JPH0928220A - Production of mutated plant body - Google Patents

Abstract

PROBLEM TO BE SOLVED: To induce a mutated plant body exhibiting various forms in high yield, with suppressing the damage to the plant body minimum by irradiating the ovary of a plant after its pollination with a heavy ion beam. SOLUTION: This method for producing a mutated plant body is constituted by irradiating the ovary of a plant such as a tobacoo, a white whitlow grass, a rice and an orchid after their pollination with a heavy ion beam such as a nitrogen ion beam, a carbon ion beam, a neon ion beam and an argon ion beam at preferably a radiation dose of 5-100 Gy in an irradiating range of 5-20mm diameter, and selecting the plants generating an abnormality from among the plants treated with the irradiation.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for minimizing plant damage and inducing a mutant plant exhibiting various morphologies at a high rate. More specifically, the present invention relates to a mutant plant irradiated with heavy ion beams. About the method of production.

[0002]

BACKGROUND OF THE INVENTION Irradiation of plants with gamma rays or X-rays or treatment of the plants with agents such as ethyl methanesulfonate (EMS) can artificially mutate plants [Euphytica 69: 95 to 101 (1
993)]. Since some of such mutants have excellent traits not found in conventional plants, plant breeding using mutations has been conventionally performed.

However, since it is difficult to accurately irradiate specific tissues of a plant with radiations such as gamma rays and X-rays, it is very difficult to fix gene mutations occurring in some cells as genetic traits of the whole individual. Requires effort.
In addition, drugs such as EMS induce mutations and damage the plants themselves and reduce the germination rate, so that it is difficult to obtain a large number of mutants.

Meanwhile, radiation called a heavy ion beam can be produced by ionizing and accelerating various elements such as carbon and nitrogen using a cycloton. As an example of producing a mutant of a plant by heavy ion beam irradiation, there is an example in which a rice seed is irradiated with a heavy ion beam to obtain a mutant strain resistant to white blight of rice (R
IKEN Accel. Prog. Rep. 26: 109, 1992).

However, in this method, the seeds are irradiated with a heavy ion beam, so that the seed embryo part which grows as a plant has already differentiated into many cells, and it is difficult to cause mutation only in the cells that carry out genetic control. It is. In addition, genes are fixed in a stable structure in dried seeds, and have a drawback such that when the state is changed by mutation, they often become lethal.

[0006]

As described above, artificially inducing mutation is an extremely useful technique for producing a trait-excellent plant. Mutants could not yet be induced with a satisfactory frequency. The present invention has been made in view of such problems of the prior art, and aims at minimizing plant damage and inducing mutants exhibiting various morphologies at a high rate. It is to provide a method that can be performed.

[0007]

The present inventors have made intensive studies to solve the above-mentioned problems, and as a result, by irradiating the plant ovary after pollination with a heavy ion beam, it is possible to use various varieties at a very high frequency. The present inventors have found that a mutant showing a morphology can be induced, and completed the present invention. That is, the present invention provides a method for producing a mutant plant, which comprises irradiating a plant ovary after pollination with a heavy ion beam and selecting a plant having a morphological abnormality from the irradiated plants. It is.

[0008] The present invention also provides a method for producing a chlorophyll-deficient plant, comprising irradiating a pollinated plant ovary with a heavy ion beam and selecting a chlorophyll-deficient plant from the irradiated plants. is there. Hereinafter, the present invention will be described in detail. The heavy ion beam used in the present invention is not particularly limited as long as it can induce a mutant. For example, a nitrogen ion beam, a carbon ion beam, a neon ion beam, an argon ion beam, or the like can be used. The dose of the heavy ion beam may be determined according to the type of ion beam used, the type of plant to be irradiated, and the like.
In addition, since the induction frequency of the mutant is low at 5 Gray or less, the range is preferably 5 to 100 Gray. If the irradiation range of the heavy ion beam is too wide, damage to the plant body becomes large. Therefore, it is preferable that the irradiation range is about 5 to 20 mm in diameter.

The plant to be subjected to heavy ion beam irradiation is not particularly limited, but preferred examples include tobacco, Arabidopsis, rice, and orchid. The irradiation time of the heavy ion beam is preferably to irradiate the egg cell immediately after fertilization, where mutation is likely to occur. More specifically, 2 to 108 hours after pollination (just after fertilization to 80 hours)
Is preferred.

After irradiating the plant ovary after pollination with a heavy ion beam, seeds of the plant are sown and germinated. At the germination stage, a mutant can be obtained by selecting an individual having a morphological or physiological abnormality. The morphological abnormality here means different from the normal plant in appearance, for example, chlorophyll deficiency, needle-like leaves, gravity insensitivity, dwarfism, early birth, etc., and physiological abnormalities include drug resistance, environmental stress resistance, etc. It is. Among the deficiencies of chlorophyll mentioned here, in addition to those that do not contain chlorophyll at all, those that lack so-called variegated chlorophyll and those that exhibit light green color because the amount of chlorophyll is less than that of normal plants are also included. Including.

[0011]

【Example】

[Example 1] Nitrogen ions (7+: 135 MeV / u) were accelerated by a RIKEN cyclotron and irradiated to the ovary of a tobacco variety BY-4 at a diameter of 10 mm (see FIG. 1). In the experiment, the irradiation time was set to 36 hours after pollination, 48 to 60 hours,
Time, 96-108 hours, divided into 5 doses
0, 50, 100, and 200 grays were performed in five stages. At this time, LET (linear energy transfer)
Was 28.5 KeV / mm. One month after irradiation with the nitrogen ion beam, M1 seeds were harvested, and the germination rate of the seeds was determined. The result is shown in FIG. As shown in FIG. 2, when the dose is 200 Gray, the germination rate is extremely low, and it is estimated that a large damage has occurred to the plant.

The germinated plants contained individuals showing morphological abnormalities. These appearances are shown in FIGS. FIG. 3 shows an individual irradiated with a nitrogen ion beam at a dose of 10 Grays 96 to 108 hours after pollination, in a state containing no chlorophyll (albino). FIG. 4 shows an individual irradiated with a nitrogen ion beam at a dose of 10 Grays 72 to 84 hours after pollination, showing a small amount of chlorophyll and a pale green color. FIG.
Shows an individual irradiated with a nitrogen ion beam at a dose of 10 Grays 72 to 84 hours after pollination, and had needle-like leaves. FIG. 6 shows that 200 to 96 to 108 after pollination.
This shows an individual irradiated with a nitrogen ion beam at a gray dose, and had needle-like leaves. FIG. 7 shows an individual irradiated with a nitrogen ion beam at a dose of 10 Grays 72 to 84 hours after pollination, and showed a normal form.

The plants that germinated as described above exhibited various morphological abnormalities. Of these morphological abnormalities, attention was paid only to the chlorophyll deficiency, and the proportion of chlorophyll-deficient individuals in all germinated individuals was determined. The result is shown in FIG. FIG.
As shown, 48 to 60 hours after pollination and 96 to 10
When irradiated 8 hours later, the proportion of chlorophyll-deficient individuals was highest when the dose was 100 Gray, but when irradiated 72 to 84 hours after pollination, the highest percentage of chlorophyll-deficient individuals was obtained when the dose was 10 Gray. Accounted for a large percentage of the total. Irradiation 32 hours after pollination, the occupancy of chlorophyll-deficient individuals was low at all doses. [Comparative Example 1] As test plants, two tobacco varieties (Xant
hi and BY-4) were used.

The ovary from which the style was removed was covered with absorbent cotton impregnated with 0.1% EMS, and allowed to stand for 1 day. At this time, the treated portion was covered with a microtube to prevent drying.
This EMS treatment is performed after pollination at 36, 48, 54, 6
The test was performed in five stages after 0 hour. One month after the treatment with the mutagen, M1 seeds were harvested and the germination rate was determined. The result is shown in FIG. As shown in FIG. 9, the germination rate was not reduced at any stage in Xanthi,
In BY-4, when the treatment was performed 36 hours and 48 hours after pollination, a decrease in the germination rate was observed.

The germinated individuals included individuals showing various morphological abnormalities as in Example 1, but no chlorophyll-deficient individuals were found. The percentage of morphologically abnormal individuals in all germinated individuals was determined. In addition, the morphological abnormalities were classified into A) one-leaf type, B) three-leaf type, C) and other three types, and the indicated ratios were determined. The results are shown in FIGS. As shown in FIG. 109, the most morphologically abnormal individuals were the case where the processing was performed 36 hours in BY-4 and 48 hours in Xanthi. The observed morphological abnormalities were the one-lobe type abnormalities in Xanthi, and B
In Y-4, many trilobular morphological abnormalities were observed.

[0016]

The present invention provides a novel method for producing a mutant plant. This method has the following features. (1) Since the ovary is irradiated with a heavy ion beam, a large amount of mutant seeds can be reliably produced. Therefore, new functions can be confirmed in plant solids. (2) Since the heavy ion beam is irradiated during the gene growth phase after fertilization, lethal mutation can be reduced. (3) The heavy ion beam is limited to the ovary and is irradiated immediately, so that lethal mutation can be reduced. (4) Albino mutant plants can be efficiently produced. (5) Since the primordial (egg) cells are irradiated with a heavy ion beam, it is easy to obtain a homozygote having a uniform genetic trait.

As described above, the present production method is capable of minimizing plant damage and inducing mutant plants exhibiting various morphologies at a high rate, and is a very useful technique in the field of plant breeding. It is.

[Brief description of the drawings]

FIG. 1 is a diagram showing a method of irradiating a plant ovary after pollination with a heavy ion beam.

FIG. 2 is a diagram showing a relationship between a dose of a nitrogen ion beam and a germination rate.

FIG. 3 is a photograph showing the morphology of a morphologically abnormal individual.

FIG. 4 is a photograph showing the morphology of a morphologically abnormal individual.

FIG. 5 is a photograph showing a morphology of a morphologically abnormal individual.

FIG. 6 is a photograph showing the morphology of a morphologically abnormal individual.

FIG. 7 is a photograph showing the form of a normal individual organism.

FIG. 8 is a diagram showing the relationship between the dose of a nitrogen ion beam and the frequency of occurrence of chlorophyll-deficient individuals.

FIG. 9 is a diagram showing the relationship between the EMS treatment time and the germination rate.

FIG. 10 is a diagram showing the relationship between the processing time of EMS and the frequency of occurrence of abnormal morphological individuals.

FIG. 11 is a diagram showing the proportion of single-lobe morphologically abnormal individuals and three-lobe morphologically abnormal individuals in all morphologically abnormal individuals.

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Masayuki Kase 2-1 Hirosawa, Wako-shi, Saitama Prefecture, RIKEN (72) Inventor Akira Goto 2-1 Hirosawa, Wako-shi, Saitama RIKEN, (72) Inventor Yasushi Yano 2-1 Hirosawa, Wako-shi, Saitama Pref. RIKEN

Claims (2)

[Claims] 1. A method for producing a mutant plant, comprising irradiating a pollinated plant ovary with a heavy ion beam and selecting a plant having a morphological abnormality from the irradiated plants. 2. A method for producing a chlorophyll-deficient plant, comprising irradiating a pollinated plant ovary with a heavy ion beam and selecting a chlorophyll-deficient plant from the irradiated plants.