Asparagus material with high saponin content and breeding method thereof

Abstract

The invention relates to the technical field of asparagus cultivation, and provides a high-saponin-content asparagus material and a breeding method thereof, and the breeding method comprises the following steps: (1) taking a high-saponin-content diploid asparagus sibling female plant as a material, inducing to form a seedling, doubling by using colchicine, and creating a tetraploid asparagus sibling female parent; (2) performing anther culture to induce callus by taking diploid asparagus anther as a material, and doubling by colchicine to obtain a tetraploid supermale plant as a male parent; and (3) hybridizing the female parent in the step (1) with the male parent in the step (2) to obtain the tetraploid asparagus all-male new material with high saponin content. According to the asparagus material with high saponin content and the breeding method thereof, the saponin content in asparagus can be quickly and effectively increased.

Description

Technical Field

[0001] The invention relates to the technical field of asparagus cultivation, and in particular to an asparagus material with high saponin content and a breeding method thereof. Background Art

[0002] asparagus( Asparagus officinalis L.), belonging to the genus Asparagus, is a functional vegetable with high nutritional and medicinal value. Asparagus is rich in saponins, flavonoids, polysaccharides and other active substances, thus having good antioxidant, anti-tumor, and immunity-enhancing effects. Asparagus's closest relative, Asparagus officinalis, is a traditional Chinese medicine, and steroidal saponins are one of its main active ingredients.

[0003] Saponin is an important terpene compound with a complex chemical structure. It is named because its aqueous solution can produce soap-like foam after shaking. According to the structure of sapogenin generated after saponin hydrolysis, it can be divided into two categories: triterpenoidal saponins and steroidal saponins. Steroidal saponins are mainly found in asparagus and its related species, and have multiple biological functions. Studies have shown that they have the effects of inhibiting tumors, promoting blood circulation and anti-inflammation, and anti-oxidation.

[0004] Studies have shown that after a diploid plant doubles to become a polyploid, the increase in chromosome number, additional genome interactions and genetic changes often make the polyploid plant's traits superior to those of a diploid plant. Polyploidy is beneficial to the increase in the content of metabolites of some medicinal plants, thereby increasing the effective ingredients of the medicinal plants.

[0005] Asparagus is a dioecious plant, with male plants producing more than female plants, and the content of flavonoids, soluble protein, and iron in tender stems being higher than that in female plants. Therefore, breeding all-male tetraploid varieties can not only effectively increase asparagus yields but also increase the content of active ingredients.

[0006] There are no reports on new tetraploid all-male asparagus varieties with high saponin content and their breeding methods. The main technical problems are as follows: 1. The existing methods for doubling asparagus chromosomes mainly include treating asparagus seeds with colchicine and treating asparagus sprouts or stem tips with different mutagens, but the doubling efficiency is low.

[0007] 2. The existing breeding methods have not introduced the excellent genes of the medicinal plant Asparagus cochinchinensis, a close relative of asparagus, and have not changed the saponin synthesis genes from the genetic material.

[0008] 3. There is a lack of hybridization technology between tetraploid asparagus and closely related species. Summary of the invention

[0009] The object of the present invention is to provide an asparagus material with high saponin content and a breeding method thereof, which can quickly and effectively increase the saponin content in asparagus.

[0010] The present invention provides a method for breeding asparagus materials with high saponin content, comprising the following steps: (1) Using high-saponin-content diploid female plants of asparagus relatives as materials, inducing seedling formation, doubling with colchicine, and creating tetraploid asparagus relatives as female parents; (2) Using diploid asparagus anthers as materials, anther culture was performed to induce callus, and then colchicine was used to double the tetraploid super-male plants, which were used as the male parent; (3) hybridizing the female parent in step (1) with the male parent in step (2) to obtain a new tetraploid asparagus all-male variety with a high saponin content.

[0011] Furthermore, in step (1), the saponin content of a large number of diploid female plants of related species of asparagus is tested, and female plants of related species of asparagus with high saponin content are screened out. The tender stems of the female plants are used as materials to induce the formation of seedlings, which are doubled with colchicine to create tetraploid female plants of related species of asparagus.

[0012] Furthermore, in step (2), anthers of diploid male asparagus plants are used as materials for anther culture, which is doubled with colchicine, and tetraploid super male plants are obtained by flow cytometry and chromosome pressing. The tetraploid super male asparagus plants finally obtained are used as the male parent.

[0013] Furthermore, in step (1), female plants of asparagus related species with high saponin content are screened out, and the tender stems of the female plants are used as materials to induce the formation of seedlings, which are doubled with colchicine to create tetraploid asparagus related species mother plants.

[0014] Furthermore, in step (2), the anther size of the male asparagus plant with high saponin content is 2.0-2.5 mm, and the microspore development is in the uninucleate marginal stage.

[0015] Furthermore, in steps (1) and (2), the tetraploid parents were obtained by treating callus with 1.0-3.0 g / L colchicine.

[0016] The present invention also provides asparagus materials prepared by the above-mentioned asparagus material breeding method with high saponin content.

[0017] The technical solution of the present invention has at least the following advantages and beneficial effects: 1. Through anther culture technology, the anthers of asparagus in the mononuclear marginal stage are used as materials to induce callus, and colchicine is used to double the callus. The doubling efficiency is high, and it is easier to obtain tetraploid asparagus super male plants.

[0018] 2. By introducing excellent genes from closely related species of the medicinal plant Asparagus, the saponin synthesis genes are changed genetically, thereby increasing the saponin content in new asparagus varieties.

[0019] 3. By screening a large number of female asparagus plants with high saponin content, inducing the formation of seedlings, and doubling them with colchicine, tetraploid female asparagus plants were obtained.

[0020] 4. Through the hybridization of a large number of tetraploid asparagus super male plants and asparagus female plants, tetraploid asparagus super male plants with high affinity and high saponin content are screened out. DETAILED DESCRIPTION

[0021] The present invention is further described below in conjunction with examples, but the embodiments of the present invention are not limited thereto. The experimental methods used in the following examples are conventional methods unless otherwise specified.

[0022] Example 1 Effect of colchicine on the doubling effect of seedlings The tender stems of female plants of asparagus relatives with high saponin content were used as explants. The induction medium for seedlings was MS+6-BA1.0 mg / L+IAA 0.5 mg / L+KT 0.5 mg / L (6-BA is 6-benzylaminoadenine, IAA is indoleacetic acid, and KT is kinetin). The concentration and treatment time of colchicine were used as induction factors, and multiple gradient orthogonal experiments were designed to study their effects on the induction rate of asparagus tetraploid. The seedlings were treated as follows, with colchicine concentrations of 1, 3, and 6 g / L, and the soaking time of seedlings was 3h, 6h, and 24h. Four seedlings were inoculated in each bottle of each material, and each treatment was repeated 3 times. After different treatments, all materials were inoculated on the proliferation medium, MS+6-BA 0.6 mg / L +IAA 0.3 mg / L. Other culture conditions were the same. Tetraploids were identified by flow cytometry and chromosome counting.

[0023] Table 1 The induction rate of tetraploidy in seedlings of asparagus related species by different concentrations of colchicine

[0024] The doubling effect of colchicine on asparagus seedlings under tissue culture conditions is shown in Table 1. The variation index is based on the obvious thickening of the plant as a rough standard, and the chromosome identification of the tender stems of polyploid materials is used as the identification standard. The results show that the effect of colchicine concentration on the doubling of asparagus seedlings is greater than the treatment time. There was no dead seedling at a concentration of 1 g / L colchicine, but the doubling rate was not high; the number of dead seedlings was the same at different treatment times at a concentration of 3 g / L colchicine, and the doubling rate was the highest when the treatment was 6h; at a concentration of 6 g / L colchicine, the germination of tender stems of seedlings was inhibited, resulting in a significant reduction in the number of treated plants and a significant increase in the number of dead seedlings. Therefore, the best concentration of colchicine for the doubling effect of asparagus seedlings under tissue culture conditions is 3 g / L, 6h.

[0025] Example 2 Analysis of saponin content of different multiples of asparagus A female diploid asparagus relative with high saponin content was used as the female parent and a male diploid asparagus was used as the male parent to prepare a hybrid combination named ZJ1; a tetraploid female plant obtained by doubling a female diploid asparagus relative with high saponin content was used as the female parent and a tetraploid super male plant created by anther culture was used as the male parent to prepare a hybrid combination named ZJ2. The saponin content of the three asparagus materials was detected by RP-HPLC using the common asparagus variety as the control (CK), and the detection sites included asparagus roots, tender stems, and mother stems.

[0026] Table 2 Saponin content in asparagus of each group

[0027] From the data in Table 2, it can be seen that the tetraploid asparagus all-male combination obtained by the cultivation method of the present application has the highest saponin content, wherein the content of asparagus roots is 17.91% higher than that of ordinary asparagus, the content of tender stems is 28.20% higher, and the content of mother stems is 57.14% higher.

[0028] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for breeding asparagus materials with high saponin content, characterized in that: The steps include: (1) Using high-saponin-content diploid female plants of asparagus relatives as materials, inducing seedling formation, doubling with colchicine, and creating tetraploid asparagus relatives as female parents; (2) Using diploid asparagus anthers as materials, anther culture was performed to induce callus, and then colchicine was used to double the tetraploid super-male plants, which were used as the male parent; (3) hybridizing the female parent in step (1) with the male parent in step (2) to obtain a tetraploid all-male asparagus new material with a high saponin content.

2. The method for breeding asparagus materials with high saponin content according to claim 1, characterized in that: In step (1), the saponin content of the diploid asparagus related species female plants is tested, and the asparagus related species female plants with high saponin content are screened out. The tender stems of the female plants are used as materials to induce the formation of seedlings, which are doubled with colchicine to create tetraploid asparagus related species mother plants.

3. The method for breeding asparagus materials with high saponin content according to claim 1, characterized in that: In step (2), anthers of diploid male asparagus plants are used as materials for anther culture, which is then doubled with colchicine, and tetraploid super male plants are obtained by flow cytometry and chromosome pressing. The tetraploid super male asparagus plants finally obtained are used as the male parent.

4. The method for breeding asparagus materials with high saponin content according to claim 1, characterized in that: Step (1) screens out female plants of asparagus related species with high saponin content, uses the tender stems of the female plants as materials, induces the formation of seedlings, doubles them with colchicine, and creates tetraploid asparagus related species mother plants.

5. The method for breeding asparagus materials with high saponin content according to claim 1, characterized in that: Step (2) uses anthers of male asparagus plants with high saponin content, the size of which is 2.0-2.5 mm, and the anthers of which the microspore development is in the mononuclear marginal stage.

6. The method for breeding asparagus materials with high saponin content according to claim 1, characterized in that: In steps (1) and (2), tetraploid parents were obtained by treating callus with 1.0-3.0 g / L colchicine.

7. Asparagus material prepared by the asparagus material breeding method with high saponin content according to any one of claims 1 to 6.

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