A method for utilizing delay of flowering of a wild species of sugarcane with thin stalk to realize hybridization with a cultivated species

Abstract

The application discloses a method for delaying the flowering of a wild sugarcane variety to realize hybridization with a cultivated variety, which comprises the following steps: firstly, determining the germplasm of the wild sugarcane variety (commonly known as the Qushenmi variety), and determining the type of the selected germplasm according to the breeding target before utilization; secondly, delaying the cultivation of the selected Qushenmi variety, that is, taking the seed stalks from a seedling nursery in early June to mid-July to perform the delaying cultivation, and cutting off the main stem in early or middle September, and focusing on cultivating the tiller stems until the Qushenmi variety flowers; and finally, processing the field difficult-to-flower sugarcane parent plants in the parent generation, such as the tropical variety, the sugarcane breeding variety or the parent plant, cultivating the parent plants by means of barrel cultivation, starting to induce the flowering of the parent plants by means of artificial light period on about June 20, realizing the flowering period of the Qushenmi variety, performing the warm water emasculation treatment on the inflorescence of the female parent, and then performing the parent combination, so that the hybridization utilization of the excellent Qushenmi variety can be realized.

Description

Technical Field

[0001] This invention belongs to the field of agricultural planting technology, specifically relating to a method for delaying the flowering of wild sugarcane slender stems to achieve hybridization with cultivated varieties. Background Technology

[0002] Spontanm (also known as the slender-stemmed wild sugarcane) is a wild species within the genus *Saccharum*. The genus *Saccharum* includes cultivated species and wild species. Cultivated species include the tropical species *S. officinarum*, the Indian species *S. barberi*, and the Chinese species *S. sinense*. Wild species include the large-stemmed wild species *S. rodusfum*, the fleshy-spike wild species *S. edule*, and the slender-stemmed wild species. The slender-stemmed wild species is widely distributed, highly adaptable, and exhibits diverse ecological types. It possesses resistance to drought, poor soil, cold, flooding, disease, and insects, making it a major source of stress resistance in sugarcane breeding. It is the earliest utilized and most successfully bred wild species in sugarcane genetic breeding. Since the 1920s, in Indonesia, breeders have crossed Javanese sugarcane with tropical varieties to develop the "King of Sugarcane," POJ2878. In India, they have crossed Indian sugarcane with tropical and Indian varieties to breed world-renowned parent lines or varieties such as Co213, Co281, Co290, and Nco310, laying the genetic foundation for sugarcane breeding worldwide. Currently, the vast majority of cultivated varieties are descendants of POJ and CO. To further broaden the genetic basis of sugarcane breeding, break the long-standing practice of inbreeding, enhance the stress and disease resistance of varieties, and achieve a second breakthrough in sugarcane varietal development, countries around the world have actively researched and utilized sugarcane germplasm resources. Since the 1960s, the United States has used the Burmese variety *Gesellschaft* to breed early-maturing, high-sugar, and cold-resistant varieties such as CP65-357 and the main cultivated variety LCP85-384. In the 1970s, Australia used Burmese *Gesellschaft* to breed popular varieties such as Q138, Q170, and Q234. In the mid-to-late 20th century, Taiwan used Taiwan *Gesellschaft* to breed popular varieties such as ROC16, ROC23, and ROC24. Since its establishment in 1953, the Hainan Sugarcane Breeding Farm in my country has used Yacheng *Gesellschaft*, Lingshui *Gesellschaft*, and Yunnan *Gesellschaft* 75-2-11 to breed 32 new sugarcane varieties, including Yuetang 89-113, Tiancheng 16, and Guitang 24. Yunnan Province in my country has used Yunnan *Gesellschaft* to breed Yunzhe 99-155. The use of *Gesellschaft* varieties has promoted the development of sugarcane breeding. However, existing technologies have the following drawbacks: 1. The *Gesellus chinensis* population is rich in types, with multiple chromosome types and a wide distribution, and the germplasm used has not yet been used to breed breakthrough sugarcane varieties; 2. *Gesellus chinensis* flowers relatively early, mostly concentrated in September and October, making it difficult to encounter sugarcane varieties or parents that flower from December to January of the following year, thus hindering hybridization and making it difficult to obtain the planned ideal combination. Therefore, the discovery and utilization of superior *Gesellus chinensis* varieties has potential breeding value for improving sugarcane varieties. Summary of the Invention

[0003] To address the aforementioned technical problems, this invention presents a method for delaying flowering of wild sugarcane slender shoots to facilitate hybridization with cultivated varieties, specifically comprising the following steps:

[0004] S1: Collect and preserve the superior slender-stemmed wild species (Gnaphalium affine) of the Sugarcane genus from different ecotypes at home and abroad in the resource nursery;

[0005] S2: In the spring when the weather warms up in February or March, take single-bud seed stems with dense cuttings from the resource nursery and plant them in a bucket. Fill the bucket with soil, place the seed, fertilize, cover with soil, water, and cover with film. Remove the film after one month, weed, fertilize and apply pesticides, mound the soil, and water as needed, and manage normally.

[0006] S3: From early June to mid-July, take the single-bud seed stems of sugarcane with dense cuttings from S2 for delayed barrel planting. Plant them in barrels. When planting, they need to be disinfected by soaking in 2% quicklime water or 2% carbendazim solution. Fill the soil, place the seed, fertilize, cover with soil, water, and cover with film. After half a month, remove the film, weed, fertilize and apply pesticides, and water in time to maintain normal growth.

[0007] S4: In early to mid-September, in the S3 delayed-planting tank, the main stem branches and diseased or abnormally growing plants are cut off from the root. The main stems that have already developed spikes or produced spikes are encouraged to develop tillers. Appropriate fertilization, soil mounding, and timely watering are carried out. In November, the plants are moved into the greenhouse for warmth until they develop spikes and flower from December to January of the following year. The flowering period coincides with that of sugarcane production varieties or parent plants. Through crossbreeding, the sugarcane can be harvested more efficiently.

[0008] Furthermore, the method for delaying flowering during delayed barrel planting and delayed hand-cutting in S4 is as follows:

[0009] S4.1: First, determine the germplasm of the cut hand sugarcane. Then, carry out cultivation treatment on the selected cut hand sugarcane, that is, take seed stems from the seed collection nursery from early June to mid-July for delayed planting treatment; and cut off the main stem in early to mid-September to focus on cultivating tillers until delayed flowering.

[0010] S4.2: Treat the sugarcane parent that is difficult to flower in the field in the recurrent parent, cultivate it in barrel, and start artificial photoperiod to induce flowering in mid-to-late June to achieve the same flowering period as the female parent. The flower spikes are treated with warm water to kill the males, and then the two parents are bred to obtain the hybrid utilization of the superior female parent.

[0011] Furthermore, during the cultivation of the hand-cut dense tillers, 4-5 pot-grown tillers are retained during the treatment in September. During the cultivation of the tillers, they are cultivated under natural conditions in November. When the temperature drops to 13℃, they are moved to a 20℃ greenhouse for insulation to ensure normal heading and flowering in the later stage.

[0012] Furthermore, the photoperiod induction method in S4.2 is as follows: after fixing the light time at noon and treating for 120-150 days, the light is gradually reduced by 1 min / day, and then the light is fixed until flowering to form a good induction effect.

[0013] Furthermore, the method for killing male pollen in the female parent's flower spikes in S4.2 is as follows: when the sugarcane flower spikes emerge and the florets begin to open, soak the flower spikes in a 50℃ constant temperature water tank for 3-5 minutes to kill the pollen, prevent the female parent from self-pollinating and producing seeds, and ensure that a true hybrid offspring of the female and male parents is obtained.

[0014] The beneficial effects of this invention are:

[0015] This invention delays the flowering of *Gynostemma pentaphyllum* by delaying its planting in barrels and cultivating tillers, thus enabling hybridization with sugarcane varieties or parents and utilizing different superior *Gynostemma pentaphyllum* germplasm resources. Furthermore, this invention achieves a significant and stable effect in delaying flowering, ensuring planned hybridization with sugarcane varieties or parents. In addition, the delayed flowering of *Gynostemma pentaphyllum* produced by this invention results in well-developed, abundant, and full pollen, leading to a high fruit set rate when hybridized with sugarcane and yielding numerous seedlings. Detailed Implementation

[0016] Example 1

[0017] This invention discloses a method for delaying flowering in wild sugarcane slender shoots to achieve hybridization with cultivated varieties, comprising:

[0018] S1: Ruili RL2023-14, collected from Ruili, Yunnan, with a stem diameter of 2.1cm, is a type of extra-large stem with dense cutting.

[0019] S2: In the spring when the weather warms up in February or March, take the single-bud seed stem of Ruili RL2023-14 and plant it in a bucket. Fill the bucket with soil, place the seed, fertilize, cover with soil, water, and cover with film. Remove the film after one month, weed, fertilize and apply pesticides, mound the soil, and water as needed. Manage it normally. The normal treatment in this example is the conventional treatment technology for sugarcane, which is existing technology and will not be explained in detail here.

[0020] S3: From early June to mid-July, take the single-bud stem of Ruili RL2023-14 from S2 for delayed barrel planting. Plant in barrels. When planting, soak in 2% carbendazim solution for disinfection, fill with soil, place the seed, fertilize, cover with soil, water, and cover with film. After half a month, remove the film, weed, fertilize and apply pesticides, and water as needed to maintain normal growth.

[0021] S4: In early to mid-September, prune the main stem of the potted plant, retain the tillers, fertilize, mound soil around the plant, and cultivate the tillers. Under natural conditions in November, when the temperature drops to 13℃, move the plant to a 20℃ greenhouse for warm cultivation.

[0022] S5: On December 29, 2023, the hybrid combination Yunzhe 08-1609 × Ruili RL2023-14 was obtained. On January 2, 2024, an investigation revealed that there was still 1 plant in the booting stage and 1 plant in the heading stage.

[0023] The above-mentioned hybrid combination Yunzhe 08-1609 × Ruili RL2023-14 is to realize the hybrid utilization of Geshoumi Ruili RL2023-14 and sugarcane varieties.

[0024] After delayed planting, the Ruili RL2023-14 variety flowered on December 29, 2023, coinciding with the flowering period of Yunzhe 08-1609. At this time, the flower spikes of Yunzhe 08-1609 were soaked in a 50℃ constant temperature water tank for 4 minutes to kill the pollen and prevent Yunzhe 08-1609 from producing self-pollinating seeds as the female parent. Yunzhe 08-1609, after being sterilized by warm water, was used as the female parent and crossed with Ruili RL2023-14 to obtain the hybrid combination Yunzhe 08-1609 × Ruili RL2023-14. The results are shown in Table 1 below:

[0025] Table 1. Delayed flowering and fruiting after treatment with Mireille RL2023-14 (a type of citrus herb).

[0026]

[0027] Note: The flowering time is based on the time when the florets open after the sugarcane heads emerge.

[0028] The planting date for the Ruili RL2023-14 sugarcane variety was June 19, 2023, which was 86 days later than the control. After the main stem was pruned on September 10, 2023, and the tillers were retained, the tillers flowered on December 29, 2023, which was more than 58 days later than the control. Using Yunzhe 08-1609 as the female parent and Ruili RL2023-14 as the male parent, the hybrid combination Yunzhe 08-1609 × Ruili RL2023-14 was obtained, realizing the hybrid utilization of Ruili RL2023-14 and a superior sugarcane variety. However, the traditional method of utilizing Ruili RL2023-14 without treatment cannot obtain the hybrid combination Yunzhe 08-1609 × Ruili RL2023-14. Finally, by sowing hybrid seeds, 78 seedlings were obtained from the hybrid combination Yunzhe 08-1609 × Ruili RL2023-14. The results showed that the treated Ruili RL2023-14 had good pollen development and a good fruit set rate when hybridized with sugarcane.

[0029] Example 2

[0030] A method for delaying flowering in wild sugarcane slender shoots to facilitate hybridization with cultivated varieties includes:

[0031] S1: Yunnan Agricultural University 2015-83, collected from Myanmar and preserved in the Yunnan Agricultural University Resource Nursery, was introduced to the Ruili Resource Nursery in 2018. The preserved stem diameter is 1.27cm and the grain size is 16.1%. It is a germplasm resource of large stem, high yield and high sugar content of *Gesellschaft*.

[0032] S2: In the spring when the weather warms up in February or March, take the single-bud seed stems of Yunnan Agricultural University 2015-83 and plant them in a bucket. Fill the bucket with soil, place the seed, fertilize, cover with soil, water, and cover with film. Remove the film after one month, weed, fertilize and apply pesticides, mound the soil, and water as needed. Manage normally. The normal treatment in this example is the conventional treatment technology for sugarcane, which is existing technology and will not be explained in detail here.

[0033] S3: From early June to mid-July, take the single-bud stems of Yunnan Agricultural University 2015-83 from S2 for delayed barrel planting. Plant in barrels. When planting, soak in 2% carbendazim solution for disinfection, fill with soil, place the seed, fertilize, cover with soil, water, and cover with film. After half a month, remove the film, weed, fertilize and apply pesticides, and water as needed to maintain normal growth.

[0034] In early to mid-September, the main stem of S4 plants in pots was pruned, leaving the tillers. Fertilizer was applied, soil was added, and tillers were cultivated. Under natural conditions in November, when the temperature dropped to 13℃, the plants were moved to a 20℃ greenhouse for warm cultivation.

[0035] S5: On December 13, 2023, the hybrid combination Yunnan Agricultural University 2015-83 × Yunzhe 05-51 was obtained. Observation showed that the natural flowering time of Yunnan Agricultural University 2015-83 was relatively late and concentrated.

[0036] The above-mentioned hybrid combination, Yunnan Agricultural University 2015-83 × Yunzhe 05-51, is to realize the hybridization and utilization of Yunnan Agricultural University 2015-83 and sugarcane varieties.

[0037] After delayed planting, the Yunnan Agricultural University 2015-83 variety, known as "Geshoumi," flowered on December 13, 2023, coinciding with the flowering period of Yunzhe 05-51. At this time, the flower spikes of Yunnan Agricultural University 2015-83 were soaked in a 50℃ constant temperature water bath for 4 minutes to kill the pollen and prevent self-pollination and seed production as the female parent. Yunnan Agricultural University 2015-83, after being sterilized by warm water, was used as the female parent and crossed with Yunzhe 05-51 to obtain the hybrid combination Yunnan Agricultural University 2015-83 × Yunzhe 05-51. The results are shown in Table 2 below.

[0038] Table 2. Delayed flowering and fruiting after treatment with Yunnan Agricultural University 2015-83 (a type of herb).

[0039]

[0040] Note: The flowering time is based on the time when the florets open after the sugarcane heads emerge.

[0041] The planting date for the Yunnan Nongda 2015-83 sugarcane variety was July 17, 2023, which was 136 days later than the 2020 planting (control). After the main stem was pruned on September 15, 2023, and the tillers were retained, the tillers flowered on December 13, 2023, which was 15 days later than the 2020 planting (control). Using Yunnan Nongda 2015-83 as the female parent and Yunzhe 05-51 as the male parent, a hybrid combination Yunnan Nongda 2015-83 × Yunzhe 05-51 was obtained, realizing the hybrid utilization of Yunnan Nongda 2015-83 and a superior sugarcane variety. However, the traditional method of utilizing Yunnan Nongda 2015-83 without treatment cannot obtain the hybrid combination Yunnan Nongda 2015-83 × Yunzhe 05-51. Finally, by sowing hybrid seeds, 26 seedlings were obtained from the hybrid combination Yunnan Agricultural University 2015-83 × Yunzhe 05-51. The results showed that the treated Yunnan Agricultural University 2015-83 had good pollen development and could produce fruit when hybridized with sugarcane.

[0042] Example 3

[0043] A method for delaying flowering in wild sugarcane slender shoots to facilitate hybridization with cultivated varieties includes:

[0044] S1: Cut Hand Secret Vietnam No. 2, collected from Lao Cai, Vietnam, with a plant height of 285cm, stem diameter of 0.78cm, and a yield of 8.43%. It is a typical high-yield, high-sugar, humid and hot ecological type of Cut Hand Secret.

[0045] S2: In the spring when the weather warms up in February or March, take the single-bud stem of Vietnam No. 2 and plant it in a bucket. Fill the bucket with soil, place the seed, fertilize, cover with soil, water, and cover with film. Remove the film after one month, weed, fertilize and apply pesticides, mound the soil, and water as needed. Manage it normally. The normal treatment in this example is the conventional treatment technology for sugarcane, which is existing technology and will not be explained in detail here.

[0046] S3: From early June to mid-July, take the single-bud stems of Vietnam No. 2 from S2 for delayed barrel planting. Plant in barrels. When planting, soak in 2% carbendazim solution for disinfection, fill with soil, place the seed, fertilize, cover with soil, water, and cover with film. After half a month, remove the film, weed, fertilize and apply pesticides, and water as needed to maintain normal growth.

[0047] In early to mid-September, the main stem of S4 plants in pots was pruned, leaving the tillers. Fertilizer was applied, soil was added, and tillers were cultivated. Under natural conditions in November, when the temperature dropped to 13℃, the plants were moved to a 20℃ greenhouse for warm cultivation.

[0048] S5: On December 20, 2022, the hybrid combination Yunzhe 05-51 × Vietnam No. 2 was obtained.

[0049] The aforementioned hybrid combination Yunzhe 05-51 × Vietnam No. 2 is intended to achieve the hybridization and utilization of Geshoumi Vietnam No. 2 and sugarcane varieties.

[0050] After delayed planting, the Vietnam No. 2 cultivar, also known as Ge Shou Mi, flowered on December 20, 2022, coinciding with the flowering period of Yunzhe 05-51. At this time, the flower spikes of Yunzhe 05-51 were soaked in a 50℃ constant temperature water bath for 4 minutes to kill the pollen and prevent self-pollination and seed production as the female parent. Yunzhe 05-51, after undergoing warm water emasculation treatment, was used as the female parent and crossed with Vietnam No. 2 to obtain the hybrid combination Yunzhe 05-51 × Vietnam No. 2. The results are shown in Table 3 below.

[0051] Table 3. Delayed flowering and fruiting after treatment with *Gnaphalium affine* Vietnam No. 2.

[0052]

[0053] The planting date for the "Ge Shou Mi Vietnam 2" sugarcane variety was July 4, 2022, 123 days later than the control. After pruning the main stem and retaining the tillers on September 5, 2023, the tillers flowered on December 20, 2022, 54 days later than the control. Using Yunzhe 05-51 as the female parent and Vietnam 2 as the male parent, a hybrid combination Yunzhe 05-51 × Vietnam 2 was obtained, realizing the cross-fertilization of Ge Shou Mi Vietnam 2 with a superior sugarcane variety. Traditional, untreated Ge Shou Mi Vietnam 2 methods cannot achieve the hybrid combination Yunzhe 05-51 × Vietnam 2. Finally, by sowing hybrid seeds, 527 seedlings of the hybrid combination Yunzhe 05-51 × Vietnam 2 were obtained. The results showed that the treated Ge Shou Mi Vietnam 2 had excellent pollen development and a high seed setting rate when crossed with sugarcane.

[0054] The preferred embodiments of the present invention disclosed above are only for the purpose of illustrating the present invention. The preferred embodiments do not describe all the details in detail, nor do they limit the invention to the specific implementation described.

Claims

1. A method for delaying flowering of wild sugarcane slender shoots to achieve hybridization with cultivated varieties, characterized in that, Specifically, the following steps are included: S1: Collect and preserve high-quality sugarcane cultivars of different ecotypes from home and abroad in the resource nursery; S2: In the spring when the weather warms up in February or March, take single-bud seed stems with dense cuttings from the resource nursery and plant them in a bucket. Fill the bucket with soil, place the seed, fertilize, cover with soil, water, and cover with film. Remove the film after one month, weed, fertilize and apply pesticides, mound the soil, and water as needed, and manage normally. S3: From early June to mid-July, take the single-bud seed stems of sugarcane with dense cuttings from S2 for delayed barrel planting. Plant them in barrels. When planting, they need to be disinfected by soaking in 2% quicklime water or 2% carbendazim solution. Fill the soil, place the seed, fertilize, cover with soil, water, and cover with film. After half a month, remove the film, weed, fertilize and apply pesticides, and water in time to maintain normal growth. S4: In early to mid-September, in the S3 delayed-planting tank, the main stem branches and diseased or abnormally growing plants are cut off from the root. The main stems that have already developed spikes or produced spikes are encouraged to develop tillers. Appropriate fertilization, soil mounding, and timely watering are carried out. In November, the plants are moved into the greenhouse for warmth until they develop spikes and flower from December to January of the following year. The flowering period coincides with that of sugarcane production varieties or parent plants. Through crossbreeding, the sugarcane can be harvested more efficiently. In S3, S3.1: First, determine the germplasm of the cut hand sugarcane. Then, carry out cultivation treatment on the selected cut hand sugarcane, that is, take seed stems from the seed collection nursery from early June to mid-July for delayed planting treatment; and cut off the main stem in early to mid-September to focus on cultivating tillers until delayed flowering. S3.2: Treat the sugarcane parent that is difficult to flower in the field in the recurrent parent line, and start artificial photoperiod to induce flowering in mid-to-late June through barrel cultivation, so as to achieve the same flowering period as the female parent. The flower spikes are treated with warm water to kill the males, and then the two parents are bred to obtain the hybrid utilization of the superior female parent.

2. The method for utilizing delayed flowering of wild sugarcane slender shoots to achieve hybridization with cultivated varieties according to claim 1, characterized in that, When cultivating the hand-cut dense tillers, retain 4-5 pot-grown tillers during the treatment in September. During the tiller cultivation process, cultivate under natural conditions in November. When the temperature drops to 13℃, move to a 20℃ greenhouse for insulation to ensure normal heading and flowering in the later stage.

3. The method for utilizing delayed flowering of wild sugarcane slender shoots to achieve hybridization with cultivated varieties according to claim 1, characterized in that, The photoperiod-induced method in S3.2 is as follows: after fixing the light time at noon and treating for 120-150 days, the light is gradually reduced by 1 min / day, and then the light is fixed until flowering to form a good induction effect.

4. The method for utilizing delayed flowering of wild sugarcane slender shoots to achieve hybridization with cultivated varieties according to claim 1, characterized in that, The method for killing male pollen in the female parent's flower spikes in S3.2 is as follows: after the sugarcane flower spikes emerge and the florets begin to open, soak the flower spikes in a 50℃ constant temperature water bath for 3-5 minutes to kill the pollen, prevent the female parent from self-pollinating and producing seeds, and ensure that a true hybrid offspring of the female and male parents is obtained.

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