CN114451303A - Tissue culture method of salix mongolica - Google Patents
Tissue culture method of salix mongolica Download PDFInfo
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- CN114451303A CN114451303A CN202011238488.3A CN202011238488A CN114451303A CN 114451303 A CN114451303 A CN 114451303A CN 202011238488 A CN202011238488 A CN 202011238488A CN 114451303 A CN114451303 A CN 114451303A
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- 241000124033 Salix Species 0.000 title claims abstract description 23
- ZZUFCTLCJUWOSV-UHFFFAOYSA-N furosemide Chemical compound C1=C(Cl)C(S(=O)(=O)N)=CC(C(O)=O)=C1NCC1=CC=CO1 ZZUFCTLCJUWOSV-UHFFFAOYSA-N 0.000 title claims abstract description 22
- 238000012136 culture method Methods 0.000 title claims abstract description 7
- 238000000034 method Methods 0.000 claims abstract description 16
- 239000001963 growth medium Substances 0.000 claims description 28
- 238000011081 inoculation Methods 0.000 claims description 10
- 239000004576 sand Substances 0.000 claims description 10
- 238000004659 sterilization and disinfection Methods 0.000 claims description 9
- 238000012258 culturing Methods 0.000 claims description 7
- 239000000203 mixture Substances 0.000 claims description 7
- 239000002689 soil Substances 0.000 claims description 7
- 238000004140 cleaning Methods 0.000 claims description 6
- 238000005286 illumination Methods 0.000 claims description 6
- 230000035755 proliferation Effects 0.000 claims description 5
- 230000001954 sterilising effect Effects 0.000 claims description 5
- 239000012883 rooting culture medium Substances 0.000 claims description 4
- 244000025254 Cannabis sativa Species 0.000 claims description 3
- TWFZGCMQGLPBSX-UHFFFAOYSA-N Carbendazim Natural products C1=CC=C2NC(NC(=O)OC)=NC2=C1 TWFZGCMQGLPBSX-UHFFFAOYSA-N 0.000 claims description 3
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 3
- JNPZQRQPIHJYNM-UHFFFAOYSA-N carbendazim Chemical compound C1=C[CH]C2=NC(NC(=O)OC)=NC2=C1 JNPZQRQPIHJYNM-UHFFFAOYSA-N 0.000 claims description 3
- 239000006013 carbendazim Substances 0.000 claims description 3
- 229910052799 carbon Inorganic materials 0.000 claims description 3
- CRQQGFGUEAVUIL-UHFFFAOYSA-N chlorothalonil Chemical compound ClC1=C(Cl)C(C#N)=C(Cl)C(C#N)=C1Cl CRQQGFGUEAVUIL-UHFFFAOYSA-N 0.000 claims description 3
- 230000000249 desinfective effect Effects 0.000 claims description 3
- 238000002791 soaking Methods 0.000 claims description 3
- 239000012499 inoculation medium Substances 0.000 claims description 2
- 239000002609 medium Substances 0.000 claims description 2
- 239000012882 rooting medium Substances 0.000 claims description 2
- 238000010008 shearing Methods 0.000 claims description 2
- 241000196324 Embryophyta Species 0.000 abstract description 5
- 239000000463 material Substances 0.000 abstract description 4
- 238000011031 large-scale manufacturing process Methods 0.000 abstract description 3
- 238000005406 washing Methods 0.000 description 8
- 238000005520 cutting process Methods 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 6
- 239000008223 sterile water Substances 0.000 description 4
- 239000008399 tap water Substances 0.000 description 4
- 235000020679 tap water Nutrition 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 241000418534 Salix lutea Species 0.000 description 3
- 241000233779 Cyclocarya paliurus Species 0.000 description 2
- 244000071131 Salix caprea Species 0.000 description 2
- 235000000516 Salix caprea Nutrition 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 239000006260 foam Substances 0.000 description 2
- 229960002523 mercuric chloride Drugs 0.000 description 2
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 description 2
- 229920000136 polysorbate Polymers 0.000 description 2
- 241000238631 Hexapoda Species 0.000 description 1
- 241000218998 Salicaceae Species 0.000 description 1
- 241001299682 Salix purpurea Species 0.000 description 1
- 241000607479 Yersinia pestis Species 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
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Classifications
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- 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
- A01H4/001—Culture apparatus for tissue culture
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G31/00—Soilless cultivation, e.g. hydroponics
-
- 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
- A01H4/008—Methods for regeneration to complete plants
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- Life Sciences & Earth Sciences (AREA)
- Developmental Biology & Embryology (AREA)
- Environmental Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a tissue culture method of salix mongolica. The invention realizes the mass propagation of the wickers by using less plant branches through a tissue culture propagation method. The method can reduce the damage to the existing vegetation, provides technical support for the large-scale production of desertification control materials, and has important application value.
Description
Technical Field
The invention belongs to the field of plants, and relates to a tissue culture method of salix mongolica.
Background
Salix mongolica (Salix gordejeviii) is a deciduous and erect shrub of Salix genus of Salicaceae family, and is widely distributed on mobile and semi-mobile sandy land of grassland. The salix mongolica is suitable for quicksand, has strong sprouting power, is easy to grow adventitious roots and adventitious buds when being buried by quicksand to form new branches and root systems, is easy to reproduce asexually, has strong wind resistance, is an excellent pioneer sand fixing species, is widely applied to artificial sand control, and has good effect. Within a certain range, the growth speed of the salix mongolica trunk is positively correlated with the sand burying depth, and as long as the sand burying does not exceed the crown, the thicker the sand layer is, the more vigorous the growth is, and finally, a tall salix mongolica sand bag is often formed.
In the desertification grassland treatment process of the eastern part of the inner Mongolia, branch cutting seedling is often adopted for sand land treatment. In the actual operation process, 1-2 year old branches which are robust and have no plant diseases and insect pests are cut and taken after the flow of the salix mongolica sap is stopped in winter, and are stored in a cellar in a low-temperature and dark place in winter, and are taken out for cutting seedling after soil is unfrozen in spring of the next year. The yellow willow cutting required by cutting needs to cut a large amount of adult willow twigs, so that the original vegetation is easily damaged, and the protection of the original vegetation is not facilitated.
Disclosure of Invention
The invention aims to provide a tissue culture method of salix mongolica.
The tissue culture method of salix mongolica provided by the invention comprises the following steps:
1) explant sterilization
Taking a tender branch of the current year of the salix mongolica as an explant, and disinfecting to obtain a disinfected explant;
2) culturing the sterilized explant obtained in the step 1) by using an inoculation culture medium, transferring the explant into a proliferation culture medium after culturing for 3-7 days until a bud mass is formed, transferring the bud mass into a strong seedling culture medium, shearing off a big seedling, and placing the big seedling into a rooting culture medium for rooting to obtain a tissue culture seedling;
transferring the original bud mass to a new strong seedling culture medium every 30 days or so;
the composition of the inoculation medium is as follows: MS +6-BA (1mg/L) + NAA (0.3mg/L) + sugar (30g/L)
The proliferation medium had the following composition: MS +6-BA (1.5-2.0mg/L) + NAA (0.3mg/L) + sugar (30g/L)
The strong seedling culture medium comprises the following components: MS +6-BA (0.05mg/L) + NAA (0.3mg/L) + sugar (30g/L) the composition of the rooting medium is as follows: 1/2MS + NAA (0.3mg/L-0.5mg/L) + sugar (20 g/L);
in the above-mentioned culture medium, the percentage of each component refers to the concentration in the whole culture medium;
3) taking out the tissue culture seedlings obtained in the step 2) when the root length is 2cm, hardening the seedlings for one week, taking out the seedlings from the bottles, cleaning a culture medium, and planting to finish the tissue culture of the salix mongolica.
In the step 1) of the method, the explant is terminal buds and/or axillary buds on tender branches of the current-year salix mongolica.
Specifically, the sterilization may be performed according to various conventional methods, for example, according to the following method: putting the tender branches into a beaker, adding a few drops of washing agent, adding water, washing the tender branches under tap water until no foam exists, and repeating the steps once. The tap water washing time is not easy to be too long so as to avoid the secondary pollution of the explants. And (5) taking the clean bench to prepare for disinfection and inoculation. Cutting tender branches to 3-4cm length with scissors, placing into a ground bottle for disinfection, soaking in 70% ethanol in a super clean bench for 1 min, washing with sterile water for 2 times, disinfecting with 0.2% mercuric chloride (added with two drops of Tween) for 10 min, and washing with sterile water for 6 times. The specific disinfection time can be adjusted according to the tenderness degree and the size of the material.
In the step 2), the culture step is carried out by using an inoculation culture medium under the following conditions: the illumination time is 12 hours at 23 +/-1 ℃, the illumination intensity is 2000Lx, and the pH value is 5.8-5.9;
the step 3) further comprises the following steps: after the step of taking out the bottle and cleaning the culture medium and before field planting, cleaning the seedlings by using 1000 times of carbendazim or thalonil and soaking the roots of the seedlings.
The culture medium used for field planting is a mixture obtained by mixing disinfected grass carbon soil, sand and the like in mass.
In the planting step, the planting environment conditions are as follows: the plastic shed keeps the relative humidity of air at 60-80%, and appropriately shades the air, and the ambient temperature is 25 +/-1 ℃.
The invention realizes the mass propagation of the wickers by using less plant branches through a tissue culture propagation method. The method can reduce the damage to the original vegetation, provides technical support for the large-scale production of desertification control materials, and has important application value.
Drawings
FIG. 1 shows tissue culture seedling of Salix purpurea. Wherein, 1, 3: explants, shoots and leaves of salix caprea; 2: overground seedlings formed after explant inoculation; 4. 5: rooting culture of tissue culture seedling; 6: rooting the cyclocarya paliurus plant; 7: and (3) culturing the yellow willow seedlings in soil.
Detailed Description
The present invention will be further illustrated with reference to the following specific examples, but the present invention is not limited to the following examples. The method is a conventional method unless otherwise specified. The starting materials are commercially available from the open literature unless otherwise specified.
Examples 1,
Explant sterilization
Selecting terminal buds and axillary buds on the tender branches of the current year as explants. Firstly, taking the tender branches of the current year of the salix mongolica, putting the tender branches into a beaker, adding a few drops of washing agent, adding water, washing the tender branches under tap water until no foam exists, and repeating the steps once. The tap water washing time is not easy to be too long so as to avoid the secondary pollution of the explants. And (5) taking the clean bench to prepare for disinfection and inoculation. The tender branches are cut to 3-4cm in length by scissors, put into a ground bottle for disinfection, soaked in 70% alcohol for 1 minute in a super clean bench, washed with sterile water for 2 times, disinfected with 0.2% mercuric chloride (added with two drops of tween) for about 10 minutes, and finally washed with sterile water for 6 times.
Culture medium
Cutting off redundant leaves and stem segments, reserving 1.5-2.0cm of stem segments with terminal buds and lateral buds for inoculation, and selecting an inoculation culture medium. The culture conditions are as follows: the illumination time is 12 hours at 23 +/-1 ℃, the illumination intensity is about 2000Lx, and the pH value is 5.8; the culture starts to grow about 3-7 days, then the culture is transferred into a multiplication culture medium, and a bud mass is formed about 30 days to generate a large number of buds. Because the seedlings are short and grow slowly, the whole bud mass is transferred to a strong seedling culture medium every time of proliferation, the buds grow up in about 20 days and are transferred every 30 days. When the seedling amount reaches a certain amount, strong and big seedlings can be cut off, and the strong and big seedlings are transferred into a rooting culture medium to induce rooting for about 15 to 20 days.
Inoculating a culture medium: MS +6-BA (1mg/L) + NAA (0.3mg/L) + sugar (30g/L)
And (3) adding a culture medium: MS +6-BA (1.5-2.0mg/L) + NAA (0.3mg/L) + sugar (30g/L)
Strong seedling culture medium: MS +6-BA (0.05mg/L) + NAA (0.3mg/L) + sugar (30g/L)
Rooting culture medium: 1/2MS + NAA (0.3mg/L-0.5mg/L) + sugar (20g/L)
Transplanting of tissue culture seedlings
The tissue culture seedling can be taken out of the bottle when the root grows to 2cm, and the seedling is hardened for one week before being taken out of the bottle. After the seedlings are taken out of bottles and the culture medium is cleaned, the roots of the seedlings are cleaned and soaked by 1000 times of carbendazim or thalonil, so that the occurrence of diseases can be prevented and controlled. The culture medium is prepared by mixing grass carbon soil and sand one by one, and sterilizing the soil. Watering the seedlings for one time by using a watering can after field planting, and arranging a plastic shed to keep the air humidity (the relative air humidity is about 60-80% to be proper) in the early stage of transplanting and appropriately shade, wherein the environmental temperature is controlled to be about 25 ℃.
FIG. 1 shows the main stage of the tissue culture and seedling raising of the salix mongolica. Wherein, 1, 3: explants, shoots and leaves of salix caprea; 2: overground seedlings formed after explant inoculation; 4. 5: rooting culture of tissue culture seedling; 6: rooting the cyclocarya paliurus plant; 7: and (3) culturing the yellow willow seedlings in soil.
The invention realizes the mass propagation of the wickers by using less plant branches through a tissue culture propagation method. The method can reduce the damage to the existing vegetation, provides technical support for the large-scale production of desertification control materials, and has important application value.
Claims (6)
1. A tissue culture method of salix mongolica comprises the following steps:
1) explant sterilization
Taking a tender branch of the current year of the salix mongolica as an explant, and disinfecting to obtain a disinfected explant;
2) selecting an inoculation culture medium for culturing the sterilized explant obtained in the step 1), transferring the explant into a proliferation culture medium after culturing for 3-7 days until a bud mass is formed, transferring the bud mass into a strong seedling culture medium, shearing off a big seedling, and placing the big seedling in a rooting culture medium for rooting to obtain a tissue culture seedling;
the composition of the inoculation medium is as follows: MS +6-BA (1mg/L) + NAA (0.3mg/L) + sugar (30g/L)
The proliferation medium had the following composition: MS +6-BA (1.5-2.0mg/L) + NAA (0.3mg/L) + sugar (30g/L)
The strong seedling culture medium comprises the following components: MS +6-BA (0.05mg/L) + NAA (0.3mg/L) + sugar (30g/L)
The rooting medium comprises the following components: 1/2MS + NAA (0.3mg/L-0.5mg/L) + sugar (20 g/L);
3) taking out the tissue culture seedlings obtained in the step 2) when the root length is 2cm, hardening the seedlings for one week, taking out the seedlings from the bottles, cleaning a culture medium, and planting to finish the tissue culture of the salix mongolica.
2. The method of claim 1, wherein: in the step 1), the explant is terminal buds and/or axillary buds on the tender branch of the current-year salix mongolica.
3. The method according to claim 1 or 2, characterized in that: in the step 2), in the step of culturing with the inoculation culture medium, the conditions are as follows: the illumination time is 12 hours at 23 +/-1 ℃, the illumination intensity is 2000Lx, and the pH value is 5.8-5.9.
4. A method according to any one of claims 1 to 3, wherein: the step 3) further comprises the following steps: after the step of taking out the bottle and cleaning the culture medium and before field planting, cleaning the seedlings by using 1000 times of carbendazim or thalonil and soaking the roots of the seedlings.
5. The method of claim 4, wherein: the culture medium used for field planting is a mixture obtained by mixing disinfected grass carbon soil, sand and the like in mass.
6. The method according to claim 4 or 5, characterized in that: in the planting step, the planting environment conditions are as follows: the plastic shed keeps the relative humidity of air at 60-80%, and appropriately shades the air, and the ambient temperature is 25 +/-1 ℃.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011238488.3A CN114451303A (en) | 2020-11-09 | 2020-11-09 | Tissue culture method of salix mongolica |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202011238488.3A CN114451303A (en) | 2020-11-09 | 2020-11-09 | Tissue culture method of salix mongolica |
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| CN114451303A true CN114451303A (en) | 2022-05-10 |
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| CN202011238488.3A Pending CN114451303A (en) | 2020-11-09 | 2020-11-09 | Tissue culture method of salix mongolica |
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| UA72275U (en) * | 2012-02-15 | 2012-08-10 | Национальный Университет Биоресурсов И Природопользования Украины | Method for propagating in vitro and adapting propagation material of basket willow (salix viminalis l.) for bioenergetics |
| KR20130084028A (en) * | 2012-01-16 | 2013-07-24 | 대한민국(관리부서 : 산림청 국립산림과학원장) | The method of shoot regeneration from salix spp. and its improvement |
| CN103733997A (en) * | 2013-12-24 | 2014-04-23 | 镇江山水湾生态农业开发有限公司 | Fast reproduction method for energy willow tissue culture |
| CN104396760A (en) * | 2014-12-15 | 2015-03-11 | 中国林业科学研究院林业研究所 | Method for tissue culture and rapid propagation of salix dasyclados |
| CN110178726A (en) * | 2019-05-09 | 2019-08-30 | 北京林业大学 | A kind of root media and weeping willow tissue culture and rapid propagation method for weeping willow tissue-culturing rapid propagation |
-
2020
- 2020-11-09 CN CN202011238488.3A patent/CN114451303A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20130084028A (en) * | 2012-01-16 | 2013-07-24 | 대한민국(관리부서 : 산림청 국립산림과학원장) | The method of shoot regeneration from salix spp. and its improvement |
| UA72275U (en) * | 2012-02-15 | 2012-08-10 | Национальный Университет Биоресурсов И Природопользования Украины | Method for propagating in vitro and adapting propagation material of basket willow (salix viminalis l.) for bioenergetics |
| CN103733997A (en) * | 2013-12-24 | 2014-04-23 | 镇江山水湾生态农业开发有限公司 | Fast reproduction method for energy willow tissue culture |
| CN104396760A (en) * | 2014-12-15 | 2015-03-11 | 中国林业科学研究院林业研究所 | Method for tissue culture and rapid propagation of salix dasyclados |
| CN110178726A (en) * | 2019-05-09 | 2019-08-30 | 北京林业大学 | A kind of root media and weeping willow tissue culture and rapid propagation method for weeping willow tissue-culturing rapid propagation |
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Application publication date: 20220510 |