormones and the obtained callus is proliferated by repeating the division and transplantation on a callus-proliferation medium. The proliferated callus is transferred to a medium for inducing indefinite bud. The induced indefinite bud is transferred to a rooting medium and rooted. Finally, the rooted young seedling is grown to reproduced the plant body.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention relates to a method for raising seedlings of plants of the genus Dicentra, and in particular, a method of raising seedlings of plants of the genus Dicentra, which is suitable for obtaining a large number of seedlings in a short period of time by organ culture. It is related to.

[Prior Art] Dicentra is a typical alpine plant of the poppy family, a perennial plant, and blooms pale pink or white flowers around July or August. In other words, Dicentra humus is popular as an ornamental plant;
Moreover, the alkaloids decentrin (C2゜H2□04N) and protopine (C2, H, ○S N) are present in the whole plant.
) is a useful plant that contains ingredients with anesthetic properties.

Methods of propagating Dicentra humus include propagation by seedlings and multiplication by division. However, it is difficult to germinate seedlings, and both of these methods of dividing seedlings into two
Its proliferation rate was extremely low.

[Problems to be Solved by the Invention] As mentioned above, the multiplication rate of Dicentra humus is extremely low in both methods of dividing seedlings into two, and it takes more than two years to grow into a flowering plant. Mass production of Dicentra humus is extremely difficult.

Furthermore, conventional methods are affected by the seasons and can only produce seedlings in the spring.

Currently, large quantities of young seedlings are obtained by tissue culture of plants in Western culture, etc., but it is completely unknown to obtain young seedlings from Dicentra humus by such a method.

The present invention was made in order to solve the problems of the above-mentioned conventional technology.
Moreover, it is an object of the present invention to provide a method for raising Dicentra humus seedlings that can be produced throughout the year without being affected by the seasons.

[Means for Solving the Problem] In order to achieve the above object, the structure of the method for raising Dicentra humic plant seedlings according to the present invention is such that a part of the organ of Dicentra humic plant is collected, and the collected material is A process of inducing callus by culturing in a callus induction medium containing at least auxins as a plant hormone, a process of multiplying the callus by repeating dividing and transplanting using a callus propagation medium, and inducing adventitious buds from the proliferated callus. a step of transferring the adventitious buds to a rooting medium to induce adventitious buds, and a step of transferring the adventitious buds to a rooting medium to root and produce seedlings, and grow the rooted seedlings to regenerate the plant body. It is something.

The drawings are process explanatory diagrams of a method for raising seedlings of Dicentra humicus.

As shown in the drawings, the present invention relates to organs (leaves) of dicentra
Collecting the callus (Step 2), inducing callus with a callus induction medium (Step 2), and repeating division and transplantation of the callus to proliferate in large quantities (Step 2). Transfer the proliferated callus to another adventitious bud induction medium to induce adventitious buds (Step 2), transfer the adventitious buds to a rooting medium to root (Step 2), and grow the seedlings to grow Dicentra plants. Regenerating the body (Step ①) This is a method consisting of each step.

Note that the plant organs here include roots, stems, leaves, and flowers.

In fruits, etc., a collection of cells with similar shapes and functions is generally called a tissue, and several tissues come together to form an organ.

In addition, callus in botanical terms refers to an amorphous plant cell mass formed experimentally by cell division.

Additionally, the culture medium is formed into test tubes or culture vessels.

Next, the method of the present invention will be explained in more detail.

First, a part of an organ such as a leaf, stem, or root of a Dicentra humicus plant, such as a leaf piece, is cut off, and this is cultured in a callus induction medium to induce callus.

As the callus induction medium, Gamborg's B5 medium, Murashige-Skoog's medium, White's medium, Linsmeyer-Skoog's medium, and modified media of these, which are commonly used for plant tissue culture, are used, and auxins are added to these medium. , cytokinin-type plant growth hormones are added.

In addition, Gunbolder B5 in the above and the following explanations
Regarding the culture medium, Gamborg, Miller Ojima (0, L,
Gamborg, R, A, Mil 1 e r, to,
Exp, Ce11 Res, 5 by Oj ima)
0. Pages 151-158, published in 1968.

Here, at least auxin is always added as a plant growth hormone, and examples of auxin include, for example, 2,4-dichlorophenoxyacetic acid (2,4-
D), indole acetic acid (Technical AA), naphthalene acetic acid (N
AA), indolebutyric acid (IBA), etc. Examples of the cytokinins include kinetin, benzyladenine, and zeatin.

Auxins usually have a concentration of 0.01 to 10 pPm.
Be sure to add it to the culture medium at the following ratio. Further, cytokinins are added to the medium at a concentration of 10 pPm or less, but do not necessarily need to be added.

On the other hand, if the amount (concentration) added is too large exceeding IQppm, death, growth inhibition, etc. will be observed.

Furthermore, when the amount of auxins is less than O, Olppm, organs such as adventitious roots other than callus may directly differentiate, die, or be inhibited in growth.

Dicentra leaf pieces were placed in a callus induction medium at an ambient temperature of 15 to
When cultured at 30°C in the dark, callus is formed on the cut surface of the leaf after 15 to 30 days.

Next, the cultured callus is transplanted to a callus growth medium and the callus is grown.

As the callus growth medium, a medium commonly used for plant tissue culture such as Gamborg's B5 medium is used, and a solid medium such as an agar medium containing agar or a liquid medium containing water is used. Alternatively, the callus induction medium described above may be used.

As plant hormones that promote proliferation, auxins and cytokinins may be added at a concentration of 10 ppm or less, or may not be added.

If these amounts are too large (more than 10 ppm), death, growth inhibition, etc. will be observed.

Cultivation is usually carried out in the dark at an ambient temperature of 18-30°C. The proliferated callus is further proliferated by repeating division and transplantation as shown in steps ① and ② of the drawing.

Next, the proliferated callus is transferred to an adventitious bud induction medium to induce adventitious buds.

As the adventitious bud induction medium, a Murashige-Skoog medium commonly used for plant tissue culture is used. Murashige・
The composition of Skoog's medium is T.

physi by Murashige, F., Skoog
ol Plantarum, 15e pages 473-4
97. Described in 1962.

The adventitious bud induction medium is the Murashige-Skoog medium, etc., containing cytokinins such as benzyladenine and kinetin at a concentration of 5p as plant hormones for promoting adventitious bud induction.
Add at pm or less, or do not add at all. Auxins should not be added, or if added,
The concentration should be 2 ppm or less, which is lower than the @ ground for callus propagation. Here, the addition of cytokinins produces better results in inducing adventitious buds, while the addition of auxins produces better results. In addition, if the concentration of cytokinins exceeds 5 Ppm, they turn brown and die, and auxins have a concentration of 2 Ppm.
If the amount exceeds ppm, differentiation of adventitious buds will not occur and only callus will proliferate.

If callus is placed on such an adventitious bud induction medium and cultured at an ambient temperature of 15 to 30°C in the dark or under irradiation of 20°,000 lux or less, adventitious buds will be induced in 15 to 40 days. be done.

Next, the adventitious shoots thus induced are transferred to a rooting medium and allowed to root.

Gamborg's B5 medium, which is commonly used for plant tissue culture, is used as a rooting medium, and indolebutyric acid (IBA) and indoleacetic acid (I) are used as plant hormones to promote rooting.
Auxins such as AA) are added at a concentration of 5 ppm or less, or are not added at all. Further, cytokinins are usually not added, or if added, the amount is 0.5 ppm or less.

Here, if a large amount of auxins is added in a concentration exceeding 5 ppm, adventitious shoots will wither, and if a large amount of cytokinins is added in a concentration exceeding 0.5 ppm, rooting will not occur.

Adventitious shoots are planted in such a rooting medium, and the ambient temperature is 15-3.
When cultured under irradiation with light of 1,000 to 15,000 lux at a temperature of 0°C, roots will develop in 20 to 50 days and young Dicentra seedlings will be obtained, and by growing them, the plant will be regenerated. Ru.

[Function] The function of the above technical means (each step of the seedling raising method) is as follows.

In the conventional method of dividing one seedling, the proliferation rate of plants of the genus Dicentra is extremely low, but in the method of the present invention, callus is effectively induced in a medium containing plant hormones. Proliferation occurs rapidly in this form of callus. Therefore,
By inducing adventitious buds from this multiplied callus and rooting them, a large number of young seedlings of plants of the genus Dicentra can be obtained in a short period of time.

In addition, in conventional methods, seedlings can only be obtained in spring due to the influence of the season, but with the method of the present invention, seedlings can be obtained throughout the year without being affected by the season.

[Example] Examples of the present invention will be described below.

Lost salmon (1) Collection of Dicentra leaf pieces (refer to the process in the drawing) Cut Dicentra seedlings to an appropriate size, immerse them in a 70% ethanol aqueous solution for 60 seconds, and then soak them in a sodium hypochlorite aqueous solution (C1 - Concentration: 2°0%) for 12 minutes for sterilization.

This was washed three times with sterile water and then cut into three leaves.

(2) Induction of callus (see step ① in the drawing) The above-mentioned leaf piece with three leaves attached was placed on a callus induction medium.

As a callus induction medium, 2.4 auxins are recommended.
A Gamborg B5 solid medium (Gelrite 0.2%) containing D in a Q ratio of 2 ppm and benzyladenine as a cytokinin in an O lppm ratio was used.

The callus induction medium on which the leaf pieces were placed was heated to an ambient temperature of 24°C.
When cultured in the dark at ℃ for 20 days, callus was formed on the cut end of the leaf.

(3) Proliferation of callus (see step ① in the drawing) The callus formed in (2) was transplanted into a callus growth medium.

As a callus growth medium, 2.4 auxins are recommended.
A Gamborg B5 solid medium (Gelrite 0.2%) containing D in a Q ratio of 2 ppm and benzyladenine as a cytokinin in a Q ratio of 2 ppm was used.

The callus growth medium in which the callus was transplanted was heated to an ambient temperature of 2.
Culture was carried out in the dark at 4°C for 30 days.

The callus was propagated by repeating dividing and transplanting the callus as shown in steps ① and ② of the drawing.

(4) Induction of adventitious buds (see step ① in the drawing) The proliferated callus obtained in (3) was transplanted to an adventitious bud induction medium.

As a medium for inducing adventitious buds, Murashige-Skoog's solid medium (
Gelrite (0.2%) containing 0.1% of casamino acid and 25 ppm of adenine sulfate was used.

The adventitious bud induction medium to which the proliferated callus had been transplanted was cultured at an ambient temperature of 24° C. and in the dark for 12 days, and then under irradiation of 3000 lux. After Wl floor, about 2
Adventitious bud induction started in 0 days.

More than 20 adventitious buds were induced in a test tube with a diameter of 25 m5 in about 30 days.

(5) Rooting (see step ■ in the drawing) The adventitious shoots induced in (4) were cut from the base and transplanted onto a rooting medium.

As a rooting medium, a solid medium of Gamborg B5 (Gelrite 0.2
%) was used.

The rooting medium to which the adventitious shoots were transplanted was grown at an ambient temperature of 20°C for 3
Culture was performed under light conditions of 000 lux. Approximately 2 after placing the bed
Rooting began in 0 days, and several roots of 20 to 40 m in length were developed in 40 days, yielding young Dicentra seedlings that could be transplanted into pots.

As described above, according to this example, seedlings can be obtained in a shorter period of time than in the conventional method of germinating seedlings or dividing the seedlings.

lost sight triumphant master (1) Collection of Dicentra leaf pieces This is the same as in Example 1 (1).

(2) Induction of callus This is the same as in Example 1 (2).

(3) The callus formed in callus multiplication (2) is treated as auxin.
The cells were transplanted into a Gamborg B5 liquid medium containing 4D Q at a rate of 2 ppm and no cytokinins, and cultured in a reciprocating shaking incubator.

By replanting the callus in the same medium every two weeks,
The callus becomes suspended in the liquid medium.

The growth rate was higher than that on solid medium. Other culture conditions are the same as in Example 1 (3).

(4) Induction of adventitious buds The suspended callus obtained in (3) above was used in Example 1 (4).
), and cultured under the same temperature and light conditions as in Example 1 (4).

Adventitious buds began to be induced about 20 days after transplantation, and many adventitious buds of approximately uniform size were induced about 30 days later.

(5) Rooting When the adventitious shoots obtained in (4) above were cultured in the same manner as in Example 1 (5), young Dicentra seedlings that could be transplanted into pots were obtained in about 40 days.

[Effects of the Invention] As explained in detail above, the present invention provides a method for raising Dicentus plants that can produce large quantities of Dicenta plant seedlings in a short period of time and throughout the year without being affected by the seasons. I was able to provide it.

[Brief explanation of drawings]

The drawings are process explanatory diagrams of a method for raising seedlings of plants of the genus Dicentra.

Claims (1)

[Scope of Claims] 1. A step of collecting a part of an organ of a plant of the genus Dicentra and culturing the collected material in a callus-inducing medium containing at least auxins as a plant hormone to induce callus; A step of multiplying the callus by repeating dividing and transplanting using a callus propagation medium, a step of transferring the proliferated callus to an adventitious bud induction medium to induce adventitious buds, and a step of transferring the adventitious buds to a rooting medium. A method for raising seedlings of plants of the genus Dicentra, which comprises a step of transferring and rooting the seedlings, and nurturing the rooted seedlings to regenerate the plant body. 2. The callus induction medium has an auxin concentration of 0.0.
1 to 10 ppm, and the concentration of cytokinins is 1
The method for raising seedlings of plants of the genus Dicentra according to claim 1, characterized in that the concentration is 0 ppm or less. 3. The method for raising seedlings of plants of the genus Dicentra according to claim 1, wherein the callus propagation medium has a concentration of each of auxins and cytokinins of 10 ppm or less. 4. The adventitious bud induction medium has an auxin concentration of 2pp.
2. The method for raising seedlings of plants of the genus Dicentra according to claim 1, wherein the concentration of cytokinins is 5 ppm or less. 5. The method for raising seedlings of plants of the genus Dicentra according to claim 1, wherein the rooting medium has a concentration of auxins of 5 ppm or less and a concentration of cytokinins of 0.5 ppm or less.