High-efficiency propagation method of Eurai poplar 107 poplar

Abstract

The invention relates to the technical field of plant breeding, in particular to a high-efficiency breeding method of Europe and America poplar 107 poplar. The method comprises the following steps of: cutting by taking annual European and American poplar 107 branches as a material, inducing a large number of adventitious buds, cutting the adventitious buds into stem sections, cutting the stem sections in a micro cutting device, inducing rooted seedlings, and cutting the rooted seedlings into sections after the rooted seedlings grow high to continue cutting propagation. Compared with conventional hard wood cutting, the number of the seedlings cultivated through the method is remarkably increased and is 3n times that of the conventional hard wood cutting, the efficient propagation coefficient is achieved, the land space used for propagation is saved, technical support is provided for efficient popularization and utilization of the 107 poplar and research on genetic transformation of adventitious buds based on field callus regeneration, and the important application value is achieved.

Description

Technical Field

[0001] The present invention relates to the technical field of plant breeding, and particularly relates to an efficient propagation method for Populus×euramericana cv.‘Neva’. Background Art

[0002] Populus×euramericana cv.‘Neva’ belongs to the genus Populus of the family Salicaceae, and is an excellent clone selected from the clones of “Populus deltoides×Populus nigra” introduced from Italy. Populus×euramericana cv.‘Neva’ is suitable for a wide range of planting areas and is widely distributed in the Huanghuai and Haihe River basins. Currently, it is widely planted in the northeast and northwest regions. It has various uses, significant characteristics of fast growth and high yield, cold resistance, good quality and high density, straight trunks, beautiful trunk shapes, small lateral branches and dense branches and leaves, strong wind and pest resistance, and easy propagation and high survival rate. Moreover, Populus×euramericana cv.‘Neva’ belongs to a plant with strong sprouting ability and can be used to cultivate seedlings on a large scale through asexual propagation methods such as cutting, grafting, layering, and tissue culture, providing a basis for the construction of an in vitro regeneration system for poplars. Compared with the method of regenerating adventitious buds through tissue culture, the adventitious buds regenerated from callus in the field have strong adaptability. It not only omits multiple links such as rooting, acclimatization, transplanting to the field, and sterilization after inducing adventitious buds through tissue culture, but also can directly obtain cuttings and be directly applied to production in one step. In addition, the adventitious buds regenerated from cuttings not only have strong adaptability but also can grow in clusters, greatly improving the propagation efficiency. In addition, there are still problems in the asexual propagation of Populus×euramericana cv.‘Neva’ in the prior art, such as low efficiency and large land space occupation. Therefore, establishing an efficient propagation method for Populus×euramericana cv.‘Neva’ is of great significance for its efficient promotion and utilization and genetic transformation research based on adventitious bud regeneration from callus in the field. Summary of the Invention

[0003] In view of the above technical problems, the present invention provides an efficient asexual propagation method using one-year-old branches of Populus×euramericana cv.‘Neva’ as cuttings in combination with a micro-cuttage device, which has a high propagation coefficient and occupies less land space, providing technical support for the efficient promotion and utilization of Populus×euramericana cv.‘Neva’ and genetic transformation research based on adventitious bud regeneration from callus in the field.

[0004] In order to achieve the above-mentioned invention purpose, the present invention provides the following technical solutions:

[0005] The present invention provides an efficient propagation method for Populus×euramericana cv.‘Neva’, comprising the following steps:

[0006] (1) Preparation of materials before propagation

[0007] Collect one-year-old ungerminated Populus×euramericana cv.‘Neva’ as cuttings; soak the top of the cuttings with a solution containing cytokinin, and seal with wax for standby;

[0008] (2) Cutting and cultivation

[0009] Insert the cuttings processed in step (1) into flower pots containing culture substrates; after dark cultivation for 8 - 12 days, place them under light for cultivation for 26 - 30 days to obtain adventitious buds;

[0010] (3) Adventitious bud cutting

[0011] When the adventitious buds obtained in step (2) grow to 2 - 3 cm, cut them from the cuttings and insert them into a container filled with culture substrate, and after cultivation for 1 - 2 weeks, obtain adventitious buds with roots;

[0012] (4) Repeated cutting of adventitious buds

[0013] Continue to cultivate the adventitious buds with roots until the plant height reaches 8 - 9 cm, cut them and cut them into stem segments of 2 - 3 cm and re - insert them into a container filled with culture substrate, and the cultivation method is the same as in step (3), and after continuing to cultivate for 1 - 2 weeks, obtain rooted seedlings;

[0014] (5) Transplanting

[0015] Continue to cultivate the rooted seedlings until the plant height reaches 6 - 8 cm, and then transplant them to the field.

[0016] Preferably, in step (1), the length of the cuttings is 12 - 16 cm and the thickness is 0.7 - 2.5 cm.

[0017] Preferably, in step (1), the solution containing cytokinin contains 0.2 - 0.8 mg / L cytokinin and 0.05 - 0.15 mg / L NAA.

[0018] Preferably, in step (1), the soaking time is 12 - 17 min.

[0019] Preferably, in step (2), the dark cultivation temperature is 25℃ - 26℃ and the humidity is 45% - 65%.

[0020] Preferably, in step (2), the temperature for cultivation under light is 25℃ - 26℃ and the humidity is 45% - 65%.

[0021] Preferably, the culture substrates in steps (2), (3) and (4) are all sandy soil, native soil and vermiculite, and the mass ratio is 4 - 6:0.8 - 1.2:0.8 - 1.2.

[0022] Preferably, the cultivation temperature in step (3) is 25℃ - 26℃ and the humidity is 45% - 65%.

[0023] Preferably, the containers filled with culture substrate in steps (3) and (4) are both micro - cutting devices.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] In the present invention, taking the branches of Populus×euramericana cv. 'Neva' in the first year as materials for cutting, after inducing a large number of adventitious buds, they are cut into stem segments and cut in a micro-cuttage device to induce rooting seedlings. After the rooting seedlings grow taller, they can also be cut into segments for continuous cuttage propagation. Compared with conventional hardwood cuttage, the number of seedlings cultivated by this method is significantly increased, which is 3 n times (n is the number of repeated cuttings). The present invention provides an efficient asexual propagation method using the first-year branches of Populus×euramericana cv. 'Neva' as cuttings in combination with a micro-cuttage device, which has a high propagation coefficient, saves the land space used for propagation, provides technical support for the efficient promotion and utilization of Populus×euramericana cv. 'Neva' and the genetic transformation research of adventitious bud regeneration based on callus in the field, and has important application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only the embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can also be obtained according to the provided drawings.

[0027] Figure 1 It is a state diagram of the branches of Populus×euramericana cv. 'Neva' cut into flower pots;

[0028] Figure 2 It is a state diagram of callus growing from the cut of the branches of Populus×euramericana cv. 'Neva'; where Figure (A) is the overall state of the flower pot, and Figure (B) is the state of the callus of a single branch;

[0029] Figure 3 It is a state diagram of adventitious buds growing from the cut of the branches of Populus×euramericana cv. 'Neva'; where Figure (A) is the overall state of the flower pot, Figure (B) is the state of adventitious buds of a single branch; Figure (C) is the state of adventitious buds when they grow to about 2 cm; (D) is the state of adventitious buds when they grow to about 3 cm;

[0030] Figure 4 It is a state diagram of the cuttage culture of adventitious buds on a micro-cuttage device;

[0031] Figure 5 It is a state diagram of adventitious bud rooting; DETAILED DESCRIPTION OF THE EMBODIMENTS

[0032] The following will describe in detail the technical solutions provided by the present invention in combination with the embodiments, but they cannot be understood as limiting the protection scope of the present invention.

[0033] In the following experimental methods of the examples, unless otherwise specified, they are all conventional methods. The materials, reagents, etc. used in the following examples can all be obtained from commercial sources unless otherwise specified. The micro-cuttage device used in the following examples is derived from ZL202321141212.2.

[0034] Example 1

[0035] An efficient propagation method for Populus × euramericana cv. 'Neva' is as follows:

[0036] (1) Collection and treatment of Populus × euramericana cv. 'Neva' branches

[0037] On March 7, 2023, before the germination of Populus × euramericana cv. 'Neva' in spring, in the Yanshan nursery in Cangzhou, Hebei Province, cut the one-year-old cuttings of Populus × euramericana cv. 'Neva' from the base. None of them had germinated. Remove the small branches and keep the cuttings about 2 m long. Cut the cuttings into three parts and soak them in water at the base in the artificial climate chamber of Hebei Agricultural University for 2 days.

[0038] (2) Preparation of culture medium and cuttings

[0039] Culture medium: Sandy soil: Original soil: Vermiculite (5:1:1);

[0040] Preparation of cuttings: Cut the soaked Populus × euramericana cv. 'Neva' branches into cuttings 15 cm long and 1.5 cm thick, keep the cross-section smooth and the integrity of the cambium.

[0041] (3) Treatment of cuttings

[0042] Immerse the top cross-section of the cuttings in a hormone solution containing 6-BA 0.5 mg / L + NAA 0.1 mg / L for 15 min, and then seal them with wax.

[0043] (4) Cuttage and culture

[0044] Mix the culture medium thoroughly and fill it into flower pots with an inner diameter of 15×15×10, ensure that the amount of culture medium in each flower pot is the same, insert 8 treated cuttings in each flower pot, keep the cutting depth of the cuttings consistent, carry out dark treatment for 10 days, the culture temperature is 25°C, and the humidity is 50% (see Figure 1 ). Then place them in the artificial climate chamber for light culture, the culture temperature is 25°C, and the humidity is 50%. Culture for a total of 28 days, and callus (see Figure 2 ) and adventitious buds (see Figure 3 ) are induced. On average, each cutting produces (43 + 81 + 74) / 3 = 66 buds.

[0045] (5) Cuttage of adventitious buds

[0046] When the adventitious buds growing on the cuttings reach 3 cm in length, cut them off from the cuttings and insert them into a micro-cutter filled with culture substrate. Appropriately prune the cuttings with more leaves and retain 3 leaves, and culture them in an artificial climate chamber (see Figure 4 ). The culture temperature is 25 °C and the humidity is 50%. Cover with a plastic film at the initial stage and pay attention to maintaining the humidity, and remove it after 1 week. Then carry out normal watering management. After rooting in 2 weeks, continue to culture in the artificial climate chamber.

[0047] (6) Repeated cutting of adventitious buds

[0048] After 60 days, when the plant height of the plants in the micro-cutter grows to 9 cm after rooting, cut them off, cut them into stem segments of 3 cm, and re-insert them into the micro-cutter and continue to culture for 2 weeks. The culture method is the same as in step (5). By mid-June, it can be cut again, and finally obtain rooted seedlings (see Figure 5 ).

[0049] (7) Transplanting rooted seedlings to the field

[0050] When the rooted seedlings grow to 7 cm, they can be transplanted to the field and normal water and fertilizer management can be carried out.

[0051] (8) Calculation of propagation coefficient

[0052] For 1 one-year-old Populus × euramericana cv. 'Neva' plant, the number of rooted seedlings of Populus × euramericana cv. 'Neva' that can be propagated by this propagation method is expressed by the propagation coefficient. The formula is 200 / 15×the average number of adventitious buds germinated per cutting×(9 / 3) n , after calculation, the propagation coefficient of Example 1 of the present invention = 200 / 15×66×(9 / 3) 1 = 2640.

[0053] The above is only the preferred embodiment of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. An efficient propagation method for Populus europaeus 107, characterized in that: The following steps are involved: (1) Preparation of materials before breeding One-year-old ungerminated Populus occidentalis 107 was collected as cuttings; Soak the top of the cuttings in a solution containing cytokinin and seal with wax for later use; (2) Cutting and cultivation The cuttings treated in step (1) are inserted into a flower pot containing a culture medium; after dark culture for 8 to 12 days, they are placed under light for culture for 26 to 30 days to obtain adventitious buds; (3) Adventitious bud cuttings When the adventitious buds obtained in step (2) grow to 2 to 3 cm, they are cut off from the cuttings and inserted into a container containing a culture medium. After culturing for 1 to 2 weeks, rooted adventitious buds are obtained; (4) Repeated cuttings of adventitious buds The rooted adventitious buds are further cultured until the plant height reaches 8-9 cm, and then cut into 2-3 cm long stem segments and re-inserted into a container containing a culture medium. The culture method is the same as step (3), and rooted seedlings are obtained after continuing to culture for 1-2 weeks. (5) Transplantation Continue to cultivate the rooted seedlings until they are 6 to 8 cm tall and then transplant them to the field.

2. The efficient breeding method according to claim 1, characterized in that: In the step (1), the length of the cuttings is 12 to 16 cm, and the thickness is 0.7 to 2.5 cm.

3. The efficient breeding method according to claim 1, characterized in that: The cytokinin-containing solution in step (1) contains 0.2-0.8 mg / L cytokinin and 0.05-0.15 mg / L NAA.

4. The efficient breeding method according to claim 1, characterized in that: The soaking time in step (1) is 12 to 17 minutes.

5. The efficient breeding method according to claim 1, characterized in that: In the step (2), the dark culture temperature is 25° C. to 26° C., and the humidity is 45% to 65%.

6. The efficient breeding method according to claim 1, characterized in that: The temperature of the culture under light in step (2) is 25° C. to 26° C., and the humidity is 45% to 65%.

7. The efficient breeding method according to claim 1, characterized in that: The culture medium in step (2), step (3) and step (4) is sand, original soil and vermiculite, and the mass ratio is 4-6:0.8-1.2:0.8-1.

2.

8. The efficient breeding method according to claim 1, characterized in that: The culture temperature of step (3) is 25° C. to 26° C., and the humidity is 45% to 65%.

9. The efficient breeding method according to claim 1, characterized in that: The containers containing the culture matrix in step (3) and step (4) are both micro-cutting containers.

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