CN112106649A - Rapid breeding method of imidazolinone herbicide-resistant rice germplasm - Google Patents
Rapid breeding method of imidazolinone herbicide-resistant rice germplasm Download PDFInfo
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- CN112106649A CN112106649A CN202010975939.5A CN202010975939A CN112106649A CN 112106649 A CN112106649 A CN 112106649A CN 202010975939 A CN202010975939 A CN 202010975939A CN 112106649 A CN112106649 A CN 112106649A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/04—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection
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Abstract
The invention discloses a method for quickly breeding rice germplasm with an imidazolinone herbicide resistant function, which utilizes the seed soaking of the herbicide with a specific concentration and the spraying of the hybridization separation progeny of the herbicide resistant material and the non-resistant material in the seedling stage to quickly and accurately remove the progeny material without resistance, so that the surviving plants are herbicide resistant and can normally grow, and the seed selection is convenient. The method comprises the steps of soaking seeds by utilizing the herbicide with a specific concentration, spraying the herbicide with the specific concentration in a seedling stage, wherein the seed soaking concentration is 0.03%, and the seed soaking time is 72 hours. The spraying concentration is 0.1 percent, and the spraying amount is 30ml/m2. The method can improve the efficiency and the accuracy of the herbicide-resistant germplasm breeding process.
Description
Technical Field
The invention belongs to the technical field of resistant rice breeding, and particularly relates to a rapid breeding method of rice germplasm with an imidazolinone herbicide resistance.
Background
Rice is the main food crop in China and even the world, and is the main heat source of nearly half of the population in the world. In all rice fields in rice growing areas of China, a large amount of weeds occur all the time. According to statistics, more than 200 kinds of weeds exist in the national rice field, and about 40 kinds of weeds (10-20 kinds distributed in each rice area) which are common, serious and most common occur. In our country, the means of manual weeding, farm weeding, chemical weeding and the like are mainly used for preventing and killing weeds in rice fields and weedy rice. The manual weeding and the farm tool weeding have high labor consumption and low work efficiency, can not prevent and remove weeds in a large area in time, can not remove field weedy rice by utilizing the existing chemical means, and can influence the growth of rice when being improperly sprayed. Therefore, the method for planting herbicide-resistant rice to prevent and kill weeds in fields and weedy rice by chemical means is the most effective way. The imidazolinone herbicide resistant rice germplasm is characterized in that the ALS gene generates basic group variation, acetolactate synthase (ALS) is the primary key enzyme in the branched chain amino acid synthetic pathway, and the herbicides taking the acetolactate synthase (ALS) as a target are divided into: sulfonylureas, imidazolinones, pyrimidine salicylates, and sulfonamides. Imidazolinone herbicides have excellent herbicidal activity against cyperaceae weeds, annual or perennial monocotyledonous weeds, broad-leaved weeds and weedy rice. The herbicide has the characteristics of high activity, broad weed control spectrum, strong selectivity and the like, and is an ideal herbicide at present. In recent years, scientists have obtained a batch of non-transgenic imidazolinone herbicide-resistant rice mutants through various ways, and some germplasm materials with better resistance performance have been successfully applied to rice breeding.
The prior invention patent 201710728532.0 discloses a method for culturing herbicide-resistant imazethapyr japonica rice variety with the assistance of molecular markers, which is characterized in that molecular markers closely linked with herbicide-resistant genes are utilized to amplify detected material DNA, the detected material and a donor Jinjing 818 banding pattern are compared by an electrophoresis means, and the banding pattern is consistent and has herbicide-resistant genes. But it suffers from the following drawbacks: (1) the detection coverage of the offspring population is less, and the offspring resources are easy to lose. (2) The late seed selection is time-consuming and labor-consuming. (3) True and false hybrids after crossing cannot be tested. (4) A situation occurs in which the detection is inaccurate. The main reasons for this are: (1) the molecular marker is used for detecting a large amount of progeny materials, which is time-consuming and labor-consuming, and valuable progeny resources are easily lost if the detection is not complete. (2) Experiments can be carried out only after the F1 generation grows out, and after identification, non-resistant single plants need to be manually removed in a field, and then the selection of the germplasm of the herbicide-resistant golden rice needs to be carried out. (3) The band pattern was examined only to see the presence or absence of the resistance gene, and a true hybrid and a false hybrid could not be examined in the F1 generation. (4) Since a linkage marker is used, a gene may be linked, and thus, detection may be inaccurate.
Disclosure of Invention
The invention aims to provide a rapid breeding method of rice germplasm with an imidazolinone herbicide resistance, which utilizes materials such as Jinjing 818, Jindao 372 and the like containing ALS genes as parents to carry out hybridization breeding on multiple types of non-transgenic herbicide resistance germplasm resources with high-quality and high-yield rice varieties, and can improve the efficiency and the accuracy of a breeding process of herbicide resistance planting.
In order to achieve the technical purpose, the invention is specifically realized by the following technical scheme:
a rapid breeding method of rice germplasm resistant to imidazolinone herbicides comprises the following steps:
1) selecting an imidazolinone herbicide-resistant variety as a male parent, selecting a high-quality and high-yield conventional non-resistant japonica rice variety as a female parent, and hybridizing to obtain F1Seed generation;
2) f is to be1The seeds are soaked in imazethapyr solution for 72 hours and then sown and raised, so that the germination of false hybrids is inhibited;
3) when the seedlings grow to the period of two leaves and one heart, the imidazole acetic niacin solution is used for spraying the seedlings to remove the false hybrid;
4) true hybrid F1Planting in rows and selfing to obtain FnOr backcrossing to obtain BCnFn；
5) F is to benOr BCnFnSeed generation is carried out by soaking seeds for 72 hours by using imazethapyr solution, and then seeding and seedling raising are carried out, so as to inhibit the seed of the nonreactive materialGerminating the seeds;
6) wait for FnOr BCnFnThe seedlings grow to the period of two leaves and one heart, and the imazethapyr solution is used for spraying the seedlings to kill the non-resistant materials.
Further, the imidazolinone herbicide resistant variety is selected from Jinjing 818 or Jindao 372.
Further, the mass concentration of the imazethapyr solution in the steps 2) and 5) is 0.03%, and the seed soaking time is 72 hours.
Further, the mass concentration of the imazethapyr solution in the steps 3) and 6) is 0.1 percent, and the spraying amount is 30ml/m2。
Further, the imazethapyr solution in the steps 3) and 6) is continuously sprayed twice at an interval of 3 days.
The invention has the beneficial effects that:
the method can improve the efficiency and the accuracy of the breeding process of herbicide-resistant planting, and the principle is that 0.03 percent of herbicide solution is used for soaking separated offspring for 72 hours, the nonresistant seeds after sowing are inhibited to germinate and can not grow into seedlings, and 30ml/m of herbicide solution with the concentration of 0.1 percent is sprayed again in the two-leaf one-heart period of the seedlings2(20L/mu in field), and killing residual non-resistant materials. Thereafter, the individual plants transferred to the field (offspring selection nursery) were resistant to the herbicide.
The method has obvious high efficiency in removing the non-resistant single plants from a large amount of progeny materials, hardly increases the workload compared with the normal seeding and seedling raising process, and has extremely high accuracy in removing the non-resistant single plants through experimental verification. In the later period, the existence of resistance does not need to be considered again in the seed selection in the selection nursery, and the seed selection efficiency is improved.
In addition, all the steps of the method are completed in the seedling raising period, so that the labor time is saved, and the transplanted single plant has resistance. And test processes such as sampling after transplanting, molecular identification and the like are not needed; after transplanting, manual cutting or pesticide spraying for killing in the field is not needed again, normal planting and breeding of surrounding common rice are not affected, and the later-stage seed selection efficiency is improved.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to specific embodiments of the present invention, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
The invention provides a method for quickly breeding rice germplasm with an imidazolinone herbicide resistance, which is characterized in that a herbicide-resistant material and a non-herbicide-resistant material are hybridized and separated to obtain progeny, the herbicide with a specific concentration is used for seed soaking and seedling stage spraying, the non-resistant progeny material is quickly and accurately removed, the surviving plants are herbicide-resistant and can normally grow, and the selection is convenient. The method comprises the steps of soaking seeds by using the herbicide with a specific concentration, spraying the herbicide with the specific concentration in a seedling stage, wherein the seed soaking concentration is 0.03 percent, the seed soaking time is 72 hours, and the spraying concentration is 0.1 percent. The method specifically comprises the following steps:
a. the screened imidazolinone herbicide resistant variety Jinjing 818 or Jindao 372 is used as a resistance gene donor parent, the other parent is selected according to breeding requirements, herbicide resistant material is used as a male parent, and the two parents are hybridized to obtain F1。
b. Hybrid F1Soaking seeds in 0.03% imazethapyr solution for 72 hr, taking out the seeds and sowing in seedling trays, and managing other water and fertilizer. Spraying imazethapyr solution with concentration of 0.1% to seedlings when the seedlings grow to the period of two leaves and one heart, wherein the spraying amount is 30ml/m2(20L/acre in the field), at which time the seedlings started to wither after 48 hours if there was no resistant material. This process can remove false hybrids that fail hybridization.
c. True hybrid F1Planting into lines, selfing to obtain FnOr backcrossing to obtain BCnFnStable single plants can be selected in each generation, and the progeny which is not resistant to herbicide in each generation is removed by the method in the seed soaking and seedling raising processes of each generation.
The operation steps of the invention are all in the rice seedling raising process. When seeds are soaked, the herbicide solution with specific concentration is added to inhibit the germination of the seeds of the offspring materials without resistance, and seedlings which can germinate and normally grow in a seedling tray or a seedling bed have the herbicide resistance. And spraying a certain amount of herbicide solution with specific concentration on the seedlings when the seedlings grow to the stage with two leaves and one heart (before transplanting), and rejecting few uninhibited non-resistant plants in the germination stage. Therefore, the invention can cover all progeny materials in the process of breeding herbicide-resistant rice germplasm, and is convenient and easy to implement.
Imidazolinone herbicide resistant gene as dominant gene in hybrid F1Expression is in time, so that F can be detected by using the present invention by crossing one parent having a herbicide-resistant gene as a male parent1The true and false of the hybrid.
The invention uses a phenotype identification method, and experiments prove that more than 99.9 percent of non-resistant offspring can be removed by utilizing the concentration and the time of the herbicide during seed soaking and the concentration of pesticide spraying in the two-leaf one-heart period, so that the accuracy is extremely high.
Example 2
Utilizing herbicides with different concentrations to soak seeds to kill non-herbicide-resistant materials, and setting the herbicide concentrations: 0. 0.0005%, 0.001%, 0.003%, 0.005%, 0.007%, 0.01%, 0.03%, 0.05%, 0.07%, 0.1%, 0.3%, 0.5%, 0.7%. As shown in tables 1 and 2, 0 to 13 means 0 to 0.7% from left to right, respectively.
TABLE 1 survival of materials at 14 concentration gradients from 0 to 0.7% (50 seeds sown)
| Concentration of | 0 | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | 13 |
| Cultivation 60 | 50 | 50 | 40 | 37 | 28 | 22 | 5 | 1 | 0 | 0 | 0 | 0 | 0 | 0 |
| Jindao 372 | 50 | 50 | 50 | 50 | 50 | 50 | 50 | 50 | 40 | 35 | 30 | 21 | 10 | 10 |
TABLE 2 survival of material at 3 seed soak times (50 seeds sown) with 0.01% -0.07% 4 concentration gradients
Experimental results show that by using 0.03% of herbicide to soak seeds for 72 hours, the herbicide-resistant materials can be effectively killed, and the effect is optimal.
Example 3
Spraying herbicide to remove non-resistant materials in a seedling stage of rice seedling tray planting simulation, and setting the concentration of the herbicide: 0. 0.04%, 0.06%, 0.08%, 0.1% and 0.12%, and spraying the herbicide twice when the rice grows to have two leaves and one core, wherein the interval is 3 days. The results are shown in Table 3.
TABLE 3 survival rates of seedlings after herbicide application in the seedling trays
| Variety/concentration | 0 | 0.04％ | 0.06％ | 0.08％ | 0.10％ | 0.12％ |
| Cultivation 60 | 100％ | 78％ | 43％ | 5％ | 0％ | 0％ |
| Jindao 372 | 100％ | 100％ | 100％ | 100％ | 100％ | 95％ |
The experimental result shows that the herbicide with the concentration of 0.1 percent can be used for killing the materials which are not resistant to the herbicide in the seedling stage.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (3)
1. A rapid breeding method of rice germplasm resistant to imidazolinone herbicides is characterized by comprising the following steps:
1) selecting an imidazolinone herbicide-resistant variety as a male parent, selecting a high-quality and high-yield conventional non-resistant japonica rice variety as a female parent, and hybridizing to obtain F1Seed generation;
2) f is to be1Soaking seeds with imazethapyr solution, and sowing and raising seedlings;
3) when the seedlings grow to the period of two leaves and one heart, the imidazole acetic niacin solution is used for spraying the seedlings to remove the false hybrid;
4) true hybrid F1Planting in rows and selfing to obtain FnOr backcrossing to obtain BCnFn；
5) F is to benOr BCnFnSoaking seeds with imazethapyr solution, and sowing and raising seedlings;
6) wait for FnOr BCnFnThe seedlings grow to the period of two leaves and one heart, and the imazethapyr solution is used for spraying the seedlings to kill the non-resistant materials.
2. The method for rapidly breeding the rice germplasm resistant to the imidazolinone herbicides according to claim 1, wherein the mass concentration of the imazethapyr solution in the steps 2) and 5) is 0.03%, and the seed soaking time is 72 hours.
3. The rapid breeding method of rice germplasm with imidazolinone herbicide resistant rice as claimed in claim 1, wherein the imazethapyr solution mass concentration in steps 3) and 6) is 0.1%, and the spraying amount is 30ml/m2。
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