CN108935079B - Simplified seed production method of rape hybrid F1 seed - Google Patents
Simplified seed production method of rape hybrid F1 seed Download PDFInfo
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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
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Abstract
The invention provides a simplified method for producing seeds of rape hybrid F1, which comprises selecting a Bolima cytoplasmic male sterile line or a conventional inbred line (or a conventional variety) as a female parent, selecting a restorer line or a conventional inbred line (or a conventional variety) which has self-incompatibility characteristic and resistance to sulfonylurea herbicide as a male parent, mixing the female parent and the male parent under isolation conditions, sowing the uniformly mixed seeds in a field of a rape seed production area, spraying 1-time male sterilant containing sulfonylurea herbicide at the initial stage of bud emergence, spraying 1 time after 15-20 days, mixing the female parent and the male parent during maturation, and producing the hybrid F1 seeds. The invention has simple breeding program, saves labor cost, money and labor.
Description
Technical Field
The invention belongs to the technical field of rape hybrid seed production, and particularly relates to a hybrid seed production method for producing hybrid F1 seeds by using a Polima cytoplasmic male sterile line or a conventional inbred line (or a conventional variety) and a Polima cytoplasmic male sterile restoration line or a conventional inbred line (or a conventional variety) which simultaneously has self-incompatibility characteristics and resistance to sulfonylurea herbicides due to gene mutation of acetolactate synthase.
Background
The rape hybrid vigor is obvious, the yield can be increased by 20-30% generally, and the yield increase amplitude can reach 50% after good combination hybridization, so that the popularization and application of the hybrid rape are rapidly developed, and only the area of the hybrid rape in China accounts for more than 70% of the total area of the hybrid rape. Heterosis utilization is an important way to increase crop yield. Rape heterosis utilization approaches are many, and mainly include cytoplasmic male sterility, nuclear male sterility, self-incompatibility, chemical male killing and the like. Among cytoplasmic male sterility, bolima cytoplasmic male sterility is an important type of cytoplasmic male sterility. In chemical male killing, the chemical male killing agent currently used in production is mainly a mixture containing tribenuron-methyl in low concentration. Researches prove that the target of tribenuron-methyl is acetolactate synthase, the tribenuron-methyl solution with high concentration can kill rape, and the tribenuron-methyl solution with low concentration can induce male sterility without obvious influence on the growth of the rape. When some sites of the acetolactate synthase gene are mutated, the rape can generate resistance to tribenuron-methyl.
At present, the method for producing seeds of rape Bolima cytoplasmic male sterile hybrid F1 is to plant the Bolima cytoplasmic male sterile line (female parent) and the restorer line (male parent) in a certain row ratio in the field, and harvest F1 hybrid seeds on the male sterile line (female parent) when the seeds are mature. The method for producing seeds of hybrid F1 by chemical emasculation includes such steps as planting the selfing line A (or conventional variety A) (female parent) and selfing line B (or conventional variety B) (male parent) in field according to a certain row ratio, spraying 1-time male sterilant containing tribenuron-methyl at the early stage of budding, spraying again after 15-20 days, and harvesting F1 hybrid seeds from the selfing line A (or conventional variety A) (female parent) when the seeds are mature. The hybrid F1 is produced by using the cytoplasmic male sterile line of Bolima and chemical male killing, and has the disadvantages that the female parent and the male parent need to be separately planted according to a certain row ratio during planting, the male parent needs to be separately harvested during harvesting, or the male parent is cut off after the male parent withers flowers, and F1 hybrid seeds are harvested from the female parent when the seeds are mature. At the same time, strict supervision is required during seed harvest to avoid mixing the male parent seed into the hybrid F1 seed that has grown on the female parent. Obviously, the existing method wastes time and labor and has higher labor cost. In addition, fertility of the balima cytoplasmic male sterile line is often affected by temperature, and different amounts of fertile pollen are generated, so that the sterile line can self-cross or cross and fruit, the purity of the F1 seed is reduced, and certain production risk exists.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provide a simplified method for producing rape hybrid F1 seeds by using a Bolima cytoplasmic male sterile line or a conventional inbred line (or a conventional variety).
In order to realize the purpose of the invention, the inventor finally obtains the following simple and efficient technical scheme by combining the research experience of rape male sterility hybridization for many years:
a simplified method for preparing the hybrid F1 seeds of rape includes such steps as choosing the cytoplasmic male sterile line of Polima or conventional selfing line (or conventional variety) (female parent) and the restoring line with self-incompatibility and resistance to sulfonylurea herbicide caused by the mutation of acetolactate synthetase gene or selfing line (or conventional variety) (male parent) to obtain the hybrid F1 seeds, mixing the female parent with male parent in a certain proportion under isolation condition, sowing the uniformly mixed seeds in the field of rape seed preparing area, spraying 1-time formulated male sterilant containing sulfonylurea herbicide at the beginning of bud, spraying 1-time herbicide after 15-20 days, and mixing female parent with male parent when it is mature to obtain the hybrid F1 seeds.
Further preferably, the suitable proportion of the female parent seeds to the male parent seeds in the method is 70-90%: 10 to 30 percent.
Further preferably, the method for preparing the hybrid rape F1 seed is simplified as described above, wherein the isolation conditions comprise natural isolation and artificial isolation.
Further preferably, the simplified method for producing the hybrid rape F1 seed is as described above, wherein the artificial broadcast sowing is carried out at 6000-30000 plants per mu, the mechanical or artificial drill sowing is carried out at a row spacing of 10-25 cm and a plant spacing of 5-15 cm.
Further preferably, the male sterilant with the formula containing tribenuron-methyl is sprayed at the early budding stage, and the same male sterilant is sprayed again after 15 to 20 days.
Compared with the prior art, the hybrid seed production method has the advantages and the progressiveness that:
(1) in the sowing process, the traditional method is to separately plant the female parent and the male parent according to a certain row ratio, which takes labor and time; the invention has the advantages of mixed planting of female parent and male parent, simple breeding procedure, labor cost saving, money saving and labor saving.
(2) When harvesting, the traditional method is to separately harvest the female parent and the male parent, or to cut off the male parent in advance; the invention directly mixes the female parent and the male parent for harvest, has simple operation and further saves the labor cost.
(3) The male parent has the characteristic of self-incompatibility, so that the male parent cannot normally fruit, the flowering period is prolonged, pollination is facilitated, and the yield of F1 seeds is increased.
(4) The male parent cannot fruit normally due to the self-incompatibility characteristic, and the risk of mixing the male parent seeds into the F1 seeds is reduced fundamentally.
(5) In the traditional method, when the polima cytoplasmic male sterile line is used for producing seeds, hybrid plants can pollinate the sterile line due to incomplete impurity removal of a female parent, so that the purity of the hybrid is influenced; meanwhile, the fertility of the Bolima cytoplasmic male sterile line is easily influenced by temperature, and the fertility instability of the sterile line can be self-bred and the brother and sister cross fructification can also influence the purity of the hybrid. The invention can simultaneously perform emasculation on the hybrid plant and the female parent in the female parent row by spraying the emasculation agent, eliminates the risks of pollination of the female parent by the hybrid plant and sterile line selfing or brother-sister crossing, is beneficial to improving the seed purity and reduces the production risk.
(6) In the traditional method, when the male killing is carried out by chemical male killing, the male killing agent is sprayed to the female parent row to avoid spraying the male parent row, otherwise, the male parent is killed, the pollen amount of the male parent is reduced, and the seed production yield is influenced. In order to avoid spraying the male killing agent on the male parent, a certain isolation measure is adopted in the traditional method. The male parent has sulfonylurea herbicide resistance gene, so that the male fertility of the male parent is not affected by the tribenuron-methyl spraying, and isolation measures are not needed.
Detailed Description
The present invention is further illustrated by the following specific examples. In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention. The present invention may be embodied in many different forms without departing from the spirit or essential characteristics thereof, and it should be understood that various changes in form and details can be made therein by those skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims. The following experimental methods are conventional methods and techniques in the art unless otherwise specified. In addition, in the examples, the birima cytoplasmic male sterile line 616A is a new variety cultivated by the present inventors, and the right of new plant variety is obtained in 2018, 1 month and 2 days, and the right of variety is CNA 20141083.8.
Example 1: breeding of Bolima cytoplasmic male sterile restorer line 7-5STR with self-incompatibility characteristic and resistance to sulfonylurea herbicide caused by acetolactate synthase gene mutation
(1) Crossing with a self-incompatible single plant (the genotype is BnS-1300BnS-1300) separated from an F2 progeny of Hua YOUZAO 95 by using Hua YOUZAO No. 12 restorer line 7-5 as a female parent (the genotype is BnS-7BnS-7) to obtain F1.
(2) Backcrossing the F1 as female parent with Hua-YOU-ZAO No. 12 restorer line 7-5 to obtain BC 1.
(3) And (3) carrying out DNA molecular detection on the BC1 group single plants by using a DNA molecular marker linked with the self-incompatibility gene BnS-1300, and screening the single plants with heterozygous genotypes (BnS-1300BnS-7) at the self-incompatibility gene locus.
(4) F1 was obtained by crossing Hua-YOU hybrid No. 12 restorer line 7-5 (genotype: BnaAHAS3_ BnaAHAS3) as female parent with 655 (genotype: Bnaahas3_ Bnaahas3) which is resistant to sulfonylurea herbicides by acetolactate synthase gene mutation.
(5) Backcrossing 7-5 by using F1 obtained in the step (4) as a female parent to obtain BC 1.
(6) The individual plants of the BC1 group are detected by using molecular markers linked with the acetolactate synthase gene BnaAHAS3, and the individual plants with heterozygous genotypes (BnaAHAS3Bnaahas3_) at the acetolactate synthase gene locus are screened out.
(7) Individuals selected from the two BC1 populations were crossed to obtain a double cross F1.
(8) The DNA molecular markers linked with the self-incompatible gene BnS-1300 and acetolactate synthase gene BnaAHAS3 are respectively used for DNA molecular detection of the single plant of the compound cross F1 group, and the single plant which is heterozygous genotype (BnS-1300BnS-7+ BnaAHAS3Bnaahas3) at the gene locus of the self-incompatible acetolactate synthase is screened out.
(9) And (4) carrying out backcross on selected individuals in the backcross F1 with 7-5 to obtain the backcross BC 1.
(10) The individual plants of the double-cross BC1 population are subjected to DNA molecular detection by using DNA molecular markers linked with self-incompatible genes BnS-1300 and acetolactate synthase genes BnaAHAS3 respectively, and the individual plants which are heterozygous genotypes (BnS-1300BnS-7+ BnaAHAS3Bnaahas3) at the self-incompatible acetolactate synthase gene sites are screened out at the same time.
(11) And (3) carrying out backcross on selected single plants in the compound cross BC1 with 7-5 to obtain compound cross BC 2.
(12) And (3) carrying out DNA molecular detection on the single plants of the double-cross BC2 population by using DNA molecular marker pairs linked with self-incompatible genes BnS-1300 and acetolactate synthase genes BnaAHAS3 respectively, and screening out the single plants of which the gene loci of the self-incompatible acetolactate synthase genes are heterozygous genotypes (BnS-1300BnS-7+ BnaAHAS3Bnaahas 3).
(13) And (3) selecting single plants from the compound BC2 for selfing to obtain compound BC2F 2.
(14) And (3) carrying out DNA molecular detection on the single plants of the double-cross BC2F2 population by using DNA molecular marker pairs linked with self-incompatible genes BnS-1300 and acetolactate synthase genes BnaAHAS3 respectively, and screening out the single plants which are homozygous genotypes (BnS-1300BnS-1300+ BnaAHAS3BnaAHAS3) at the gene loci of the self-incompatible acetolactate synthase genes.
(15) And (3) selfing (artificial supplementary pollination) is carried out on the single plants which are in the selfing incompatible acetolactate synthase gene locus as the homozygous genotype (BnS-1300BnS-1300+ BnaAHAS3BnaAHAS3) in the double-cross BC2F2 population at the same time to obtain the double-cross BC2F 3.
(16) And (3) carrying out agronomic character, self-incompatibility and tribenuron-methyl resistance identification on the compound cross BC2F3 family, screening out a family which is self-incompatible and has tribenuron-methyl resistance and a phenotype closest to 7-5, carrying out mixed propagation (artificial supplementary pollination) and mixed harvesting to obtain 7-5 STR.
Example 2: hybrid seed production of rape by using cytoplasmic male sterility
The method of the invention comprises the steps of producing hybrid F1 seeds by using a Polima cytoplasmic male sterile line 616A and a restorer line 7-5STR which simultaneously has self-incompatibility characteristic and resistance to sulfonylurea herbicides caused by acetolactate synthase gene mutation, and the hybrid seed production method comprises the following specific steps:
(1) 616A and 7-5STR seeds were mixed in 80%: mixing at a ratio of 20%.
(2) And under the condition of isolating an artificial net room, planting the uniformly mixed seeds according to the row spacing of 20cm and the plant spacing of 10 cm.
(3) Spraying chemical pesticide WP2 (the first step is to compound 5 g chemical pesticide WP2 wettable powder and 1000ml water into mother liquid, the second step is to compound 50ml mother liquid and 15000ml water into diluent, and the third step is to spray the diluent directly onto rape leaf). After 15 days, spraying again on sunny days.
(4) When the seeds are mature, the sterile line and the restorer line are harvested in a mixed mode, and statistics shows that the purity of the hybrid is 95.6%.
Example 3: hybrid seed production of rape by using cytoplasmic male sterility
(1) 616A and 7-5STR seeds were mixed in 80%: mixing at a ratio of 20%.
(2) Under the condition of isolating an artificial net room, the seeds which are uniformly mixed are planted according to the density of 25000 plants per mu.
(3) Spraying chemical pesticide WP2 (the first step is to compound 5 g chemical pesticide WP2 wettable powder and 1000ml water into mother liquid, the second step is to compound 50ml mother liquid and 15000ml water into diluent, and the third step is to spray the diluent directly onto rape leaf). After 15 days, spraying again on sunny days.
(4) When the seeds are mature, the sterile line and the restorer line are harvested in a mixed mode, and statistics shows that the purity of the hybrid is 94.8%.
Example 4: chemical male-killing hybrid seed production for rape
The method of the invention comprises the steps of producing hybrid F1 seeds by utilizing the conventional rape variety 'Zhongshuang 11' and a restorer line 7-5STR which simultaneously has self-incompatibility characteristic and resistance to sulfonylurea herbicides caused by gene mutation of acetolactate synthase, and the hybrid seed production method comprises the following specific steps:
(1) the method comprises the following steps of mixing 80% of conventional rape variety 'Zhongshuang 11' and 80% of restorer line 7-5STR seeds with self-incompatibility characteristics: mixing at a ratio of 20%.
(2) Under the condition of natural isolation, the seeds which are uniformly mixed are planted according to the row spacing of 20cm and the plant spacing of 10 cm.
(3) Spraying chemical pesticide WP2 (the first step is to compound 5 g chemical pesticide WP2 wettable powder and 1000ml water into mother liquid, the second step is to compound the mother liquid into 110ml water and 15000ml water into diluted liquid, and the third step is to spray the diluted liquid onto rape leaf directly and homogeneously). After 15 days, 80ml of mother liquor and 15000ml of water are taken to prepare a diluent, and the diluent is sprayed once again on sunny days.
(4) When the seeds are mature, the sterile line and the restorer line are harvested in a mixed mode, and statistics shows that the purity of the hybrid is 91.2%.
Example 5: chemical male-killing hybrid seed production for rape
The method of the invention comprises the steps of producing hybrid F1 seeds by utilizing the conventional rape variety 'Zhongshuang 11' and a restorer line 7-5STR which simultaneously has self-incompatibility characteristic and resistance to sulfonylurea herbicides caused by gene mutation of acetolactate synthase, and the hybrid seed production method comprises the following specific steps:
(1) the method comprises the following steps of mixing 80% of conventional rape variety 'Zhongshuang 11' and 80% of restorer line 7-5STR seeds with self-incompatibility characteristics: mixing at a ratio of 20%.
(2) Under the condition of natural isolation, the seeds mixed uniformly are planted according to the density of 25000 plants per mu.
(3) Spraying chemical pesticide WP2 (the first step is to compound 5 g chemical pesticide WP2 wettable powder and 1000ml water into mother liquid, the second step is to compound the mother liquid into 110ml water and 15000ml water into diluted liquid, and the third step is to spray the diluted liquid onto rape leaf directly and homogeneously). After 15 days, 80ml of mother liquor and 15000ml of water are taken to prepare a diluent, and the diluent is sprayed once again on sunny days.
(4) When the seeds are mature, the sterile line and the restorer line are harvested in a mixed mode, and statistics shows that the purity of the hybrid is 90.8%.
Claims (6)
1. A simplified method for preparing the seeds of rape F1 hybrid includes such steps as choosing the cytoplasmic male sterile line or selfing line or conventional variety of Polima as female parent, choosing the restoring line with self-incompatibility and resistance to sulfonylurea herbicide, selfing line with self-incompatibility and resistance to sulfonylurea herbicide, or conventional variety with self-incompatibility and resistance to sulfonylurea herbicide as male parent, mixing the female parent with male parent seeds, sowing the uniformly mixed seeds in the field of rape seed preparing area, spraying 1-time male killer containing sulfonylurea herbicide at the beginning of bud, spraying again after 15-20 days, and mixing female parent with male parent to obtain hybrid F1.
2. The simplified method for producing hybrid rape hybrid F1 seed according to claim 1, wherein the ratio of the female parent to the male parent is 70-90%: and (3) uniformly mixing 10-30% of the mixture and then sowing.
3. The simplified method for producing hybrid rapeseed plant F1 seed as claimed in claim 1, wherein said isolation conditions include natural isolation and artificial isolation.
4. The simplified method for producing seeds of rape hybrid F1 as claimed in claim 1, wherein 6000-30000 plants per mu are planted by manual sowing, the row spacing of mechanical or manual drilling is 10-25 cm, and the row spacing is 5-15 cm.
5. The method for producing simplified canola hybrid F1 seed as claimed in claim 1, wherein the sulfonylurea herbicide is tribenuron-methyl.
6. The simplified seed production method of rape hybrid F1 seeds as claimed in claim 1, wherein the male sterilant containing tribenuron-methyl is sprayed 1 time at the early stage of budding, and the male sterilant containing tribenuron-methyl is sprayed 1 time at intervals of 15-20 days.
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| CN110741927B (en) * | 2019-11-18 | 2021-04-09 | 江苏省农业科学院 | Chemical emasculation method of herbicide-resistant rape mutant M342 and application thereof |
| CN112056205A (en) * | 2020-09-21 | 2020-12-11 | 夏立山 | Method for mechanically producing seeds of rapes based on seed coat color genetic marker |
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Effective date of registration: 20220914 Address after: Buildings B7 and B8, Bio-industry (Jiufeng) Innovation Base, No. 666 Gaoxin Avenue, Donghu New Technology Development Zone, Wuhan City, Hubei Province 430014 Patentee after: WUHAN LIANNONG SEED TECHNOLOGY Co.,Ltd. Address before: 430070 No. 1 Lion Rock street, Hongshan District, Hubei, Wuhan Patentee before: HUAZHONG AGRICULTURAL University Patentee before: WUHAN LIANNONG SEED TECHNOLOGY Co.,Ltd. |