Method for creating flowering Chinese cabbage cytoplasmic male sterile line

Abstract

The invention discloses a method for creating a cytoplasmic male sterile line of flowering Chinese cabbage, which comprises the following steps of: (1) selfing Zhengcheng tip leaf flowering Chinese cabbage to find a sterile plant; (2) carrying out test cross on sterile plants and sister plants appearing in the Zhengcheng sharp-leaf flowering cabbage inbred line to obtain test cross seeds; (3) planting the test-cross seeds, observing fertility and nectary conditions in a flowering period, backcrossing sterile plants and corresponding sister plants in a group with the test-cross offspring sterility rate of more than 50%, screening out a group with the sterility rate of 100% from backcross offsprings, and selfing the sterile plants in the group with the corresponding sister plants to obtain the sister plants with the infertility rate of more than 50% and the infertility rate of more than 100% in the group with anther degradation and normal pistil and nectary. Selfing the fertile sister plants to obtain offspring to form a maintainer line, and performing test crossing on the sterile plants and the maintainer line to obtain a matched sterile line. According to the method, transformation of a sterile source is not needed, time is saved, cost is low, the sterile line preserves completeness of nectary genes, nectary is normal, the podding rate is high, the maturing rate is high, and the seed yield is high.

Description

Technical Field

[0001] The invention relates to a method for creating a cauliflower sterile line, in particular to a method for creating a cauliflower cytoplasmic male sterile line. Background Art

[0002] Chinese cabbage Brassica parachinensis Bailey ), its botanical morphological characteristics are basically similar to other cabbage vegetables, and its flower stalk is the edible organ. Due to its unique flavor, short growth cycle and high multiple cropping index, it has become one of the main vegetables in South China. It can be exported overseas and earn foreign exchange, and is known as the "Queen of Vegetables" and "Crown of Vegetables".

[0003] Chinese cabbage is a cross-pollination vegetable with bisexual flowers (monosexual flowers). The breeding methods include selective breeding, hybrid breeding, haploid breeding, transgenic breeding, etc. The hybrid first generation shows strong hybrid vigor, but its flower organs are small. To utilize hybrid vigor, the problem of hybrid seed production must be solved first. At present, Chinese cabbage hybrid breeding is mainly carried out through the following ways:

[0004] (1) Self-incompatible lines are used for hybrid breeding. However, there are still some problems in the reproduction of self-incompatible lines that are difficult to overcome. For example, the bud stage mainly relies on manual bud pollination and emasculation, which is time-consuming, labor-intensive and costly. It is difficult to breed 100% self-incompatible parents, and the purity of the first generation of hybrids is not high. Multiple generations of self-pollination are also prone to plant vitality decline. (2) The use of cytoplasmic male sterile lines to produce first-generation hybrids can eliminate the need for artificial detasseling, and has shown obvious advantages in the reproduction and preservation of parents and the purity of first-generation hybrid seeds. For breeders and breeding institutions, it increases the controllability of new varieties, prevents the loss of parents, and enhances commerciality and competitiveness.

[0005] Therefore, using cytoplasmic male sterile lines to formulate new hybrid varieties has become the current research status of breeders.

[0006] Cytoplasmic male sterility (CMS) is a phenomenon that is based on maternal inheritance and cannot produce viable or fertile pollen. Cytoplasmic male sterility is first manifested in the morphology of floral organs, most of which are characterized by degeneration of stamens, short corollas, and filaments. The main ones currently used are derived from Pol CMS and Ogu CMS (References: Breeding of a new Chinese cabbage variety Nongfeng 70-day Chinese cabbage heart, Chinese Vegetables, 2024(3): 128-130; Breeding of a new Chinese cabbage variety Yutian No. 3 Chinese cabbage heart, Chinese Vegetables, 2024(1): 118-120). The breeding process is time-consuming and generally requires 6 to 7 generations of breeding. In addition, the continuous backcrossing process using the target material as the male parent can easily cause the loss of nectary genes. Chinese cabbage is a cross-pollinating plant of the Cruciferae family, and the presence of nectaries can increase insect pollination. Therefore, the current sterile seed production generally has a low fruit set rate, which directly leads to high seed production costs and seriously hinders the promotion and application of hybrids.

[0007] It should be noted that the information disclosed in the above background technology section is only used to enhance the understanding of the background of the present disclosure, and therefore may include information that does not constitute the prior art known to ordinary technicians in the field. Summary of the invention

[0008] The purpose of the present invention is to provide a method for creating a cytoplasmic male sterile line of Chinese cabbage, which method does not require the transfer of sterile sources, is time-saving and low-cost, and the sterile line preserves the integrity of the nectary gene, the nectary is normal, the pod setting rate is high, the fruit setting rate is high, the seed yield is high, the hybrid purity is high, and the line can be stably transplanted to other materials for utilization.

[0009] In order to achieve the above object, the present invention provides a method for creating a cytoplasmic male sterile line of Chinese cabbage, the method comprising the following steps: (1) Self-pollination of Zengcheng Jianyecaixin revealed sterile plants, the anthers of which were degenerate, but the pistils and nectaries were normal; (2) Test crosses were conducted with sterile plants and sister plants from the Zengcheng Jianye Chinese cabbage inbred line to obtain test cross seeds, which were then planted to observe fertility and nectary conditions during the flowering period. The sterility rate of the test cross offspring was greater than 50%. (3) In the population with a sterility rate greater than 50% in the test cross progeny in step (2), backcrossing is performed between the sterile plants in the population and the corresponding sister plants, and a population with a sterility rate of 100% is screened out from the backcross progeny. The anthers in the population are degenerate, and the pistils and nectaries are normal. The sterile plants in the population are self-pollinated with their corresponding sister plants, and the fertile sister plants are self-pollinated to obtain progeny to form a maintainer line. The sterile plants are test-crossed with the maintainer line to obtain a matching sterile line.

[0010] Preferably, in step (1), the Zengcheng sharp-leaved Chinese cabbage variety is planted, bagged before the flower buds open during the flowering period to prevent pollen cross-pollination, artificially assisted self-pollination is performed several times, and the seeds are threshed and harvested after they mature. In the next planting season, the self-pollinated first-generation seeds are planted and self-pollinated again to obtain the second-generation self-pollinated materials; in the next planting season, the second-generation self-pollinated materials are planted, and sterile plants with degenerated anthers and normal pistils and nectaries appear in the population.

[0011] Preferably, in step (2), a number of sister plants are selected from the group where sterile plants are found and bagged with sulfuric acid paper, and flower branches of the sterile plants that are the same number as the sister plants are selected and bagged, and the sterile branches of the sterile plants are test-crossed with the sister plants one by one, and the test-cross seeds are harvested after the seeds mature, and a group with a sterility rate greater than 50% is selected from the test-cross offspring.

[0012] Preferably, in step (2), in the next planting season, several test cross seeds harvested from the maintenance of sterile plants are planted in several plots, and the corresponding several self-pollinated materials are planted in adjacent plots; the fertility and nectary conditions are observed during the flowering period, and if the sterility rate in the test cross offspring group is found to be >50%, several sterile plants are selected from the group, and several flower branches are bagged, and at the same time, several plants are selected from the corresponding inbred lines and backcrossed with the several sterile plants one by one; the backcross seeds are planted in several plots, and the corresponding inbred lines are planted adjacent to each other, and a 100% sterile group appears in the backcross offspring of the group, and the nectaries are normal, the sterile plants in the group are self-pollinated with their corresponding sister plants, the fertile sister plants are self-pollinated to obtain offspring to form a maintenance line, and the sterile plants are test-crossed with the maintenance line to obtain a matching sterile line.

[0013] Another object of the present invention is to provide an application of the method for creating a cytoplasmic male sterile line of rapeseed, wherein the sterile line obtained by the method for creating a cytoplasmic male sterile line of rapeseed and a stable inbred line are combined to select and breed a first-generation hybrid variety.

[0014] Preferably, the stable inbred line is selected from Beet Heart No. 31.

[0015] The method for creating the cauliflower cytoplasmic male sterile line of the present invention has the following advantages: (1) The method of the present invention saves time in breeding. The sterile lines and maintainer lines are obtained quickly and accurately in two planting seasons by cross-crossing the sterile plants and sister plants that appear within three generations of the Zengcheng Jianye Chinese Cabbage sterile source. (2) The method of the present invention does not require the conversion of existing sterile sources. The sterile source is obtained by high-generation self-pollination of Zengcheng Jianye Chinese cabbage heart and is quickly obtained within three generations of self-pollination. It does not need to use existing sterile sources such as Ogura and Polima for conversion; (3) The method of the present invention has stable sterility and low seed production cost. The NCMS sterile line has normal flowering, degenerate anthers, no pollen, normal pistils and nectaries, a sterility rate of 100%, high natural pollination pod and fruit setting rates, and high seed yield; (4) According to the method of the present invention, the sterile source can be stably transferred, and the sterile source in the sterile line can be stably transferred to materials such as Teyouqing 70 Chinese cabbage heart and No. 31 sugar beet heart, and transferred into a new sterile line. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The present invention is a flow chart of the method for creating the cauliflower cytoplasmic male sterile line.

[0017] Figure 2 These are plane photographs of the floral organs of the sterile Zengcheng sharp-leaved Chinese cabbage plant (left) and the corresponding fertile plant (right) obtained in Example 1 of the present invention.

[0018] Figure 3 These are side photographs of the floral organs of the sterile Zengcheng sharp-leaved Chinese cabbage plant (left) and the corresponding fertile plant (right) obtained in Example 1 of the present invention.

[0019] Figure 4 This is a plant photo of NCMS obtained in Example 1 of the present invention.

[0020] Figure 5 This is a plant photo of Zengcheng Jianye-5-7-2-3-4-1-3-4 in Application Example 1 of the present invention.

[0021] Figure 6 This is a plant photo of the early-maturing Chinese cabbage 2-1-2-1-1 in Application Example 1 of the present invention.

[0022] Figure 7 This is a photo of plant 2023048 obtained using the Zengcheng sharp-leaved Chinese cabbage sterile plant in Application Example 1 of the present invention.

[0023] Figure 8 This is a photo of the plant 2023051 obtained using the Zengcheng sharp-leaved Chinese cabbage sterile plant in Application Example 1 of the present invention.

[0024] Fig. 9 This is a plant photo of Teyouqing 70 Chinese cabbage heart-4-1-3-1 obtained by using Zengcheng sharp-leaved Chinese cabbage heart sterile plants in Application Example 2 of the present invention.

[0025] Fig.10 The following are photos of pod formation of NCMS-31 and PCMS-31 in Application Example 2 of the present invention.

[0026] Fig.11 The following are photos of the pod numbers of NCMS-31 and PCMS-31 in Application Example 2 of the present invention. DETAILED DESCRIPTION

[0027] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0028] It should be noted that if the specific conditions are not specified in the examples, the experiments were carried out according to conventional conditions or the conditions recommended by the manufacturer. If the manufacturers of the reagents or instruments are not specified, they are all conventional products that can be purchased from the market.

[0029] In the present invention, all features defined in the form of numerical ranges or percentage ranges, such as values, quantities, contents and concentrations, are only for brevity and convenience. Accordingly, the description of numerical ranges or percentage ranges should be deemed to have included and specifically disclosed all possible secondary ranges and individual values ​​within the range (including integers and fractions).

[0030] The features mentioned in the present invention can be combined arbitrarily, as long as there is no contradiction in the combination of these features, all possible combinations should be considered as the scope of this specification. Each feature disclosed in the specification can be replaced by any alternative feature that can provide the same, equal or similar purpose. Therefore, unless otherwise specified, the disclosed features are only general examples of equal or similar features.

[0031] Example 1 A method for creating a cytoplasmic male sterile line of Chinese cabbage, see Figure 1 , which specifically includes the following steps: (1) Discovery of sterile plants: Sterile plants were found in the self-pollinated materials of Zengcheng jianye caixin.

[0032] The high-quality Zengcheng Jianye Chinese Cabbage variety (Zengcheng Jianye Chinese Cabbage grown by farmers in Zengcheng District, Guangzhou City, Guangdong Province) was selected for planting. Three plants were selected from the planting population. They were covered with sulfuric acid paper bags before the buds opened during the flowering period to prevent pollen from crossing. They were tied tightly with paper clips, numbered and labeled, and artificially assisted self-pollination was performed 1 to 2 times. After the seeds matured, they were numbered, threshed and the first-generation self-pollinated seeds were harvested. In the next planting season, three self-pollinated first-generation seeds were planted in three plots respectively. Similarly, three plants were selected from the populations of the three plots for self-pollination to obtain 9 second-generation self-pollinated materials. Similarly, in the next planting season, the 9 second-generation self-pollinated materials were planted in 9 plots respectively. In the population numbered Zengcheng Jianye Chinese Cabbage-5-1, sterile plants with degenerated anthers and normal pistils and nectaries appeared. This variability was common in self-pollination for three generations or more, and the probability of occurrence increased with each generation. The sterile plants were numbered NCMS, and their plant photos can be found in Figure 4 .

[0033] (2) Maintenance of sterile plants: Test crosses were conducted between sterile plants and fertile sister plants from the Zengcheng Jianye Chinese cabbage inbred line, and a population with a sterility rate greater than 50% was selected from the test cross progeny.

[0034] After the discovery of NCMS, 8 sister plants were selected from the population of Zengcheng Jianye Caixin-5-1, bagged with sulfuric acid paper, and numbered as Zengcheng Jianye Caixin-5-1-1, Zengcheng Jianye Caixin-5-1-2, Zengcheng Jianye Caixin-5-1-3, Zengcheng Jianye Caixin-5-1-4, Zengcheng Jianye Caixin-5-1-5, Zengcheng Jianye Caixin-5-1-6, Zengcheng Jianye Caixin-5-1-7, Zengcheng Jianye Caixin-5-1-8, and artificially assisted pollination was carried out during the bud stage. In NCMS, 8 flower branches were selected and covered with sulfuric acid paper bags respectively, and the sterile branches of the sterile plant NCMS were test-crossed with sister plants one by one: NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-1, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-2, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-3, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-4, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-5, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-6, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-7, NCMS×Zengcheng Sharp-leaved Chinese Cabbage Heart-5-1-8. After the seeds matured, 8 test-cross seeds were harvested.

[0035] In the next planting season, the eight test cross seeds harvested above were planted in eight plots, and the corresponding eight self-pollinated materials were planted in adjacent plots. During the flowering period, the fertility and nectary conditions were observed, and it was found that the sterility rate in the offspring population of the NCMS×Zengchengjianyecaixin-5-1-1 (abbreviated as NCMS1) and NCMS×Zengchengjianyecaixin-5-1-4 (abbreviated as NCMS4) combinations was >50%.

[0036] (3) Breeding of sterile lines and maintainer lines: In a group with a sterility rate of test cross progeny greater than 50%, backcrossing is performed between the sterile plants in the group and the corresponding sister plants. A group with a sterility rate of 100% is selected from the backcross progeny. At this time, the sterile plants in the group are self-pollinated with their corresponding sister plants, and the fertile sister plants are self-pollinated to obtain offspring to form the maintainer line. The sterile plants are test-crossed with the maintainer line to obtain a matching sterile line.

[0037] In the NCMS1 and NCMS4 populations, 8 sterile plants were selected, and the 8 flower branches were covered with sulfuric acid paper bags and numbered. At the same time, 8 plants were selected from the corresponding inbred lines and backcrossed with the 8 sterile plants one by one; similarly, the backcross seeds were planted in 8 plots, and the corresponding inbred lines were planted adjacent to each other. In the backcross offspring of the NCMS1 and NCMS4 populations, 100% sterile populations appeared, and the nectaries were normal. At this time, the sterile population and the corresponding male parent are the sterile line and the maintenance line, and this sterile material is numbered as NCMS-11.

[0038] like Figure 2 As shown, it is a plane photograph of the flower organs of the sterile plant (left) and the corresponding fertile plant (right) of Zengcheng sharp-leaved Chinese cabbage obtained in Example 1 of the present invention, as shown in FIG. Figure 3 As shown, these are side photographs of the floral organs of the sterile Zengcheng leafy Chinese cabbage plant (left) and the corresponding fertile plant (right) obtained in Example 1 of the present invention. It can be seen that the anthers of the sterile Zengcheng leafy Chinese cabbage plant are degenerate, while the pistil and nectary are normal.

[0039] Experimental Example 1 Identification of sterile source and study of its sterility mechanism Using the second and third generation high throughput sequencing technologies, the mitochondrial genomes of fertile and sterile plants of Zengcheng Chinese cabbage were sequenced, assembled and compared to determine the loci of male sterility. The results showed that the main differences between the two genomes were in atp6 and trnM -CAT two loci, outside this region, the similarity between the two is 100%. Further comparison results show that the sterility source of Zengcheng Chinese cabbage sterility line is similar to the existing Polima sterility source sterility gene loci.

[0040] Using primers at the sterile site ORF244 and its vicinity, through PCR amplification and PCR product sequencing, it was found that the sterile site ORF244 could only be amplified in sterile plants but could not be cloned in fertile plants.

[0041] The results of high-throughput sequencing and PCR verification showed that the sterility source contained in the sterile plants of Zengcheng Chinese cabbage was the Polima sterility source, that is, the presence of ORF224, which led to the male sterility of Zengcheng Chinese cabbage.

[0042] Theoretical analysis of the method for creating the cytoplasmic male sterile line of Chinese cabbage of the present invention: the genotype of the sterile plant in step (1) is S (msms); theoretically, the genotype of the sister plant in step (2) may be N (MsMs), N (Msms) and N (msms), accounting for 1 / 4, 1 / 2 and 1 / 4 respectively. In the test cross offspring, the fertile genotype is S (Msms) and the sterile genotype is S (msms). The test cross offspring does not have a 100% sterile population, but a fertile population and a sterile population, indicating that the genotype of the sister plant may be N (MsMs) or N (Msms); the genotype of the 100% sterile population in step (3) is S (msms), so the genotype of the corresponding inbred line is N (msms), thereby obtaining a sterile line and a maintainer line.

[0043] Application Example 1 Preparation of hybrid first generation combination The sterile line NCMS-11 obtained in Example 1 of the present invention and the stable inbred line were combined to breed hybrid first-generation varieties, as follows: (1) Use Zengcheng Jianye-5-7-2-3-4-1-3-4 (see the plant photo for details) Figure 5 ) and NCMS-11 to prepare a hybrid combination, the offspring (numbered 2023048, the plant photo of which is shown in Figure 7 ) is a late-maturing, rough, pointed-leafed Chinese cabbage that exhibits strong hybrid advantage, with obvious main stalks, a high stalk-to-leaf ratio, excellent traits, high quality, and high commercial value.

[0044] (2) Using early-maturing cabbage - 2-1-2-1-1 (see the plant photo for details) Figure 6 ) and NCMS-11 to prepare a hybrid combination, the offspring (numbered 2023051, the plant photo of which is shown in Figure 8 ) shows strong hybrid advantages, excellent traits, and high commercial value, especially filling the gap in cabbage stalk varieties that can be grown in high-altitude or low-temperature areas.

[0045] The above application examples show that the sterile line of the present invention has obvious advantages in preparing combinations.

[0046] Application Example 2: Transformation and fruiting of sterile lines 1. Transformation of sterile lines 1) NCMS-11 transformed into Teyouqing 70 Chinese cabbage By backcrossing with Teyouqing 70 Chinese cabbage, NCMS-11 was successfully transferred to Teyouqing 70 Chinese cabbage, and the sterile line NCMS-Teyouqing 70 Chinese cabbage-4-1-3-1 was obtained (see the plant photo for details). Fig. 9 ), numbered NCMS-70, the sterility rate of the sterile line is 100%, and the nectaries are normal.

[0047] 2) NCMS transferred to Beet Heart No. 31 By backcrossing No. 31 beet heart, NCMS was successfully transferred to No. 31 beet heart, and the sterile line NCMS-31 beet heart-22-28-4-15-2 was obtained, numbered NCMS-31. The sterility rate of the sterile line is 100%, and the nectaries are normal.

[0048] 3) Polima to Beet Heart No. 31 By backcrossing with beet heart No. 31, the cytoplasmic male sterility source Polima of rapeseed was successfully transferred to beet heart No. 31, and the sterile line was numbered PCMS-31.

[0049] The above indicates that the NCMS of the present invention, like Polima, can be stably transfected.

[0050] 2. Fruit setting rate The fruiting rate of NCMS-31, PCMS-31 and radish cytoplasmic male sterile ogura was observed. The fruiting rate per plant of NCMS-31 was significantly higher than that of PCMS-31 and ogura: the fruiting rate per plant of NCMS-31 was about 16.04, the fruiting rate per plant of PCMS-31 was 9.15, and the fruiting rate per plant of ogura was 7.28. The normal number of pods per plant of NCMS-31 was significantly higher than that of PCMS-31 (see Fig.10 and 11 ): The normal number of pods per plant of NCMS-31 is 490, and the number of defective pods is 203; the normal number of pods per plant of PCMS-31 is 130, and the number of defective pods is 118. The average per-acre seed yield of NCMS-31 is significantly higher than that of PCMS-31: the seed yield of NCMS-31 is about 30.45kg / acre, and the seed yield of PCMS-31 is about 9.12kg / acre.

[0051] Although the content of the present invention has been described in detail through the above preferred embodiments, it should be appreciated that the above description should not be considered as a limitation of the present invention. After reading the above content, it will be apparent to those skilled in the art that various modifications and substitutions of the present invention will occur. Therefore, the protection scope of the present invention should be limited by the appended claims.

Claims

1. A method for creating a cytoplasmic male sterile line of Chinese cabbage, characterized in that: The method comprises the following steps: (1) Self-pollination of Zengcheng Jianyecaixin revealed sterile plants, the anthers of which were degenerate, but the pistils and nectaries were normal; (2) Test crosses were conducted with sterile plants and sister plants from the Zengcheng Jianye Chinese cabbage inbred line to obtain test cross seeds, which were then planted to observe fertility and nectary conditions during the flowering period. The sterility rate of the test cross offspring was greater than 50%. (3) In the population with a sterility rate greater than 50% in the test cross progeny in step (2), backcrossing is performed between the sterile plants in the population and the corresponding sister plants, and a population with a sterility rate of 100% is screened out from the backcross progeny. The anthers in the population are degenerate, and the pistils and nectaries are normal. The sterile plants in the population are self-pollinated with their corresponding sister plants, and the fertile sister plants are self-pollinated to obtain progeny to form a maintainer line. The sterile plants are test-crossed with the maintainer line to obtain a matching sterile line.

2. The method for creating a cytoplasmic male sterile line of Chinese cabbage according to claim 1, characterized in that: In step (1), the Zengcheng sharp-leaved Chinese cabbage variety is planted, bagged before the flower buds open during the flowering period to prevent pollen cross-pollination, artificially assisted self-pollination is performed several times, and the seeds are threshed and harvested after they mature. In the next planting season, the self-pollinated first-generation seeds are planted and self-pollinated again to obtain the second-generation self-pollinated materials. In the next planting season, the second-generation self-pollinated materials are planted, and sterile plants with degenerated anthers and normal pistils and nectaries appear in the population.

3. The method for creating a cytoplasmic male sterile line of Chinese cabbage according to claim 1, characterized in that: In step (2), a number of sister plants are selected from the group of sterile plants and bagged with sulfuric acid paper, and the same number of flower branches as the sister plants are selected from the sterile plants and bagged, and the sterile branches of the sterile plants are test-crossed with the sister plants one by one. After the seeds mature, the test-cross seeds are harvested, and the group with a sterility rate greater than 50% is selected from the test-cross offspring.

4. The method for creating a cytoplasmic male sterile line of Chinese cabbage according to claim 1, characterized in that: In step (2), in the next planting season, several test cross seeds harvested from the maintenance of sterile plants are planted in several plots, and the corresponding several self-pollinated materials are planted in adjacent plots; the fertility and nectary conditions are observed during the flowering period, and if the sterility rate in the test cross offspring group is found to be >50%, several sterile plants are selected from the group, and several flower branches are bagged, and at the same time, several plants are selected from the corresponding inbred lines and backcrossed with several sterile plants one by one; the backcross seeds are planted in several plots, and the corresponding inbred lines are planted adjacent to each other, and a 100% sterile group appears in the backcross offspring of the group, and the nectary is normal, the sterile plants in the group are self-pollinated with their corresponding sister plants, the fertile sister plants are self-pollinated to obtain offspring to form a maintenance line, and the sterile plants are test-crossed with the maintenance line to obtain a matching sterile line.

5. The use of the method for creating a cytoplasmic male sterile line of Chinese cabbage according to any one of claims 1 to 4, characterized in that: The sterile line obtained by the method for creating the cauliflower cytoplasmic male sterile line as claimed in claim 1 and the stable inbred line are combined to select and breed the first-generation hybrid varieties.

6. The use according to claim 5, characterized in that: The stable inbred line was selected from Beet Heart No. 31.

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