CN109006488A - A kind of red luxuriant pineapple breeding method based on using buds to propagate buds - Google Patents

Abstract

The invention discloses a method for cultivating red bud pineapple based on bud propagation, using tissue cultured whole green plants as materials, collecting them from calluses, transplanting them into medium for rooting and cultivating strong seedlings, and cutting them at the top of stems After the top treatment, move the strong seedlings into a dark environment for 40 days of stem elongation culture, and then transfer the plants with a stem length ≥ 3cm to the light condition to induce the germination of lateral buds, and then transfer them in sections to the medium that is conducive to bud germination and rooting. Subculture, and finally obtain regenerated plants with the same traits as the mother plant. The present invention obtains complete regenerated plants through in vitro culture, overcomes the disadvantages of high mutation rate in the organogenesis method of red bud pineapple tissue culture after callus redifferentiation process in the past, and ensures the regenerated plant and the mother plant. consistency.

Description

A kind of red bud pineapple cultivation method based on bud multiplication

technical field

The invention belongs to the technical field of red bud pineapple cultivation, and in particular relates to a red bud pineapple cultivation method based on bud propagation.

Background technique

In the prior art, red bud pineapple is used as material. During the process of establishing the in vitro culture and propagation system, the callus redifferentiation pathway will be used. The variation traits of adventitious buds are as high as 99%. The variation traits are many and irregular, and the variation type is mainly whole. Green plants, all-white plants, and chimeric plants; the proportions of these three plants are about 43% of all-green plants, 38% of all-white plants, and 19% of chimeric plants, and the chimeric traits are very different (Fig. 1). Therefore, in order to obtain the red bud pineapple chimera variety, it is not advisable to propagate through the approach of callus regeneration adventitious buds.

Therefore be necessary to provide a kind of new red bud pineapple cultivation method.

Contents of the invention

In view of this, the present invention provides a method for cultivating red bud pineapple based on bud propagation.

In order to solve the above-mentioned technical problems, the present invention discloses a method for cultivating red-bud pineapple based on bud propagation, using tissue-cultured whole-green plants as materials, collecting them from calluses, and transplanting them into culture medium to take root and strengthen seedlings Cultivate, after cutting the top of the stem, move the strong seedlings into a dark environment and undergo 40 days of stem elongation culture, transfer the plants with a stem length ≥ 3cm to the light condition for cultivation to induce lateral bud germination, and then transfer them in sections to facilitate bud germination and Subculture in the rooting medium, and finally obtain regenerated plants with the same traits as the mother plant.

Optionally, include the following steps:

Step 1, selection of material: collection takes pineapple stem section as explant, and the regenerated plant obtained by tissue culture is used as material; small plant is cut from the callus, transferred vertically to MS medium, strong seedling Cultivate; when the seedling grows to 3cm-4cm, the stem grows to 0.5cm, and the number of roots is 2-4, it is set aside;

Step 2, top cutting treatment: the plants cultivated by strong seedlings are cut off at the top of the stem under the ultra-clean table to eliminate the top advantage of the plant, and then transferred to MS medium for cultivation;

Step 3. Dark culture induces internode elongation: the plants after the top cutting of strong seedlings are put into a carton box for dark culture; the internode elongation length of the plant after dark culture is 1cm-4cm;

Step 4, light culture: after the end of the dark period, remove the plants with stem elongation ≥ 3 cm out of the carton, and induce the bud germination of the plants that have been in the dark environment for a long time under light conditions;

Step 5, segmented subculture: Cut the plants induced by bud germination into small sections with 2-3 nodes, which are respectively terminal bud segment, middle segment, and rooted base segment, and vertically inoculate them on the medium respectively to induce axillary buds germination;

Step 6. Formation of regenerated plants: after 1.5 months of segmental subculture, the axillary buds on the nodes will germinate, and will start to take root at the base after 2 months, and form a complete plant after 3-4 months;

Step 7, plant transplanting: the complete plant is gently cut off from the mother plant, and transplanted onto MS medium or directly into the substrate to complete plant transplantation.

Optionally, the plantlets in step 1 are healthy plants with 5-8 leaves.

Optionally, the condition for cultivating strong seedlings in the step 1 is to cultivate strong seedlings for 28 days under the condition of 25±2° C. and 12 h/d light.

Optionally, the culture condition in the MS medium in the step 2 is: culture at 25±2° C. and light for 12 h/d for one week.

Optionally, the dark culture conditions in step 3 are: culture temperature 25±2° C.; culture time 40 d, during which the culture is taken out for 1 h every 10 d.

Optionally, the condition for inducing bud germination in step 4 is light culture, specifically: culture at 25±2° C. and 12 h/d light for 15 days.

Optionally, the subculture medium in step 5 is MS+BA3mg/l+NAA2mg/l.

Optionally, the substrate in step 7 includes river sand and coconut peat at a mass ratio of 1:1.

Compared with prior art, the present invention can obtain and comprise following technical effect:

1) The present invention uses the whole green plant that comes from callus redifferentiation as material, adopts the tissue culture method, supplemented by suitable temperature, induces the elongation of the red bract pineapple internode by dark culture, and subcultures in sections on the optimized medium , Induce the germination of lateral buds, take root on the nodes to form a complete plant, and make the germination rate as high as 62% in the case of zero germination from normal culture.

2) The present invention obtains complete regenerated plants through in vitro culture, overcomes the shortcoming of high mutation rate in the organogenesis method of the red bud pineapple tissue culture process in the past through the callus redifferentiation process, and ensures that the regenerated plants are compatible with the mother. Consistency among strains.

Of course, implementing any product of the present invention does not necessarily need to achieve all the technical effects described above at the same time.

Description of drawings

The accompanying drawings described here are used to provide a further understanding of the present invention, and constitute a part of the present invention. The schematic embodiments of the present invention and their descriptions are used to explain the present invention, and do not constitute improper limitations to the present invention. In the attached picture:

Fig. 1 is the adventitious buds of different chimeric traits in the background technology of the present invention; wherein, A represents the all-green and all-white variant tissue culture seedlings with callus mainly differentiated, B represents irregular chimeras, and C represents chimeras with white leaf margins Composite (golden edge), D stands for chimera with green leaf edge and white stripes in the middle (golden heart), E stands for irregular chimera leaf, F stands for irregular chimera leaf;

Fig. 2 is the form figure of the strong green seedling in the selection of material of the present invention;

Fig. 3 is the morphological figure of the whole green seedling of stem elongation after dark cultivation of the present invention;

Fig. 4 is the comparison figure of strong seedlings before dark culture and leggy seedlings after dark culture of the present invention, wherein, A represents strong seedlings before dark culture, and B represents leggy seedlings after dark culture;

Fig. 5 is the morphological figure of long fixed buds at the leaf axils after the segmental subculture of the present invention, wherein, the long-term fixed buds at the leaf axils after the A segmental subculture, and the young and tender fixed buds during the B segmental subculture;

Figure 6 is a morphological diagram of the axillary buds on the nodes of the present invention.

Detailed ways

The implementation of the present invention will be described in detail below with examples, so as to fully understand and implement the implementation process of how the present invention uses technical means to solve technical problems and achieve technical effects.

Example 1

A kind of red bud pineapple cultivation method based on bud multiplication, comprises the following steps: the cultivation of strong seedling stage, top cutting treatment, the processing of dark cultivation, the processing of light cultivation, the cultivation of initial buds induced by segmental subculture, the cultivation of strong buds and Rooting culture process: Tissue cultured whole green plants are used as materials, and after being collected from the callus, transplanted into the culture medium to take root and cultivate strong seedlings, and after cutting the top of the stem, move the strong seedlings into a dark environment for 40 days Stem elongation culture, transfer the plants with stem length ≥ 3cm to culture under light conditions to induce the germination of lateral buds, and then transfer them into subculture medium that is conducive to bud germination and rooting, and finally obtain regenerated plants with the same traits as the mother plant. The specific operation steps are as follows:

Step 1, selection of materials: collecting the stem section of red bud pineapple as explants, and the regenerated plants obtained through tissue culture as materials; cutting healthy, small plants with about 5-8 leaves from the callus, vertically Transfer to MS medium, and cultivate strong seedlings for 28 days under the condition of 25±2°C and 12h/d light; when the seedlings are about 3cm-4cm long, the stem length is about 0.5cm, and the number of roots is 2-4 Bar, as shown in Figure 2, spare;

Step 2, Top cutting treatment: After the strong seedlings have been cultivated, the plants are cut at the top of the stem under the ultra-clean table to eliminate the top dominance of the plant, and transferred to MS medium, at 25±2°C, under the light of 12h/d cultured under the condition for one week;

Step 3. Dark culture induces internode elongation: the plants after the top cutting of strong seedlings are placed in a carton box for dark culture, the culture temperature is 25±2°C; the culture time is 40 days, and the plants are taken out for 1 hour every 10 days during the period. After dark culture, the internodes of the plants elongate obviously, and the elongation length is about 1cm-4cm (Fig. 3). The state at this time is conducive to the formation and development of axillary buds; see Figure 4. It can be seen from Figure 4 that after dark culture, the internodes of the plants are obviously elongated.

Table 1 Comparison of stem length in dark culture for 40 days

stem elongation 1cm 2cm 3cm 4cm Stem elongation ratio 4% 12% 36% 48%

Note: Stem elongation ratio = the number of plants with different stem elongation lengths after dark culture/the total number of plants with elongated stems after dark culture.

Step 4. Illumination cultivation: After the dark period ends, remove the plants with stem elongation ≥ 3cm out of the carton, so that the plants that have been in the dark environment for a long time are in a normal and healthy growth state, under the conditions of 25±2°C and 12h/d light Cultivate for 15 days to induce bud germination;

Step 5, segmented subculture: Cut the plants after light cultivation into small sections with 2-3 nodes, which are respectively terminal bud section, middle section, and rooted section, and inoculate them vertically in MS+BA3mg/ The germination of axillary buds was induced on the medium of l+NAA 2mg/l, and the germination rate of axillary buds at the rooted section was the highest, up to 68%.

Table 2 Subsection subculture axillary bud germination rate

Step 6. Formation of regenerated plants: After about 1.5 months of subculture in sections, the axillary buds on the nodes will germinate, and will start to take root at the base after 2 months, and form a complete plant after 3-4 months;

Table 3 Germination number and germination rate of regenerated plants

Step 7, plant transplanting: the complete plant is gently cut from the mother plant, transplanted to MS medium or directly transplanted into a substrate containing river sand and coconut peat at a mass ratio of 1:1, and then it can survive , Survival rate > 95%.

The above description shows and describes several preferred embodiments of the invention, but as previously stated, it should be understood that the invention is not limited to the form disclosed herein, and should not be regarded as excluding other embodiments, but can be used in various other embodiments. Combinations, modifications and circumstances, and can be modified within the scope of the inventive concept described herein, by the above teachings or by skill or knowledge in the relevant field. However, changes and changes made by those skilled in the art do not depart from the spirit and scope of the invention, and should be within the protection scope of the appended claims of the invention.

Claims (9)

1. a kind of red bud pineapple cultivation method based on bud multiplication, it is characterized in that, adopting tissue culture whole green plant is material, after gathering on callus, be transplanted in the substratum and take root strong seedling and cultivate, in stem After top-cutting treatment, move the strong seedlings into a dark environment and undergo 40 days of stem elongation culture, then transfer the plants with a stem length ≥ 3cm to culture under light conditions to induce the germination of side buds, and then move them into the medium that is conducive to bud germination and rooting in sections Subculture, and finally obtain regenerated plants with the same traits as the mother plant. 2. the red bud pineapple cultivation method according to claim 1, is characterized in that, comprises the following steps: Step 1, selection of material: collection takes pineapple stem section as explant, and the regenerated plant obtained by tissue culture is used as material; small plant is cut from the callus, transferred vertically to MS medium, strong seedling Cultivate; when the seedling grows to 3cm-4cm, the stem grows to 0.5cm, and the number of roots is 2-4, it is set aside; Step 2, top cutting treatment: the plants cultivated by strong seedlings are cut off at the top of the stem under the ultra-clean table to eliminate the top advantage of the plant, and then transferred to MS medium for cultivation; Step 3. Dark culture induces internode elongation: the plants after the top cutting of strong seedlings are put into a carton box for dark culture; the internode elongation length of the plant after dark culture is 1cm-4cm; Step 4, light culture: after the end of the dark period, remove the plants with stem elongation ≥ 3 cm out of the carton, and induce the bud germination of the plants that have been in the dark environment for a long time under light conditions; Step 5, segmented subculture: Cut the plants induced by bud germination into small sections with 2-3 nodes, which are respectively terminal bud segment, middle segment, and rooted base segment, and vertically inoculate them on the medium respectively to induce axillary buds germination; Step 6. Formation of regenerated plants: after 1.5 months of segmental subculture, the axillary buds on the nodes will germinate, and will start to take root at the base after 2 months, and form a complete plant after 3-4 months; Step 7, plant transplanting: the complete plant is gently cut off from the mother plant, and transplanted onto MS medium or directly into the substrate to complete plant transplantation. 3. The method for cultivating red bud pineapple according to claim 2, characterized in that the plantlets in the step 1 are plants with healthy growth and 5-8 leaves. 4. The method for cultivating red bud pineapple according to claim 2, characterized in that, the conditions for cultivating strong seedlings in the step 1 are at 25±2° C. and under the condition of light of 12 h/d for 28 days. 5 . The method for cultivating red bud pineapple according to claim 2 , characterized in that, the culturing condition in MS medium in the step 2 is: culturing for one week at 25±2° C. and 12 h/d of light. 6. The cultivation method of red bud pineapple according to claim 2, characterized in that the dark cultivation conditions in the step 3 are: cultivation temperature 25±2°C; cultivation time 40d, during which it is taken out for 1h every 10d. 7. The method for cultivating red bud pineapple according to claim 2, characterized in that the condition for inducing bud germination in the step 4 is light culture, specifically: culture at 25±2°C and 12h/d of light 15d. 8. The method for cultivating red bud pineapple according to claim 2, characterized in that, the subculture medium of the subculture in the step 5 is MS+BA3mg/l+NAA 2mg/l. 9. method for cultivating red bud pineapple according to claim 2, is characterized in that, the substrate in described step 7 comprises river sand and coconut bran that mass ratio is 1:1.