CN110326503B - Identification method for elongation capability of hypocotyl in dry direct-seeded rice - Google Patents
Identification method for elongation capability of hypocotyl in dry direct-seeded rice Download PDFInfo
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- CN110326503B CN110326503B CN201910719624.1A CN201910719624A CN110326503B CN 110326503 B CN110326503 B CN 110326503B CN 201910719624 A CN201910719624 A CN 201910719624A CN 110326503 B CN110326503 B CN 110326503B
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- 235000007164 Oryza sativa Nutrition 0.000 title claims abstract description 39
- 235000009566 rice Nutrition 0.000 title claims abstract description 39
- 238000000034 method Methods 0.000 title claims abstract description 18
- 240000007594 Oryza sativa Species 0.000 title 1
- 239000002689 soil Substances 0.000 claims abstract description 54
- 235000015097 nutrients Nutrition 0.000 claims abstract description 41
- 241000209094 Oryza Species 0.000 claims abstract description 38
- 230000003203 everyday effect Effects 0.000 claims abstract description 4
- 238000005406 washing Methods 0.000 claims abstract description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 21
- 238000009331 sowing Methods 0.000 claims description 12
- 239000000463 material Substances 0.000 claims description 7
- 238000005070 sampling Methods 0.000 claims description 5
- 238000005507 spraying Methods 0.000 claims description 4
- 238000005303 weighing Methods 0.000 claims description 4
- 239000008399 tap water Substances 0.000 claims description 3
- 235000020679 tap water Nutrition 0.000 claims description 3
- 238000007873 sieving Methods 0.000 claims description 2
- 238000003756 stirring Methods 0.000 claims description 2
- 230000001502 supplementing effect Effects 0.000 claims description 2
- 238000012136 culture method Methods 0.000 abstract description 12
- 238000010899 nucleation Methods 0.000 abstract description 10
- 238000012360 testing method Methods 0.000 abstract description 7
- 239000004576 sand Substances 0.000 description 8
- 230000001276 controlling effect Effects 0.000 description 3
- 229920006395 saturated elastomer Polymers 0.000 description 3
- 230000008572 axis elongation Effects 0.000 description 2
- 230000012010 growth Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000003147 molecular marker Substances 0.000 description 2
- 230000029553 photosynthesis Effects 0.000 description 2
- 238000010672 photosynthesis Methods 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 1
- ATJFFYVFTNAWJD-UHFFFAOYSA-N Tin Chemical compound [Sn] ATJFFYVFTNAWJD-UHFFFAOYSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000000875 corresponding effect Effects 0.000 description 1
- 238000012364 cultivation method Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000012252 genetic analysis Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000007180 physiological regulation Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000007226 seed germination Effects 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G22/00—Cultivation of specific crops or plants not otherwise provided for
- A01G22/20—Cereals
- A01G22/22—Rice
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G31/00—Soilless cultivation, e.g. hydroponics
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- Life Sciences & Earth Sciences (AREA)
- Environmental Sciences (AREA)
- Botany (AREA)
- Pretreatment Of Seeds And Plants (AREA)
- Cultivation Of Plants (AREA)
Abstract
The invention discloses a method for identifying mesocotyl elongation capability of dry direct-seeded rice, which adopts a nutrient soil culture method under the condition of high temperature (35 ℃, RH (50-60%) and low humidity to identify the mesocotyl elongation capability of the rice, observes the seedling emergence condition every day, records the seedling emergence sequence of different varieties, and samples and determines the mesocotyl length on the 3 rd day after the first variety emerges; taking out the seedlings from the plug tray in sequence, washing the seedlings clean, laying the seedlings on a flat plate with scales in sequence, and taking a picture; after all varieties are photographed in sequence, the mesocotyl length of each variety is measured by using Image J software according to a scale setting scale on a flat plate. By applying the method in the identification of the mesocotyl elongation capability of the dry direct seeding rice, the test period can be obviously shortened, the mesocotyl phenotype identification efficiency can be improved, and the test error caused by human factors can be reduced. Meanwhile, the repeatability and stability of the identification result of the elongation capability of the embryonic axis in the dry direct seeding rice are greatly improved.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a method for identifying the elongation capability of an embryonic axis in dry direct-seeded rice. A
Background
The dry direct seeding rice has a series of advantages of water saving, labor saving, suitability for mechanized cultivation and the like, but in the production of the dry direct seeding rice, because the root cutting depth is insufficient, the lodging phenomenon of the rice at the later growth stage often occurs, and in order to solve the problem, the seeding depth is often required to be increased in the production. Deep sowing is beneficial to the binding of the roots of rice, the absorption capacity of the root systems to water and nutrients is improved, and the lodging resistance of the rice is improved, but the rice is difficult to emerge under the deep sowing condition, the rice emergence rate is reduced, and therefore the problems of insufficient basic seedlings in the rice field and weed prevention and control are caused. And under the condition of deep sowing, the seed germination duration is long, seedlings cannot perform photosynthesis in the period, and the seedling growth is easy to weaken and the young leaves are easy to yellow. After rice germinates in a dark environment, the mesocotyl can eject the new germ out of the soil by extending, photosynthesis is carried out, and the survival rate of rice seedlings is improved. Generally, the long mesocotyl variety has strong soil-bearing capacity and high emergence rate. Therefore, the physiological regulation mechanism of mesocotyl elongation is determined, the gene for regulating mesocotyl elongation is excavated, the corresponding molecular marker is developed, and the screening and creation of long mesocotyl germplasm have important significance for molecular marker assisted breeding (MAS) of the dry direct seeding rice. However, many QTLs for controlling mesocotyl axis elongation have been localized in different studies so far, but the experimental result has poor repeatability due to the significant interaction between mesocotyl axis elongation and environmental conditions.
Disclosure of Invention
In order to solve the problems, the invention provides a method for identifying the elongation capability of the mesocotyl in the dry direct seeding rice, which realizes the stability and repeatability of the identification of the mesocotyl phenotype, thereby providing a stable phenotype for the genetic research of the elongation capability of the mesocotyl in the dry direct seeding rice.
In order to achieve the purpose, the invention adopts the technical scheme that:
a method for identifying the elongation capability of an embryonic axis in dry direct-seeded rice comprises the following steps:
s1, smashing the nutrient soil, sieving the smashed nutrient soil with a 5-mesh sieve, and uniformly stirring the smashed nutrient soil and the sieved nutrient soil for later use;
s2, filling 500g of smashed nutrient soil into the tray, paving and compacting, placing the hole tray into the tray, filling a certain amount of smashed nutrient soil into each hole of the hole tray, and compacting to enable the height difference between the soil surface and the hole opening to be 6 cm;
s3, selecting full and uniform rice seeds with intact glumes, sowing the seeds in sequence, sowing 15 seeds in each hole, and uniformly distributing the 15 seeds at the bottom of each hole;
s4, uniformly covering the smashed nutrient soil after all the materials are sowed until the soil surface is flush with the opening of the hole tray, and weighing the tray, the hole tray and the total mass of the nutrient soil and the seeds in the hole tray as W1;
s5, thoroughly pouring the nutrient soil in the hole tray in a spraying mode, taking the hole tray out of the tray after water leaks out of the holes, pouring the nutrient soil in the tray to saturation with tap water, putting the hole tray back into the tray, controlling the total amount of watering to be 2000g, then, supplementing the height of the nutrient soil in the hole tray to be level to the hole opening with smashed nutrient soil, and weighing the total weight to be W2;
s6, placing the tray and the plug tray together in a dark culture room with the indoor temperature of 35 ℃ and the relative humidity of 50% -60%, spraying once every 24 hours, and controlling the amount of sprayed water to keep the total weight of the sprayed water at the W2 level;
s7, observing the seedling emergence condition every day, recording the seedling emergence sequence of different varieties, and sampling and determining the mesocotyl length on the 3 rd day after the first variety emerges; taking out the seedlings from the plug tray in sequence, washing the seedlings clean, sequentially and flatly placing the seedlings growing uniformly in each material on a flat plate with scales, and taking a picture, wherein the camera position is required to be directly above the flat plate during taking the picture, the height is determined by placing the whole flat plate in a viewing frame, and the camera position is fixed by using a tripod; after all varieties are photographed in sequence, the mesocotyl length of each variety is measured by ImageJ software according to a scale set scale on a flat plate.
By applying the method in the identification of the mesocotyl elongation capability of the dry direct seeding rice, the test period can be obviously shortened, the mesocotyl phenotype identification efficiency can be improved, and the test error caused by human factors can be reduced. Meanwhile, compared with other methods, the method greatly improves the repeatability and stability of the identification result of the elongation capability of the embryonic axis in the dry direct seeding rice. In three other methods for simultaneously evaluating the elongation capacity of the mesocotyl of the dry direct-seeded rice, the correlation coefficient between different batches is up to 0.88 (soil from fields is used), but the method can ensure that the correlation coefficient between the test and the identification results of different batches is up to 0.95, the formula of the commercialized nutrient soil is clear, and different researchers can completely duplicate the method.
Drawings
FIG. 1 shows phenotypic variation of 208 rice varieties in four different culture modes.
FIG. 2 is a graph showing the correlation between the mesocotyl length of 208 rice materials obtained in different culture modes in two replicates.
FIG. 3 is a plot of variation range of mesocotyl length for the MAGIC population over two trials.
FIG. 4 is a graph of the correlation of mesocotyl length in two experimental replicates.
Detailed Description
The present invention will be described in detail with reference to specific examples. The following examples will assist those skilled in the art in further understanding the invention, but are not intended to limit the invention in any way. It should be noted that variations and modifications can be made by persons skilled in the art without departing from the spirit of the invention. All falling within the scope of the present invention.
Example 1
The mesocotyl elongation ability of the same rice population (208 rice germplasms introduced by international rice) is respectively identified by a water culture method, a sand culture method, a soil culture method (field loam) and a nutrient soil (Germany K brand-TS 1 type). Each cultivation method was set up for 2 independent replicates. The specifications of the used hole discs are 54cm (length) x 28cm (width) x 11cm (height), 50 holes.
a. In the water culture method, gauze is flatly paved at the bottom of each hole, 15 plump seeds are flatly paved on the gauze, a hole tray is placed in a tray, water is injected into the tray until the water surface of the hole tray is 1cm higher than the seeds, then the hole tray and the tray are sealed and shaded by tinfoil paper, finally the sealed hole tray is placed in a climatic chamber with the temperature of 30 ℃, and the length of a mesocotyl shaft is measured by a ruler after the hole tray is cultured for 7 days.
b. A sand culture method: before sowing, filling each hole in the hole tray with sand to a position 6cm away from the top of the hole tray, and placing the hole tray in a tray filled with sand, wherein the depth of the sand is 3 cm. And (3) selecting 15 full dry seeds, flatly paving the seeds at the bottom of the hole, then covering the seeds in the hole tray with sand until the seeds are flush with the top of the hole tray to ensure that the sowing depth reaches 6cm, and watering the seeds through the hole tray until the tray at the bottom is saturated in water and has no open water. After watering, the weight of each tray and each hole tray is recorded and marked, and water is supplemented once every 24 hours until the marked weight is reached. And finally, placing the plug in a climatic chamber at the temperature of 30 ℃, sampling after the seeds germinate and emerge, and measuring the length of the mesocotyl by using a ruler.
c. The soil culture method comprises filling each hole in the hole tray with soil to a position 6cm away from the top of the hole tray, and placing the hole tray in a tray filled with soil to a depth of 3 cm. And (3) selecting 15 full dry seeds, paving the seeds at the bottom of the hole, covering the seeds in the hole tray with soil until the seeds are flush with the top of the hole tray to ensure that the sowing depth reaches 6cm, and watering the seeds through the hole tray until the tray at the bottom is saturated in water and has no open water. After watering, the weight of each tray and each hole tray is recorded and marked, and water is supplemented once every 24 hours until the marked weight is reached. And finally, placing the plug tray in a climatic chamber at the temperature of 30 ℃, sampling after the seeds germinate and emerge, and measuring the length of the mesocotyl by using a ruler.
d. The nutrient soil culture method comprises filling each hole in the hole tray with nutrient soil to a position 6cm away from the top of the hole tray before sowing, and placing the hole tray in a tray filled with nutrient soil with a depth of 3cm (or 500 g). Similarly, 15 full dry seeds are selected and spread at the bottom of the hole, then the seeds in the hole tray are covered with nutrient soil until the seeds are flush with the top of the hole tray, the sowing depth reaches 6cm, then the hole tray is watered, water leaks out from the hole below the hole, then the hole tray is taken out from the tray, the nutrient soil in the tray is watered to be saturated by tap water, the hole tray is put back into the tray, the total amount of watering is controlled to be 2000g, and then the height of the nutrient soil in the hole tray is supplemented to be flush with the hole opening by the smashed nutrient soil. After watering, the weight of each tray and each hole tray is recorded and marked, and water is supplemented once every 24 hours until the marked weight is reached. Finally, the tray was placed in a dark culture chamber at a temperature of 35 ℃ and a relative humidity of 50% -60%. Observing the rice seedling emergence condition every day, recording the seedling emergence sequence, sampling and photographing on the 3 rd day after the first variety emerges, and rapidly measuring the mesocotyl lengths of different rice varieties in the culture system by using an image fixing method.
And (3) analyzing test results:
the rice mesocotyl phenotype variation under different culture modes.
FIG. 1 shows phenotypic variation of 208 rice varieties in four different culture regimes, with the abscissa representing two independent experimental replicates for each culture regime. As can be seen from FIG. 1, the phenotypic variation of the 208 rice materials was significantly different, and the variation range of the hypocotyls was 0-4.8 cm in the soil culture (FIG. 1A), 0-2.1 cm in the sand culture (FIG. 1B), except for the extreme difference (FIG. 1B), 0-4.0cm in the water culture (FIG. 1C), and the maximum variation range was 0-5.1 cm in the nutrient soil culture (FIG. 1D). The larger phenotypic variation is the basis of genetic analysis, so the nutrient soil culture method proposed in this patent is superior to other methods in view of the variation range of mesocotyl length.
FIG. 2 reflects the correlation between the mesocotyl length of 208 rice materials obtained in different culture modes in two replicates. As can be seen, the correlation between mesocotyl length in two replicates is poor under both sand and hydroponic conditions, R20.5150 (FIG. 2B) and 0.5966 (FIG. 2C), respectively, are preferred in soil culture, R20.7858 (FIG. 2A) was reached, whereas R was cultured under nutrient soil conditions20.9133 (FIG. 2D) was achieved and the slope k was closest to 1, indicating that the nutrient soil culture method proposed in this patent is the best in reproducibility and stability among the four mesocotyl length determination methods.
Example 2
The specific operation steps of detecting the variation of the hypocotyl length in 395 MAGIC rice populations by utilizing the nutrient soil culture method are the same as those of the nutrient soil culture method.
And (3) analyzing test results:
the variation in mesocotyl length of the MAGIC population ranged from 0-4.8 cm, and was evenly distributed and continuous, with results that were essentially identical in both experimental replicates (FIG. 3). At the same time, the mesocotyl length was highly correlated in both experimental replicates (R)20.8459) (fig. 4), substantially consistent with the correlation results in example 11.
The foregoing description of specific embodiments of the present invention has been presented. It is to be understood that the present invention is not limited to the specific embodiments described above, and that various changes or modifications may be made by one skilled in the art within the scope of the appended claims without departing from the spirit of the invention. The embodiments and features of the embodiments of the present application may be combined with each other arbitrarily without conflict.
Claims (1)
1. A method for identifying the elongation capability of an embryonic axis in dry direct-seeded rice is characterized by comprising the following steps: the method comprises the following steps:
s1, smashing the nutrient soil, sieving the smashed nutrient soil with a 5-mesh sieve, and uniformly stirring the smashed nutrient soil and the sieved nutrient soil for later use;
s2, filling 500g of smashed nutrient soil into the tray, paving and compacting, placing the hole tray into the tray, filling a certain amount of smashed nutrient soil into each hole of the hole tray, and compacting to enable the height difference between the soil surface and the hole opening to be 6 cm;
s3, selecting full and uniform rice seeds with intact glumes, sowing the seeds in sequence, sowing 15 seeds in each hole, and uniformly distributing the 15 seeds at the bottom of each hole;
s4, uniformly covering the smashed nutrient soil after all the materials are sowed until the soil surface is flush with the opening of the hole tray, and weighing the tray, the hole tray and the total mass of the nutrient soil and the seeds in the hole tray as W1;
s5, thoroughly pouring the nutrient soil in the hole tray in a spraying mode, taking the hole tray out of the tray after water leaks out of the holes, pouring the nutrient soil in the tray to saturation with tap water, putting the hole tray back into the tray, controlling the total amount of watering to be 2000g, then, supplementing the height of the nutrient soil in the hole tray to be level to the hole opening with smashed nutrient soil, and weighing the total weight to be W2;
s6, placing the tray and the plug tray together in a dark culture room with the temperature of 35 ℃ and the relative humidity of 50% -60%, spraying once every 24 hours, and controlling the amount of sprayed water to keep the total weight of the sprayed water at the W2 level;
s7, observing the seedling emergence condition every day, recording the seedling emergence sequence of different varieties, and sampling and determining the mesocotyl length on the 3 rd day after the first variety emerges; taking out the seedlings from the plug tray in sequence, washing the seedlings clean, sequentially and flatly placing the seedlings growing uniformly in each material on a flat plate with scales, and taking a picture, wherein the camera position is required to be directly above the flat plate during taking the picture, the height is determined by placing the whole flat plate in a viewing frame, and the camera position is fixed by using a tripod; after all varieties are photographed in sequence, the mesocotyl length of each variety is measured by using Image J software according to a scale setting scale on a flat plate.
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| CN111802203A (en) * | 2020-06-02 | 2020-10-23 | 佛山科学技术学院 | Method for evaluating pea growth condition |
| CN116574829B (en) * | 2023-04-03 | 2024-03-12 | 中国农业科学院作物科学研究所 | Molecular marker linked with rice mesocotyl elongation gene qML3 and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101103699A (en) * | 2006-07-10 | 2008-01-16 | 曹立勇 | Method for selecting and breeding new variety of direct-seeded rice with mesocotyl length as index |
| CN106212136A (en) * | 2016-07-28 | 2016-12-14 | 四川省农业科学院水稻高粱研究所 | A kind of rice varieties top soil ability authentication method |
| CN107549010A (en) * | 2017-09-28 | 2018-01-09 | 吉林省农业科学院 | A kind of method of the high sorghum variety of simple and quick screening emergence rate |
| CN208657298U (en) * | 2018-08-29 | 2019-03-29 | 甘肃农业大学 | A kind of seed top soil capability determining instrument |
-
2019
- 2019-08-06 CN CN201910719624.1A patent/CN110326503B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101103699A (en) * | 2006-07-10 | 2008-01-16 | 曹立勇 | Method for selecting and breeding new variety of direct-seeded rice with mesocotyl length as index |
| CN106212136A (en) * | 2016-07-28 | 2016-12-14 | 四川省农业科学院水稻高粱研究所 | A kind of rice varieties top soil ability authentication method |
| CN107549010A (en) * | 2017-09-28 | 2018-01-09 | 吉林省农业科学院 | A kind of method of the high sorghum variety of simple and quick screening emergence rate |
| CN208657298U (en) * | 2018-08-29 | 2019-03-29 | 甘肃农业大学 | A kind of seed top soil capability determining instrument |
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| Title |
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| Identification of molecular markers for mesocotyl elongation in weedy rice;Hyun-Sook Lee 等;《Korean Society of Breeding Science》;20121231;第44卷(第3期);238-244 * |
| 基于幼苗图像分析鉴定水稻品种旱直播耐深播特性;冯芳君 等;《农业大数据学报》;20190630;第1卷(第2期);33-39 * |
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