WO2011033658A1 - METHOD FOR CULTIVATING PEA FAMILY PLANT UTILIZING γ-RAY - Google Patents

Abstract

Disclosed is a method for cultivating a pea family plant, which enables the harvest of larger seeds than those harvested by conventional cultivation methods. The method comprises irradiating the plant that is in a post-seedling stage with a γ-ray at a dose rate of 0.04 to 0.4 Gy/day and the total dose of 0.5 to 10.0 Gy.

Description

Leguminous plant cultivation method using gamma rays

The present invention relates to a method for cultivating legumes using γ-rays, and in particular, cultivating leguminous plants including a step of irradiating a plant body with γ-rays at a dose rate in a specific range after the seedling has passed. Regarding the method.

】 Legumes such as soybeans are one of the main grains of mankind and are produced about 200 million tons worldwide. Leguminous plants such as soybean are widely used not only for food but also as vegetable oil and livestock feed, and in part, they are also being used as bioenergy, and their production is increasing year by year. One countermeasure against such an increase in demand is to produce soybean with high productivity by genetic engineering techniques. However, plants created by such genetic engineering techniques have become a global problem because they may adversely affect the conservation and sustainable use of biological diversity. There is an increasing demand for techniques that can increase productivity regardless of the above.

By the way, after the radiation hormesis effect was advocated by Thomas D. Lucky of the University of Missouri in 1978, various plants such as corn, alfalfa, soybean, pea, potato, tulip, lily, barley, green beans, etc. There have been reports on the hormesis effect of radiation such as γ rays (Non-patent Document 1). However, these reports can be understood as suggesting that effects such as the promotion of growth can be obtained by irradiating a prescribed amount of radiation in the state of seeds, tuberous roots, and seedlings. The dose rate and the irradiation period are not mentioned, and the effects obtained by the irradiation are various. For this reason, there is much debate about the validity of the data. Recently, there is a report that growth and wrinkle are reduced from about 0.4 Gy by rapid irradiation of young seedlings after germination, but there is no change in the size of wrinkles (Non-Patent Document 2). ). In addition, there is a report that, in the contaminated area around Chernobyl, the beans grown in the presence of radionuclides in the soil are small in harvested beans (Non-patent Document 3). References cited in the present specification are incorporated herein by reference.

Because of the actual state of research on the use of radiation for such crop growth, in Japan it is only used for potato bud growth control and food sterilization treatment, and radiation is used to grow plants used for food. It has not been. The same is true in other countries where the regulations on the use of radiation are relatively loose, and even in the world, radiation is not used to grow plants used for food at a practical level.

However, increasing the production of plants used for food, etc. by using radiation may solve the problems caused by plants created by genetic engineering techniques, and if clear conditions that lead to increased yields are established, the production will increase. Can provide new options for.

The present invention is based on the discovery of conditions for increasing the production of legumes that could not be predicted at all from the technical knowledge at the time of filing of the present application. In other words, not only the total dose but also the irradiation time and dose rate were examined in detail. After the seedling time passed, the total dose was 0.5-10.0 Gy and 0.04-0.4 Gy / day. When plants were irradiated with γ-rays at a dose rate in the range, the seeds were found to be larger and the average weight of the pods increased than in the case of non-irradiation, and doses in the narrower range of 0.05 to 0.08 Gy / day were found. When the plant body was irradiated with γ rays at a rate, the number of pods per individual also increased, and the weight of cocoons harvested from one individual was found to increase significantly, leading to the completion of the present invention. .

Thus, in one embodiment of the present invention, after the seedling period has elapsed, the plant body is given a total dose of 0.5-10.0 Gy, a dose rate of 0.04-0.4 Gy / day, Provided is a leguminous plant cultivation method, a legume plant grown by this method, and a seed obtained therefrom, characterized by irradiating with γ rays preferably at a dose rate of 0.05 to 0.08 Gy / day Is.

As used herein, “seedling” refers to a plant that has germinated from the seed of a seed plant and has a cotyledon remaining, and “after the seedling has passed” means that the plantlet has lost its cotyledon and is normally It refers to the season with leaves. In the present specification, “flower bud” means a growth point that has undergone vegetative growth differentiated into a growth point that undergoes reproductive growth, that is, a flower or inflorescence primordium. Means that the plant has grown and is close to flowering.

It should be noted that the presence or absence of flower bud formation can be determined by taking an autoradiography of the growth point or by observing changes in cell division images. However, since most of the cultivated plants are already known about the flower bud formation time, that is, the reproductive differentiation start time (for example, Non-Patent Document 6 states that soybean flower bud differentiation is performed around 20 days before flowering). At the practical level, you can follow it.

In addition, when viewed with individual flower buds, buds, and flowers in an individual, the formation time and flowering time are different. When viewed at an individual level, the stage before flower bud formation, the stage of flower bud formation, It should be noted that two or more stages of formation and flowering can coexist. Accordingly, when the terms “before flowering”, “before bud formation”, and “flower bud formation” are used herein, unless stated otherwise, if at least one flower is before flowering, “before flowering” "If at least one bud is formed, then" before bud is formed ", and if at least one flower bud is formed, it corresponds to" flower bud formation ".

In the present specification, “the cocoon is enriched” means a state in which the cocoon is sufficiently swollen by the seeds therein. Further, in the present specification, “until the eyelids are enriched” means until at least one eyelid is enriched, but in terms of further ensuring the effect of the method of the present invention, the irradiation of γ rays is almost (80 % Or more, preferably 90% or more) is preferably carried out until the soot is enriched.
Further, in the present specification, “seed” includes not only fully-ripened seeds but also immature seeds. Such immature seeds are edible in Japan and the like.

In addition, in this specification, the dose rate and total dose are values calculated from the absorbed air dose (Gy) measured for the ability to ionize air directly or indirectly by gamma rays emitted from a radiation source (radioactive material). means. The dose of gamma rays was defined by the source nuclide, radioactivity, distance from the source, etc., and was measured with a thermoluminescence dosimeter (TLD). More specifically, the dose rate is measured with a thermoluminescence dosimeter (TLD) at 10 m intervals from the radiation source ( ⁶⁰ Co) in the γ field at 1 m above the ground, and then a regression equation (log ₁₀ Draw Y (Gy / h) = A + B × log ₁₀ X (m)), calculate A and B, and calculate the dose rate (Gy / h) according to the distance, and the daily dose rate (Gy in this example) / 8h is expressed as a daily dose rate (Gy / day)). Attenuation from the measurement date was corrected by (Y) × (1/2) ^{t / T} , and the distance was determined. (Y is the dose rate on the day of examination, X is the distance from the source, T is the half-life of the source, and t is the elapsed time from the day of examination).

According to the cultivation method of the present invention, larger seeds can be harvested than in the normal cultivation method. In addition, according to a particularly preferred embodiment of the present invention, in addition to being able to harvest large seeds, the number of pods per individual can be increased, and the yield of seeds per individual can be increased. .
In the method according to the present invention, it is possible to achieve increased production of cultivated plants without causing a problem of food safety by genetic engineering techniques and a problem of ensuring plant diversity. In addition, in the conventional breed improvement technology, a great deal of labor and a long time are required for breed improvement, but the present invention eliminates such a barrier and enables an increase in production by a simple method.

FIG. 1 is a graph showing the average value of the weight per cocoon for each group of tests conducted in 2008. An increase in soot weight was observed in the 0.06 Gy / day dose rate group and the 0.2 Gy / day dose rate group (N = 4-9, where N indicates the number of samples, control group, 0.02 Gy / day This indicates that the number of samples in the day dose rate group, 0.06 Gy / day dose rate group, and 0.2 Gy / day dose rate group is between 4 and 9. The same applies hereinafter. FIG. 2 is a graph showing the number of pupae per individual in each group of tests conducted in 2008. The number of spikes per individual increased in the group with a dose rate of 0.06 Gy / day, whereas the number of spikes in the 0.02 Gy / day dose rate group and the 0.2 Gy / day dose rate group There was no tendency to increase (N = 4-9). FIG. 3 is a graph showing the average value of the weight of the total weight per individual in each group in the test conducted in 2008. It was the largest value in the 0.06 Gy / day dose rate group with a large weight per cage and a large number of cages (N = 4-9). A control group cultivated in Chiba at the same time is also shown (N = 3). FIG. 4 is a graph showing the average value of the weight per cocoon for each group of tests conducted in 2009. As in 2008, an increase in dredging weight was observed in the 0.06 Gy / day dose rate group and the 0.2 Gy / day dose rate group. FIG. 5 is a graph showing the number of pupae per individual in each group of tests conducted in 2009. As in 2008, an increase in the number of pupae per individual was observed in the 0.06 Gy / day dose rate group. FIG. 6 is a graph showing the average weight per cage for each group of tests conducted in both 2008 and 2009. Increased soot weight was observed in the 0.06 Gy / day dose rate group and the 0.2 Gy / day dose rate group (N = 11-17) FIG. 7 is a graph showing the number of pupae per individual in each group of tests conducted in both 2008 and 2009. The number of pupae per individual increased in the 0.06 Gy / day dose rate group, whereas the number of heels increased in the 0.02 Gy / day dose rate group and the 0.2 Gy / day dose rate group. Not recognized (N = 11-17). FIG. 8 is a photograph showing seeds harvested from each group of tests conducted in 2008. Large seeds were harvested in the 0.06 Gy / day dose rate group and the 0.2 Gy / day dose rate group (N = 4-9) FIG. 9 is a copy of a photograph showing the migration pattern of two-dimensional electrophoresis in which the protein expression pattern in seeds harvested in 2008 was analyzed. FIG. 9A shows the migration pattern of seeds harvested from the control group, and FIG. 9B shows the migration pattern of seeds harvested from the 0.2 Gy / day dose rate group.

Embodiments of the present invention will be described below, but the present invention includes other embodiments as long as it does not contradict the essence.

In the present invention, the plant body is irradiated with γ rays after the seedling time has elapsed.
Conventionally, many attempts have been made to irradiate various radiations at the seed, tuberous root, and seedling stages in various plants including legumes (Patent Document 1). However, γ-rays after the seedling stage in which differentiation has progressed have been made. There has been no report that irradiation of the plant caused an increase in legume seed production. Accordingly, the feature of the present invention that the plant body is irradiated with γ-rays at the stage after seedling is inventive with respect to the technical level at the time of filing.

As will be described later in Examples, in this test, the production of legume seeds was increased by starting irradiation with gamma rays in the vicinity of flower bud formation, specifically, when several pods were confirmed. It seems that the production of seeds can be increased if irradiation of γ rays is started at least from the time when flower buds are formed, and most seedlings are γ from the time immediately before or after flower bud formation. More preferably, the irradiation of the line is started. However, according to this test, even if the wrinkles were confirmed to some extent, it was possible to increase production, so even if γ-ray irradiation was started before or after the wrinkle formation (within a few days), seed production increased Will be possible. In addition, it is known that the size of seeds is affected by sunlight, water and nutrient conditions after the flowering period (Non-patent Documents 4 and 5), and γ-irradiation after the flowering period is related to seed development. It is expected that it will have some effect on the expression of genes. Therefore, from the viewpoint of increasing the seed size, irradiation with γ rays may be started after flower formation and before flowering.

In any case, in order to increase the yield of seeds, it is preferable to start irradiation with γ rays around several days after flower bud formation, and it is preferable to start irradiation with γ rays before wrinkles are formed at the latest. .

Similarly, in terms of increasing the number of wrinkles, radiation is considered to work in any of the period of flower bud differentiation, the period of wrinkle formation, or the period of wrinkle formation. Preferably, until a large number of wrinkles are formed (eg, 50% or more of the total number of wrinkles expected per individual), more preferably until wrinkles are formed. . In addition, from the viewpoint of forming large seeds, it is considered that seeds will be further enlarged by γ rays during the period until the pods are enriched, and irradiation of γ rays should be carried out at least until the seeds in the pods begin to expand. It is more preferable to carry out until several to 10 ridges start to swell, more preferably until at least one cocoon is filled, until 80% or more of all cocoons are filled. It will be particularly preferred to do so.

In the present invention, attention is paid to the dose rate. In one embodiment of the present invention, a dose rate of 0.04 to 0.4 Gy / day, preferably 0.04 to 0.4. Gamma rays are irradiated at 3 Gy / day, more preferably at a dose rate of 0.05 to 0.25 Gy / day. Previously, test results on the hormesis effect were reported by irradiating the seeds of various plants including legumes with gamma rays, but none of the detailed conditions other than the dose were studied, and the effects There were also various. Therefore, γ-ray irradiation at a low dose rate in a specific range proposed by the present application presents for the first time a clear condition for increasing the legume seed size (1.3 times or more by dry weight). In this respect, the present invention has inventiveness with respect to the technical level at the time of filing.

In addition to this, the present invention, in a particularly preferred embodiment, presents conditions for increasing the number of pods to be formed and increasing the yield of blisters per individual. That is, by irradiating the plant body with γ rays at a dose rate in a narrower range of 0.05 to 0.08 Gy / day, preferably 0.05 to 0.07 Gy / day, for the predetermined period, The number of pods formed can also be increased and the yield per individual can be increased. In such an extremely narrow range, the phenomenon that seeds grow and the number of pods formed increases, which has never been known so far, and it is the first time that there is a clear condition that makes it possible to increase the production of legume seeds. Present.

Irradiation with γ-rays may be performed continuously or intermittently, but it is preferable to perform γ-ray irradiation as continuously as possible. Moreover, when irradiating at intervals, the irradiation can be performed several days apart, but it is desirable that the irradiation be performed for two days or less.
The total irradiation date is preferably 10 days or more, more preferably 15 days or more, still more preferably 20 days or more, and particularly preferably 25 days or more.

The total dose of γ rays depends on the total irradiation day, but is preferably about 0.5 to 10.0 Gy, more preferably about 0.8 to 6.0 Gy. It is particularly preferably about 0.0 to 2.0 Gy. When the dose rate is 0.04 to 0.4 Gy / day or 0.04 to 0.3 Gy / day, the total dose is preferably 0.5 to 10.0 Gy, and the dose rate is In the case of 0.05 to 0.25 Gy / day, the total dose is preferably about 0.8 to 6.0 Gy, more preferably about 1.0 to 5.0 Gy, and the dose rate is When it is 0.05 to 0.08 Gy / day, the total dose is preferably about 1.0 to 2.0 Gy, and when the dose rate is 0.06 to 0.08 Gy / day, The total dose is preferably about 1.3 to 2.0 Gy.

The total dose of 10 Gy or less is used for potato germination prevention and sterilization treatment (germination suppression is about 0.05-0.15 kGy, sterilization of spoilage pathogens is 1-7 kGy, seasoning or food The sterilization of the material is 10-50 kGy), which is a very low dose range, and the present invention seems to have a low barrier to its practical use.

In general, the irradiation time per day is preferably determined according to the dose rate to be used, from the viewpoint of the effectiveness of irradiation and avoiding a high level of irradiation per unit time. For example, in the case of irradiation at a dose rate of 0.05 to 0.08 Gy / day, it is considered sufficient to ensure an irradiation time of at least 2 hours, and when irradiation is performed at a dose rate exceeding this, It is preferably 4 hours or longer, particularly preferably 7 hours or longer.

No particular limitation is imposed on the γ-ray radiation source may for example 60Co, 192Ir, 169Yb, 170Tm, 75Se, 124Sb, be mentioned 153Gd or ^{137 Cs.}

The method of the present invention is considered to be applicable to at least leguminous plants (see also Patent Document 1), and leguminous plants include, for example, plants such as soybean genus, genus pea, and pea genus. . Among them, a plant of the genus Soybean, which is considered to have a certain resistance to radiation, is preferable.

In Japan, there is an eating habit of using immature seeds such as soybeans, green beans, cowpeas, peas, peanuts, wisteria bean, jujube, winged bean, 16 cowpeas, etc. and young pods as they are. Being eaten generally. In addition, soybean varieties developed to eat as “edamame” include early varieties such as Sachio Okuhara, Swan, Sayamusume, Beer Friend, Shirako, etc .; Fuki, Ryokan, Mikawajima, Nishikiaki, Mid- and early-birth varieties such as evening coolness, hot spring girls, Fukushiko, etc .; late-generation varieties such as Akiyoshi and Toyoshirome; Black bean varieties; Dadacha beans, Kurosaki tea beans, Takihime, Fukunari etc.

In Japan, ripe seeds are used as processed foods such as tofu, miso and natto, or as raw materials for vegetable oils in Japan, and are widely used worldwide as raw materials for vegetable oils and livestock feed. Attempts have also been made to use it as bioenergy.

The present invention is considered to contribute to the utilization of such leguminous plants, and is considered to be effective when applied to soybean family plants, particularly early soybean species (note that genes related to early / late nature are not considered). (See Patent Documents 7 and 8).

The leguminous plant obtained by the method of the present invention is different from the same non-irradiated plant in terms of phenotypes such as an increase in seeds and an increase in the number of pods, as demonstrated in Examples described later. In addition, there is a difference in protein expression with respect to the same non-irradiated plant. However, such differences are not due to differences in the genes themselves, but are thought to be because the expression of the genes was affected by gamma rays.

Therefore, in one embodiment, the present invention is also directed to legumes having an unprecedented character cultivated by the method described in detail above, and seeds obtained from such legumes. Such legumes and seeds provide a new means of efficiently providing food or energy resources without concern about the adverse effects on biodiversity conservation and sustainable use by transgenic plants. obtain.

Hereinafter, the present invention will be described in more detail with reference to examples. However, the present invention should not be limited by the examples.

1. Test in fiscal year 2008 Odawara early seedlings (Sakata seedlings and Edamame are considered to be native to China) were used for the tests. In order to avoid damage to birds by sowing green soybean seeds soaked in water overnight on May 14, 2008 (Day 1) in organic culture soil (Plantation Iwamoto) in No. 6 pot (pot) Cultivated indoors for 8 days. Seeds germinated around the 5th day, and on the 9th day, the seeds were 4 to 10 cm in size. On the 9th day, it was taken out outdoors (on the roof of a five-story building) and continued to grow. On the 42nd day (June 24, 2008), it was confirmed that several cocoons were formed from several seedlings. At this point, several pots (pots) with 1 to 3 seedlings per pot (pot) are assigned to the control and three irradiation groups, each with about 10 seedlings, and the length distribution in each group Sorted out to be roughly aligned. Gamma rays that irradiate γ-rays with doses of 0.02 Gy / day, 0.06 Gy / day, and 0.2 Gy / day from about 42 TBq of ⁶⁰ Co from the 42nd day with the three irradiation groups in the pot. Installed in the field (located in Hitachiomiya City, Ibaraki Prefecture). The dose rate is measured every 10 m from the radiation source ( ⁶⁰ Co) within the γ field, and measured with a thermoluminescence dosimeter (TLD) at 1 m above the ground, and then a regression equation (log ₁₀ Y (Gy / h)) = A + B x log ₁₀ X (m)) A and B are calculated, and the dose rate (Gy / h) corresponding to the distance is calculated, and the daily dose rate (Gy / 8h in this example is set to 1 day) Dose rate (expressed as Gy / day)). Attenuation from the measurement date was corrected by (Y) × (1/2) ^{t / T} , and the distance was determined. (Y is the dose rate on the examination day, X is the distance from the radiation source, T is the half-life of the radiation source, and t is the elapsed time from the examination date).

After installation, June 28, 2008, June 29, July 5, July 6, July 12, July 13, July 16, July 19, July 20, Except on July 21, July 26, and July 27, γ rays were irradiated for 8 hours a day. The plants in each group had some flowering in almost all plants around day 50 (irradiation date about 7 days in total), and on day 60 (irradiation date about 14 days in total). Most of the cocoons were formed in most plants, and most of them were enriched on the 25th day of irradiation (July 30, 2008). At this point, irradiation was stopped. The total dose was 0.5, 1.5 and 5 Gy for each irradiation group. From the 42nd day, the control group was placed outside the gamma ray field as a pot, and was cultivated there until July 31, 2008. Another control group was placed as a pot in Chiba, away from the gamma ray field, where it was grown until July 31, 2008.
Irrigation of each group was performed 2-3 times / week throughout the cultivation period.
On the day following the end of the irradiation period (July 31, 2008, 79 days after seed sowing), the straw was harvested from the plant. In addition, individuals that died significantly due to insect damage or the like were excluded.

2. Tests in 2009 The same test was conducted in 2009, the following year. Seeds were performed on May 14, 2009 (1st day), and γ-ray irradiation was started on the 43rd day (June 25, 2009), and then on June 27, June 28, 2009, July 4, July 5, July 11, July 12, July 15, July 18, July 19, July 20, July 25, July 26 Except for γ-ray irradiation for 8 hours a day. Plants in each group grew in the same manner as in 2008, and most of the wrinkles were enriched on the 25th day of irradiation (July 31, 2009). At this point, irradiation was stopped. Thereafter, on the third day (August 3, 2009, seeds were sown and the 82nd day), the straw was harvested. The control group was only a group in which pots were installed outside the field.

3. Test results 3-1. Number of pods and weight of seeds per pod The number of pods obtained and the weight of seeds per pod were measured for each group. The test results are as shown in FIGS.

FIG. 1 shows the average weight per cocoon of each group of tests conducted in 2008, FIG. 2 shows the number of cocoons per individual in each group of tests conducted in 2008, and FIG. The total weight of FIG. 4 shows the average weight per cocoon of each group in the test conducted in 2009, and FIG. 5 shows the number of cocoons per individual in each group of the test conducted in 2009. There was much individual damage due to bad weather etc. (N = 4-9)). FIG. 6 shows the average weight per cocoon of each group of tests conducted in both years, and FIG. 7 shows the number of cocoons per individual in each group of tests conducted in both years. FIG. 8 is a photograph showing beans harvested from each group of tests conducted in 2008.

As shown in Fig.1, Fig.4, Fig.6 and Fig.8, the average weight per cage is 0.06Gy / day dose rate (total dose 1.5Gy) group, and 0.2Gy / day dose rate (total dose 5Gy) There was an increase in the group. The average seed dry weight is the control group (n = 105), 0.02 Gy / day dose rate (total dose 1.5 Gy) group (n = 64), 0.06 Gy / day dose rate (total dose 1.5 Gy) group (n = 132), 0.2Gy / day dose rate (total dose 5Gy) group (n = 117), 0.11g, 0.108g, 0.146g, 0.155g per seed, respectively (harvested in 2008) The seed dry weight of the 0.06 Gy / day dose rate (total dose 1.5 Gy) and the 0.2 Gy / day dose rate (total dose 5 Gy) relative to the average dry weight of the control group seeds) The dry weight of the seeds of the group was 1.3 and 1.44 times the weight, respectively.

In addition, as shown in FIG. 2, FIG. 5 and FIG. 7, the number of wrinkles per individual increased significantly compared to the control group in the 0.06 Gy / day dose rate (total dose 1.5 Gy) group. The Gy / day dose rate (total dose 0.5 Gy) group and the 0.2 Gy / day dose rate (total dose 5 Gy) group were not significantly different from the control group. Moreover, as shown in FIG. 3, in the 0.06 Gy / day dose rate (total dose 1.5 Gy) group, the total weight of wrinkles per individual increased significantly compared to the control group.

3-2.2 Analysis of protein expression pattern by two-dimensional electrophoresis (A) Extraction Seeds obtained from a control group harvested in 2008, a group with a dose rate of 0.2 Gy / day (total dose of 5 Gy), Put in 3% (w / v) PVP (polyvinylpyrolidone) in buffer solution (0.05M Tris-HCl pH 7.5 1mM EDTA, 1mM PMSF, 1mM DTT), crush in liquid nitrogen, 10 ml of a buffer solution (0.05 M Tris-HCl pH 7.5, 1 mM EDTA, 1 mM PMSF, 1 mM DTT) was added and centrifuged (18800 g, 30 min, 4 ° C.).
The obtained pellet is placed in a buffer (100 mM Tris-HCl pH 8.5, 4% SDS, 2% mercaptoethanol, 20% glycerol, 2 mM PMSF) for protein collection of the granular fraction, and placed in boiling water for 3 minutes. It was. After cooling, the mixture was centrifuged (18800 g, 10 min, room temperature), the supernatant was removed, passed through a 0.22 μm PVDF membrane filter, and subjected to acetone extraction (in 20 volumes of acetone, −20 ° C., overnight).
The sample subjected to acetone extraction is centrifuged (18800g, 10min, 4 ° C), the resulting pellet is re-stirred with 80% acetone, the lysate is placed at -20 ° C for 90min, and further centrifuged (18800g, 10min, 4 ° C). The obtained pellet was dissolved in a lysis buffer (5M Urea, 2M thiourea, 2% CHAPS, 2% SB3-10, 1% DTT). Finally, the lysate was subjected to ultracentrifugation (100,000 g, + 20 ° C., 1 h), and the resulting supernatant was used for electrophoresis as a granular fraction sample.

(B) Two-dimensional electrophoresis 70 μg of protein is electrophoresed in Immobiline DryStrip (GE Healthcare) with pH 4-7 (13 cm) in the first dimension, and then the second dimension electrophoresis is performed with 12% SDS-PAGE. went. After completion of the electrophoresis, SYPRO Ruby staining (Invitrogen) was performed and imaged with a FLA-5100 imaging analyzer (Fuji Film).

(C) Results As shown in FIG. 9, the expression patterns of major proteins are similar between the control group and the irradiation group, but significant increases and decreases were observed in some proteins (marked in the figure). This indicates that the irradiated and cultivated legumes were harvested at the same time as the control group but were qualitatively changed.

As described above, according to the present invention, large bean can be harvested from existing legumes without using genetic engineering techniques, and the yield of seed per individual can be greatly increased. Leguminous plants have been studied for use as energy resources and as pharmaceutical production plants in addition to their use as food, and their use in a wide range of fields is conceivable. Also, in space, radiation always exists, and it seems possible to apply the present invention by controlling it well.

Claims (14)

1. A method for cultivating leguminous plants comprising the step of irradiating a plant body with γ-rays at a dose rate of 0.04 to 0.4 Gy / day and a total dose of 0.5 to 10.0 Gy after the seedling has passed.
2. The method according to claim 1, wherein the gamma rays are irradiated at a dose rate of 0.04 to 0.3 Gy / day.
3. The method according to claim 1, wherein the gamma rays are irradiated at a dose rate of 0.05 to 0.25 Gy / day.
4. The method according to claim 1, wherein the gamma rays are irradiated at a dose rate of 0.05 to 0.08 Gy / day.
5. The method according to any one of claims 1 to 4, wherein a total dose of the γ rays is about 0.8 to 6.0 Gy.
6. The method according to any one of claims 1 to 4, wherein a total dose of the γ rays is about 1.0 to 2.0 Gy.
7. The method according to any one of claims 1 to 6, wherein the leguminous plant is a plant belonging to the genus Soybean.
8. The method according to any one of claims 1 to 7, wherein the irradiation of the gamma rays is started before flowering.
9. The method according to any one of claims 1 to 7, wherein the irradiation of the γ-ray is started before a wrinkle is formed.
10. The method according to any one of claims 1 to 8, wherein the irradiation of the gamma rays is started within a few days after flower bud formation.
11. The method according to any one of claims 1 to 10, wherein the irradiation of the γ-ray is performed at least until wrinkles are formed.
12. The method according to any one of claims 1 to 10, wherein the γ-ray irradiation is performed until the soot is enriched.
13. Legumes cultivated by the method according to claims 1-12.
14. Seeds obtained from the leguminous plant according to claim 13.

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