CN116267600B - Tissue culture seedling renewal and strengthening culture medium for asparagus plants and application thereof - Google Patents
Tissue culture seedling renewal and strengthening culture medium for asparagus plants and application thereof Download PDFInfo
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- 239000001963 growth medium Substances 0.000 title claims abstract description 38
- 238000005728 strengthening Methods 0.000 title abstract description 24
- 244000003416 Asparagus officinalis Species 0.000 title 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims abstract description 129
- 241000234427 Asparagus Species 0.000 claims abstract description 61
- 239000012137 tryptone Substances 0.000 claims abstract description 29
- OVBPIULPVIDEAO-LBPRGKRZSA-N folic acid Chemical compound C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-LBPRGKRZSA-N 0.000 claims description 70
- 235000005340 Asparagus officinalis Nutrition 0.000 claims description 38
- OVBPIULPVIDEAO-UHFFFAOYSA-N N-Pteroyl-L-glutaminsaeure Natural products C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)NC(CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-UHFFFAOYSA-N 0.000 claims description 35
- 229960000304 folic acid Drugs 0.000 claims description 35
- 235000019152 folic acid Nutrition 0.000 claims description 35
- 239000011724 folic acid Substances 0.000 claims description 35
- PRPINYUDVPFIRX-UHFFFAOYSA-N 1-naphthaleneacetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CC=CC2=C1 PRPINYUDVPFIRX-UHFFFAOYSA-N 0.000 claims description 14
- HFCYZXMHUIHAQI-UHFFFAOYSA-N Thidiazuron Chemical compound C=1C=CC=CC=1NC(=O)NC1=CN=NS1 HFCYZXMHUIHAQI-UHFFFAOYSA-N 0.000 claims description 14
- FAIXYKHYOGVFKA-UHFFFAOYSA-N Kinetin Natural products N=1C=NC=2N=CNC=2C=1N(C)C1=CC=CO1 FAIXYKHYOGVFKA-UHFFFAOYSA-N 0.000 claims description 12
- QANMHLXAZMSUEX-UHFFFAOYSA-N kinetin Chemical compound N=1C=NC=2N=CNC=2C=1NCC1=CC=CO1 QANMHLXAZMSUEX-UHFFFAOYSA-N 0.000 claims description 12
- 229960001669 kinetin Drugs 0.000 claims description 12
- 230000003716 rejuvenation Effects 0.000 claims description 12
- 241000196324 Embryophyta Species 0.000 claims description 10
- YNWVFADWVLCOPU-MDWZMJQESA-N (1E)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1H-1,2,4-triazol-1-yl)pent-1-en-3-ol Chemical compound C1=NC=NN1/C(C(O)C(C)(C)C)=C/C1=CC=C(Cl)C=C1 YNWVFADWVLCOPU-MDWZMJQESA-N 0.000 claims description 6
- 239000002609 medium Substances 0.000 claims description 5
- 239000006870 ms-medium Substances 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 10
- 238000004161 plant tissue culture Methods 0.000 abstract description 3
- 239000005648 plant growth regulator Substances 0.000 abstract description 2
- 210000001519 tissue Anatomy 0.000 description 45
- 238000012360 testing method Methods 0.000 description 20
- 230000035755 proliferation Effects 0.000 description 12
- NWBJYWHLCVSVIJ-UHFFFAOYSA-N N-benzyladenine Chemical compound N=1C=NC=2NC=NC=2C=1NCC1=CC=CC=C1 NWBJYWHLCVSVIJ-UHFFFAOYSA-N 0.000 description 10
- 235000020415 coconut juice Nutrition 0.000 description 9
- 230000004069 differentiation Effects 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 8
- 229930003756 Vitamin B7 Natural products 0.000 description 8
- 239000000306 component Substances 0.000 description 8
- 239000011735 vitamin B7 Substances 0.000 description 8
- 235000011912 vitamin B7 Nutrition 0.000 description 8
- 230000012010 growth Effects 0.000 description 7
- 230000008569 process Effects 0.000 description 5
- 230000001737 promoting effect Effects 0.000 description 5
- 238000012258 culturing Methods 0.000 description 4
- 238000009472 formulation Methods 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 229940088594 vitamin Drugs 0.000 description 4
- 229930003231 vitamin Natural products 0.000 description 4
- 235000013343 vitamin Nutrition 0.000 description 4
- 239000011782 vitamin Substances 0.000 description 4
- 235000013162 Cocos nucifera Nutrition 0.000 description 3
- 244000060011 Cocos nucifera Species 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 230000009471 action Effects 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 230000002503 metabolic effect Effects 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 150000003722 vitamin derivatives Chemical class 0.000 description 3
- 239000002699 waste material Substances 0.000 description 3
- 238000004383 yellowing Methods 0.000 description 3
- ALYNCZNDIQEVRV-UHFFFAOYSA-N 4-aminobenzoic acid Chemical compound NC1=CC=C(C(O)=O)C=C1 ALYNCZNDIQEVRV-UHFFFAOYSA-N 0.000 description 2
- 241000233932 Sparganium Species 0.000 description 2
- 238000012136 culture method Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 230000008635 plant growth Effects 0.000 description 2
- 241000234280 Liliaceae Species 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 230000003139 buffering effect Effects 0.000 description 1
- 230000009194 climbing Effects 0.000 description 1
- 230000006866 deterioration Effects 0.000 description 1
- 230000008570 general process Effects 0.000 description 1
- 235000008216 herbs Nutrition 0.000 description 1
- 210000001595 mastoid Anatomy 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 230000008719 thickening Effects 0.000 description 1
- 239000003104 tissue culture media Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/002—Culture media for tissue culture
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- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention relates to the technical field of plant tissue culture seedling renewal and strengthening, and discloses a tissue culture seedling renewal and strengthening culture medium of asparagus plants and application thereof, wherein the culture medium comprises a basic culture medium, 0.5-4mg/L plant growth regulator, 0.1-0.5g/L tryptone and 15-90mg/L citric acid, and the tissue culture old seedlings or tender seedlings of the asparagus plants can be cultivated by using the culture medium provided by the invention, so that good renewal and strengthening effects can be achieved.
Description
Technical Field
The invention relates to the technical field of rejuvenation of plant tissue culture seedlings, in particular to a tissue culture seedling rejuvenation culture medium for asparagus plants and application thereof.
Background
Radix asparagi, perennial herbs or semi-shrubs, standing or climbing, often has thick rootstocks and fleshy roots, sometimes spindle-like tuberous roots. After planting root tuber or root cluster of Asparagus plant, the germinated stems and branches are upright and tender, grow rapidly, and the leaves are coated by films and have no elongation. In the later stage of growth, the stem branches grow into leaf-shaped branches, and the leaf-shaped branches are flat, sharp, or nearly cylindrical and have a plurality of sparganium stolonifer (or sparganium stolonifer) or grooves, and are clustered frequently; transparent mastoid serrations sometimes exist on stems, branches and leaf-like branches, called soft bone serrations. The leaves degenerate into a scaly shape with the base extending somewhat into the pitch or thorns.
Asparagus is a hermaphroditic plant of Asparagus of Liliaceae, perennial herb. The male plants have more developed stems, small single quantity of young stems, but high yield. Female plants are tall and big, young stems are thick, and the yield is slightly low.
The conventional propagation method of asparagus mostly adopts the plant propagation or seed propagation, but due to the hereditary heterozygous genus of the asparagus, the uniformity of propagation offspring is poor, the propagation speed is slow, the inter-plant difference is large, and the like, the stability of the seed character of the asparagus is difficult to maintain in the propagation process, so that the phenomena of yield reduction and quality deterioration often exist. The tissue culture rapid propagation technology can better maintain the excellent properties of improved varieties, expand the number of parents, shorten the breeding period, selectively cultivate all-male plants and improve the yield.
At present, the general process of tissue culture of asparagus comprises four stages of primary culture, proliferation, differentiation and rooting, in the rooting stage, the rooting rate of asparagus is inconsistent, and according to literature reports, the rooting rate of asparagus can reach 85%, the tissue culture seedlings which are not rooted are unfavorable for planting, can only be discarded, the waste of materials is caused, and the tissue culture production cost is increased. In addition, as the subculture frequency of the tissue culture bottle seedlings of the asparagus increases, the quality of the bottle seedlings is degraded, the bottle seedlings with degraded quality need to be replaced by adopting explants for primary culture, and the primary culture cost is high.
The tissue culture elder Miao refers to rooting and seedling strengthening culture, after multiple subcultures, bottle seedlings which are not rooted and lignified are not easy to extract new branches when the tissue culture elder Miao is continued, and the old seedlings are easy to yellow when being cultured, and the tissue culture tender seedlings refer to bottle seedlings which are subjected to primary culture on plant explants, and if the tissue culture tender seedlings are too tender, the proliferation, differentiation and rooting culture of subsequent plants are not favored. Affecting the quality of the final tissue culture seedling.
The existing tissue culture medium generally comprises MS culture medium, 6-benzyl amino purine (6-BA), naphthalene Acetic Acid (NAA), kinetin (KT), thidiazuron (TDZ), tryptone and other components, and uniconazole (S-3307) can be used for proliferation and differentiation of tissue culture seedlings, and the existing tissue culture components have poor effects when being applied to updating and strengthening of the Asparagus tissue culture old seedlings and the tissue culture young seedlings.
Folic acid and citric acid are commonly used substances for tissue culture, citric acid mainly plays roles in preventing browning and pH buffering, vitamins and folic acid directly influence metabolic activities of substances such as protein, fat and sugar, and are generally used for promoting proliferation and differentiation of tissue culture seedlings, and related reports about the folic acid and the citric acid on the promotion of old seedlings and the strengthening of tender seedlings do not exist in the prior art.
Disclosure of Invention
Therefore, it is necessary to provide a culture medium for rejuvenating and strengthening tissue culture seedlings of Asparagus plants and application thereof, which increases the existing technical approach for rejuvenating and strengthening tissue culture seedlings of Asparagus plants and solves the problem of high tissue culture primary cost of Asparagus plants.
In order to achieve the aim, the invention provides a culture medium for rejuvenating tissue culture seedlings of Asparagus plants, which comprises a basic culture medium, 0.5-4mg/L plant growth regulator, 0.1-0.5g/L tryptone and 15-90mg/L citric acid.
Further, the amount of citric acid is 24-45ml/L citric acid, and the culture medium also comprises 0-1mg/L folic acid.
Further comprises MS culture medium, 0.1-1.5 mg/L6-benzylaminopurine, 0.1-0.5mg/L naphthylacetic acid, 0.1-0.5mg/L kinetin, 0.1-0.5mg/L thidiazuron, 0.1-0.5g/L tryptone and 0.1-1.0mg/L uniconazole.
Further, MS culture medium, 1.2 mg/L6-benzylaminopurine, 0.2mg/L naphthylacetic acid, 0.2mg/L kinetin, 0.2mg/L thidiazuron, 0.2g/L tryptone, 0.5mg/L uniconazole, 10ml/L citric acid and 5ml/L folic acid are included.
The application of the culture medium for renewing and strengthening the tissue culture seedlings of the asparagus plants is that the culture medium is applied to renewing and strengthening the tissue culture seedlings of the asparagus plants elder Miao.
Further, when tissue culture seedlings of Asparagus plants are updated and strengthened, the culture conditions are photoperiod of 12h/d, temperature of 25-28 ℃ and light intensity of 50-60 mu mol/m 2 ·s。
Further, the Asparagus plant is Asparagus.
The technical scheme has the following beneficial effects:
according to the invention, the updated strengthening culture medium added with citric acid and folic acid can be used for utilizing the non-rooting and lignified old seedlings in the asparagus tissue culture, the available tissue culture branches can be updated and extracted again, the updated branches can be reused in the proliferation, differentiation and rooting steps in the tissue culture step, the step of primary culture from an explant is avoided, the tissue culture production cost is reduced, the waste of materials in the existing asparagus tissue culture method is effectively reduced, the cost is reduced, and the sustainable development of the asparagus plant tissue culture industry is facilitated. The updated strengthening culture medium can also be used for strengthening the tender seedlings after the primary culture of the tissue culture, so that the diameters of branches of the tender seedlings are effectively increased, the diameters of the branches of the tender seedlings are increased, and the quality of the tissue culture seedlings is improved.
Drawings
Fig. 1 shows an asparagus elder Miao according to an embodiment.
FIG. 2 shows the results of culturing the old seedlings of asparagus in the culture medium formulations 1-6 according to the specific embodiment.
Fig. 3 shows a specific embodiment of the young asparagus seedlings.
FIG. 4 shows the results of culturing young asparagus seedlings using the medium formulations 1-6 according to the embodiment.
Detailed Description
In order to describe the technical content, constructional features, achieved objects and effects of the technical solution in detail, the following description is made in connection with the specific embodiments in conjunction with the accompanying drawings.
When the culture medium is prepared, citric acid is firstly dissolved into 3g/L citric acid solution by water, and folic acid is firstly dissolved into 0.1g/L folic acid solution by 1mg/mL sodium hydroxide solution. The vitamin solution was used at a concentration of 0.05 g/L.
1. Updating of old asparagus seedlings
1.1 selection of culture Components
(1) Selecting asparagus elder Miao, namely, bottle seedlings which are not rooted and have lignified after rooting and seedling strengthening culture, wherein the bottle seedlings are shown in figure 1;
(2) Multiple groups of culture mediums with different formulas
Formula 1: and (3) MS.
Formula 2: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5mg/L.
Formula 3: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10ml/L.
Formula 4: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
Formula 5: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 mL/L+folic acid 5 mL/L+vitamin H1mL/L.
Formula 6: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 mL/L+folic acid 5 mL/L+vitamin H1 mL/L+coconut juice 10%.
Formula 7: MS+10 ml/L of citric acid+5 ml/L of folic acid.
(3) Culturing
Respectively taking the lignified old seedlings of the asparagus on an ultra-clean workbench, transferring the lignified old seedlings of the asparagus into a prepared culture medium, and carrying out 3 sections/bottle, wherein each treatment is repeated for 5 times, and the culture conditions are as follows: photoperiod 12h/d, temperature 25-28 O C, light intensity of 50-60 mu mol/m 2 S, the culture period was 30d.
(4) Test results
The test results are shown in Table 1 below, and the tissue culture diagram is shown in FIG. 2
Table 1: different culture mediums have update effect on asparagus elder Miao
Note that: updated branches refer to the number of old branches from which new branches are drawn after cultivation;
update ratio = updated shoots/total inoculated shoots;
the new number of shoots refers to the total number of updated shoots on the inoculated old shoots.
After the new updated branches are separated, new tissue culture seedlings can be formed through proliferation, differentiation and rooting, and the new tissue culture seedlings can be used for subsequent seedling culture to form new asparagus plants.
Compared with test 1, test 3 and test 4 show that citric acid and folic acid are added into the culture medium, so that the growth of the lignified seedlings of the asparagus can be effectively promoted, and new branches can be extracted. Compared with test 2, the updating proportion of the branches and the number of new branches are increased, the new branches are effectively improved, and the old seedlings cannot be yellowing.
Vitamin H and coconut juice are often added into a culture medium in the tissue culture of plants, and in the conventional technology, coconut is used for promoting plant growth, and vitamin H directly influences the metabolic activities of substances such as protein, fat, sugar and the like, so that the coconut juice has a good promoting effect on growth.
Compared with test 4, test 5 and test 6 show that vitamin H and coconut juice cannot be added into the culture medium components, and after the vitamin H and the coconut juice are added, the update proportion of old seedlings is reduced, elder Miao is yellowing, and the update of the old seedlings is inhibited.
Therefore, citric acid and folic acid in the invention can have the effect of stimulating proliferation and growth of old seedlings of the asparagus plants in the updated and strengthened culture medium; the tissue culture elder Miao is promoted to update new branches, and elder Miao is prevented from yellowing, so that an unexpected technical effect is achieved.
1.2 selection of culture Components concentration
The media of experiment 4 were prepared at different concentrations and the best elder Miao update formulation was selected.
Formula 8: MS+6-BA 0.1mg/L+NAA 0.1mg/L+KT 0.1mg/L+TDZ 0.1 mg/L+tryptone 0.1g/L+S-3307 0.1 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
Formula 9: MS+6-BA 0.5mg/L+NAA 0.3mg/L+KT 0.3mg/L+TDZ 0.3 mg/L+tryptone 0.3g/L+S-3307 0.8 mg/L+citric acid 8 ml/L+folic acid 5ml/L.
Formula 10: MS+6-BA 1.0mg/L+NAA 0.5mg/L+KT 0.5mg/L+TDZ 0.5 mg/L+tryptone 0.5g/L+S-3307 0.3 mg/L+citric acid 15 ml/L+folic acid 5ml/L.
Formula 11: MS+6-BA 1.5mg/L+NAA 0.2mg/L+KT0.2 mg/L+TDZ 0.5 mg/L+tryptone 0.5g/L+S-3307 1.0 mg/L+citric acid 30 ml/L+folic acid 10ml/L.
Formula 4: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
The test results are shown in table 2 below,
table 2: different culture mediums update and strengthen the asparagus elder Miao
The table shows that the optimal updating formula for the old asparagus seedlings is as follows: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
2. Strengthening tender seedling of asparagus
1.1 selection of the Components of the strengthening Medium
(1) Selecting tender seedling of Germinatus Phragmitis, namely bottle seedling of explant after primary generation, as shown in figure 3;
(2) Multiple groups of culture mediums with different formulas
Formula 1: and (3) MS.
Formula 2: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5mg/L.
Formula 3: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10ml/L.
Formula 4: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
Formula 5: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 mL/L+folic acid 5 mL/L+vitamin H1mL/L.
Formula 6: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 mL/L+folic acid 5 mL/L+vitamin H1 mL/L+coconut juice 10%.
Formula 7: MS+10 ml/L of citric acid+5 ml/L of folic acid.
(3) Culturing
Respectively taking tender seedlings of the asparagus explants after primary culture on an ultra-clean workbench, transferring the tender seedlings to a prepared culture medium, and carrying out 3 sections/bottle treatment on each seedling, wherein the culture conditions are as follows: photoperiod 12h/d, temperature 25-28 O C, light intensity of 50-60 mu mol/m 2 S, the culture period was 30d.
(4) Test results
The test results are shown in Table 3 below, and the tissue culture diagram is shown in FIG. 4
Table 3: different culture mediums have effects of strengthening proliferation of young asparagus seedlings
Note that: proliferated shoots refer to the number of total newly proliferated shoots on tender shoots after cultivation.
Differentiation ratio = number of shoots differentiated/total number of shoots inoculated.
The proliferated branches can be separated and used for subsequent differentiation and rooting culture to form tissue culture seedlings, and finally, new asparagus plants are formed through seedling culture.
Compared with test 1, test 3 and test 4 show that citric acid and folic acid are added into the culture medium, so that proliferation and growth of branches of asparagus seedlings can be effectively promoted. Compared with test 2, the proliferation quantity of the branches and the diameter of the branches are increased, the branches are effectively improved, and the branches are not differentiated.
Vitamin H and coconut juice are added into a culture medium in the tissue culture of plants frequently, and in the conventional technology, coconut is used for promoting plant growth, and vitamin H directly influences the metabolic activities of substances such as protein, fat, sugar and the like, so that the coconut has a good promoting effect on growth.
Compared with test 4, test 5 and test 6 show that vitamin H and coconut juice cannot be added into the culture medium components, and after the vitamin H and the coconut juice are added, branches growing on the tender seedlings are reduced, the thickening of the branches is affected, and the strengthening of the tender seedlings is inhibited.
Therefore, citric acid and folic acid in the invention can stimulate the proliferation and growth of the asparagus plants in the updated and strengthened culture medium; the citric acid is added into the culture medium, so that the proliferation and strengthening of tissue culture seedlings of the asparagus plants are promoted, and the technical effect of no intention is achieved.
1.2 selection of culture Components concentration
The media of experiment 4 were prepared at different concentrations and the best elder Miao update formulation was selected.
Formula 8: MS+6-BA 0.1mg/L+NAA 0.1mg/L+KT 0.1mg/L+TDZ 0.1 mg/L+tryptone 0.1g/L+S-3307 0.1 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
Formula 9: MS+6-BA 0.5mg/L+NAA 0.3mg/L+KT 0.3mg/L+TDZ 0.3 mg/L+tryptone 0.3g/L+S-3307 0.8 mg/L+citric acid 8 ml/L+folic acid 5ml/L.
Formula 10: MS+6-BA 1.0mg/L+NAA 0.5mg/L+KT 0.5mg/L+TDZ 0.5 mg/L+tryptone 0.5g/L+S-3307 0.3 mg/L+citric acid 15 ml/L+folic acid 5ml/L.
Formula 11: MS+6-BA 1.5mg/L+NAA 0.2mg/L+KT0.2 mg/L+TDZ 0.5 mg/L+tryptone 0.5g/L+S-3307 1.0 mg/L+citric acid 30 ml/L+folic acid 10ml/L.
Formula 4: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
The test results are shown in the following table 4,
table 4: strengthening effect of different culture mediums on asparagus young seedlings
The table shows that the optimal updating formula for the old asparagus seedlings is as follows: MS+6-BA1.2 mg/L+NAA0.2mg/L+KT0.2 mg/L+TDZ0.2 mg/L+tryptone 0.2g/L+S-33070.5 mg/L+citric acid 10 ml/L+folic acid 5ml/L.
In conclusion, the updated strengthening culture medium added with citric acid and folic acid can be used for utilizing the non-rooting and lignified old seedlings in the asparagus tissue culture and renewing the old seedlings into available tissue culture branches, so that the waste of materials in the existing asparagus tissue culture method is effectively reduced, the cost is reduced, the sustainable development of the asparagus tissue culture industry is facilitated, the diameter of the branches can be effectively improved after the tender seedlings after primary culture are cultured by the culture medium provided by the invention, and the strengthening effect is achieved, thereby facilitating the subsequent differentiation and rooting culture.
It is noted that relational terms such as first and second, and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or terminal that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or terminal. Without further limitation, an element defined by the statement "comprising … …" or "comprising … …" does not exclude the presence of additional elements in a process, method, article or terminal device comprising the element. Further, herein, "greater than," "less than," "exceeding," and the like are understood to not include the present number; "above", "below", "within" and the like are understood to include this number.
While the embodiments have been described above, other variations and modifications will occur to those skilled in the art once the basic inventive concepts are known, and it is therefore intended that the foregoing description and drawings illustrate only embodiments of the invention and not limit the scope of the invention, and it is therefore intended that the invention not be limited to the specific embodiments described, but that the invention may be practiced with their equivalent structures or with their equivalent processes or with their use directly or indirectly in other related fields.
Claims (5)
1. The culture medium is characterized by comprising an MS culture medium, 0.1-1.5 mg/L6-benzylaminopurine, 0.1-0.5mg/L naphthylacetic acid, 0.1-0.5mg/L kinetin, 0.1-0.5mg/L thidiazuron, 0.1-0.5g/L tryptone, 0.1-1.0mg/L uniconazole and 8-45 ml/L citric acid, wherein the asparagus is asparagus.
2. The tissue culture seedling rejuvenation medium for asparagus plants according to claim 1, wherein the amount of citric acid is 24-45ml/L citric acid, and the medium further comprises 0-1 ml/L folic acid.
3. The tissue culture seedling rejuvenation medium for plants of the genus asparagus of claim 2, consisting of MS medium, 1.2 mg/L6-benzylaminopurine, 0.2mg/L naphthylacetic acid, 0.2mg/L kinetin, 0.2mg/L thidiazuron, 0.2g/L tryptone, 0.5mg/L uniconazole, 30ml/L citric acid and 0.5ml/L folic acid.
4. The use of a culture medium for rejuvenation of tissue culture seedlings of asparagus plants as defined in any one of claims 1 to 3, wherein the culture medium is used for rejuvenation of tissue culture elder Miao of asparagus plants and rejuvenation of tissue culture seedlings.
5. The use of the culture medium for rejuvenation of tissue culture seedlings of Asparagus plant as defined in claim 4, wherein the culture conditions are photoperiod 12h/d, temperature 25-28deg.C and light intensity 50-60. Mu. Mol/m when rejuvenation of tissue culture seedlings of Asparagus plant is carried out 2 •s。
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