CN115843689A - Efficient nontoxic pear germplasm resource in-vitro culture method - Google Patents
Efficient nontoxic pear germplasm resource in-vitro culture method Download PDFInfo
- Publication number
- CN115843689A CN115843689A CN202211583526.8A CN202211583526A CN115843689A CN 115843689 A CN115843689 A CN 115843689A CN 202211583526 A CN202211583526 A CN 202211583526A CN 115843689 A CN115843689 A CN 115843689A
- Authority
- CN
- China
- Prior art keywords
- pear
- culture
- vitro
- explants
- vitro culture
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses an efficient nontoxic pear germplasm resource in vitro culture method, and belongs to the field of plant in vitro culture. The method comprises the following steps: taking annual pear branches from dormancy in winter to germination for in vitro culture, taking extracted 5-10cm tender branches as explants, soaking and disinfecting the tender branches twice by using 75% ethanol, inoculating an MS solid culture medium, and culturing by illumination to obtain the pear germplasm resource in vitro tissue culture seedlings. The invention adopts tender shoots cultured indoors from dormant pear branches as explants, and the tissue culture seedlings of pear germplasm resources can be obtained by in vitro culture only twice by using 75% ethanol for disinfection and 30s each time. The method is simple and easy to implement, simple to operate, few in working procedures, fast, efficient, non-toxic and residue-free, the explants are not easy to brown, the pollution rate is low, the survival rate of the in vitro culture tissue culture seedlings can reach 100% at most, and the in vitro tissue culture seedlings of different pear germplasm resources can be efficiently obtained in 6-9 weeks.
Description
Technical Field
The invention relates to the field of in vitro culture of plants, in particular to an in vitro culture method of an efficient nontoxic pear germplasm resource.
Background
The pear is a perennial deciduous fruit tree of the genus Pyri (Pyrus L.) of the subfamily Maloideae or Maloideae of the family Rosaceae, and is one of the important fruits worldwide because of its strong adaptability and wide distribution, and is planted in more than 80 countries and regions worldwide. The original species of pears originate from the western or southwest mountain areas of China in the third era, and more than 30 species of known pear plants exist, wherein 13 species of pear plants originally produced in China exist, and the variety resources are more than 3000.
The agricultural germplasm resources are strategic resources for guaranteeing national grain safety and effective supply of important agricultural and sideline products, and are material bases for original innovation of agricultural science and technology and development of modern species industry. The germplasm resource safe preservation is directly related to variety innovation and species industry development, the preservation of the germplasm resources of fruit trees in China is mainly performed in a field garden at present, and a diversified complete preservation system consisting of a germplasm garden, an ultralow temperature warehouse, a test tube Miao Ku, a low temperature seed warehouse, a DNA warehouse and the like is successively built in developed countries in the west. The diversified preservation level of the Chinese fruit tree germplasm resources needs to be improved urgently, and the in vitro culture of the fruit tree germplasm resources is a key link for establishing a test-tube plantlet library.
At present, tissue culture test-tube plantlets of different pear varieties can be obtained through in vitro culture, for example, in vitro test-tube plantlets are obtained from varieties such as 'Laiyang pear', 'brocade pear', 'early crisp', 'duck pear', 'Qiu Baili', 'water of happiness', 'Xinbei pear seven', 'Dangshan crisp pear' and 'yellow pear', and the like, and in vitro culture methods of different varieties are different and have no unified and efficient culture method. In vitro culture of pears mostly selects spring or autumn young shoots which germinate and have no diseases and insect pests on strong plants in the field as explants. The young shoots are cleaned by water and then disinfected, wherein the disinfection is generally carried out by soaking the young shoots in 70-75% ethanol for 10-30 s, washing the young shoots with sterile water for 3-5 times and then using 0.1% HgCl 2 Sterilizing for 8-10 min, and finally washing with sterile water for 3-5 times; or 2 to 5 percent of sodium hypochlorite is soaked for 8 to 15min, washed for 5 to 6 times by sterile water and then 0.1 percent of HgCl is used 2 Sterilizing for 8-10 min, and washing with sterile water for 3-5 times. In the in vitro culture, the sterilization of the explants mostly uses mercury bichloride or sodium hypochlorite as a main disinfectant, the sterilizing effect of the mercury bichloride is excellent, but the mercury bichloride is extremely toxic and is difficult to remove residual medicaments, so that the environment is polluted, and the mortality of the in vitro culture after the sterilization is extremely high; the calcium hypochlorite has low toxicity but general bactericidal effect, the isolated culture is easy to brown and pollute, and the survival rate is low. Therefore, the method for in vitro culture of test-tube plantlets by taking the young pear shoot segments as explants and adopting mercuric chloride or sodium hypochlorite for disinfection is not efficient. At present, an in vitro culture method which is efficient, rapid, simple and nontoxic and is suitable for different variety resources is lacked.
Disclosure of Invention
The invention aims to provide a high-efficiency nontoxic pear germplasm resource in-vitro culture method, which aims to solve the problems in the prior art and realize the purpose of efficiently, simply and nontoxic obtaining pear germplasm resource in-vitro test-tube plantlets by selecting explants, screening in a disinfection mode and optimizing the culture process.
In order to achieve the purpose, the invention provides the following scheme:
the invention provides a method for in vitro culture of high-efficiency nontoxic pear germplasm resources, which comprises the following steps:
taking tender shoots cultured indoors from branches of annual pear trees after dormancy as explants, soaking and disinfecting the explants twice by using 75% ethanol, inoculating MS solid culture medium, and culturing by illumination to obtain the in vitro tissue culture seedlings of pear germplasm resources.
Preferably, the method for indoor culture of the tender tips of the pear branches comprises the following steps: culturing the pear branches by using a 1/10MS culture solution, replacing the 1/10MS culture solution every 5 days, and inducing and culturing tender tips; the culture conditions are as follows: the illumination time is 12h, the illumination intensity is 1500-2100lx, and the temperature is 25-27 ℃.
Preferably, when the tender tip is 5-10cm long, 3-5 leaves are unfolded and 2-4 visible internodes are grown, and the leaf on the tender tip is removed to be used as an explant.
Preferably, the sterilization method comprises: soaking in 75% ethanol for 30s, washing with sterile water for 3-5 times, soaking in 75% ethanol for 30s, and washing with sterile water for 3-5 times.
Preferably, the MS solid medium comprises the following concentration components: MS culture medium +30g/L sucrose +5.5g/L agar.
Preferably, the light culture conditions are: the temperature is 25-27 ℃, the illumination intensity is 2000lx and the illumination time is 16h/d.
The invention discloses the following technical effects:
the invention discloses a high-efficiency nontoxic pear germplasm resource in vitro culture method, which adopts tender shoots cultured indoors by dormant pear branches as explants, and disinfects twice by using 75% ethanol for 30s each time. The disinfection treatment is simple, convenient and fast, no toxic or side effect exists, the explant is not easy to brown, the pollution rate is low, the in vitro tissue culture seedlings of different pear germplasm resources can be efficiently obtained in 6-9 weeks, and the survival rate of the in vitro tissue culture seedlings can reach 100 percent at most. The invention provides technical support for the establishment of the pear germplasm resource test tube Miao Ku and lays a foundation for pear genetic improvement and virus-free seedling breeding.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required in the embodiments will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art that other drawings can be obtained according to the drawings without creative efforts.
FIG. 1 is a schematic diagram of different steps of a high-efficiency nontoxic pear germplasm resource in vitro culture method; a: inducing explants; b: collecting explants; c: sterilizing explants; d: culturing explants; e: obtaining the in vitro tissue culture seedling.
Detailed Description
Reference will now be made in detail to various exemplary embodiments of the invention, the detailed description should not be construed as limiting the invention but rather as a more detailed description of certain aspects, features and embodiments of the invention.
It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. In addition, for numerical ranges in the present disclosure, it is understood that each intervening value, to the upper and lower limit of that range, is also specifically disclosed. Every smaller range between any stated value or intervening value in a stated range and any other stated or intervening value in a stated range is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the documents are cited. In case of conflict with any incorporated document, the present specification will control.
It will be apparent to those skilled in the art that various modifications and variations can be made in the specific embodiments of the present disclosure without departing from the scope or spirit of the disclosure. Other embodiments will be apparent to those skilled in the art from consideration of the specification. The specification and examples are exemplary only.
As used herein, the terms "comprising," "including," "having," "containing," and the like are open-ended terms that mean including but not limited to.
Example 1
A method for in vitro culture of high-efficiency nontoxic pear germplasm resources comprises the following steps:
(1) Explant Induction (A in FIG. 1)
After dormancy in winter, strong branches without diseases and insect pests of pear germplasm resources of different species such as Chinese pears, white pears, birch pears, western pears and the like are collected from a Chinese pear germplasm resource garden (Wuhan) of a fruit tree tea research institute of the academy of agricultural sciences of Hubei province. Placing the branches in a beaker containing 1/10MS culture solution, changing fresh culture solution every 5 days, and culturing under the conditions of illumination for 12 hours every day, illumination intensity of 1500lx and temperature of 25 ℃.
(2) Explant collection (B in FIG. 1)
After the branches are cultured for 3 weeks, when new tender tips are extracted from the annual branches, collecting tender tips of 5cm extracted from the annual branches to be explants, requiring 3 leaves of the tender tips to be unfolded and 2 internodes to be visible, and removing tender leaves on the tender tips to obtain tender stems serving as the explants.
(3) Explant sterilization (C in figure 1)
On a clean bench, the explants were soaked in 75% ethanol for 30s, washed with sterile water for 3 times, then soaked in 75% ethanol for 30s, and washed with sterile water for 3 times.
(4) Isolated culture (D in FIG. 1)
The sterilized explants were inoculated on MS solid medium, MS medium +30g/L sucrose +5.5g/L agar, pH 5.8. In vitro culture is carried out under the conditions that the temperature is 25 ℃, the illumination intensity is 2000lx and the illumination is 16h/d, and strong in vitro tissue culture seedlings of pear germplasm resources are obtained after 3 weeks (E in figure 1).
Example 2
A method for in vitro culture of high-efficiency nontoxic pear germplasm resources comprises the following steps:
(1) Explant induction
After dormancy in winter, chinese pear germplasm resource gardens (Wuhan) of the institute of tea and fruit trees of academy of agricultural sciences of Hubei province collect annual robust branches without diseases and insect pests from Chinese pear germplasm resources of different species such as Chinese pear, white pear, pyrus calleryana, chinese pear and western pear. Placing the branches in a beaker containing 1/10MS culture solution, changing fresh culture solution every 5 days, and culturing under the conditions of illumination for 12 hours every day, illumination intensity of 1800lx and temperature of 26 ℃.
(2) Explant Collection
After the branches are cultured for 4 weeks, when new fresh tender tips are extracted from the annual branches, collecting tender tips of 8cm extracted from the annual branches which are cultured as explants, requiring 4 leaves and 3 visible internodes of the tender tips, and removing tender stems obtained by removing tender leaves which are slightly unfolded to be used as the explants.
(3) Explant sterilization
On a clean bench, the explants were soaked in 75% ethanol for 30s, rinsed with sterile water for 4 times, then soaked in 75% ethanol for 30s, rinsed with sterile water for 4 times.
(4) In vitro culture
The sterilized explants were inoculated on MS solid medium, MS medium +30g/L sucrose +5.5g/L agar, pH 5.9. In vitro culture is carried out under the conditions that the temperature is 26 ℃, the illumination intensity is 2000lx and the illumination is 16h/d, and the robust pear germplasm resource in vitro tissue culture seedlings are obtained after 4 weeks.
Example 3
A method for in vitro culture of high-efficiency nontoxic pear germplasm resources comprises the following steps:
(1) Explant induction
After dormancy in winter, strong branches without diseases and insect pests of pear germplasm resources of different species such as Chinese pears, white pears, birch pears, western pears and the like are collected from a Chinese pear germplasm resource garden (Wuhan) of a fruit tree tea research institute of the academy of agricultural sciences of Hubei province. Placing the branches in a beaker containing 1/10MS culture solution, changing fresh culture solution every 5 days, and culturing under the conditions of illumination for 12 hours every day, illumination intensity of 2100lx and temperature of 27 ℃.
(2) Explant Collection
After the branches are cultured for 5 weeks, when new fresh tender tips are extracted from annual branches, collecting tender shoots of 10cm from the annual branches, 5 pieces of tender tip spread leaves and 4 visible internodes are required, and tender stems obtained by removing tender tip spread leaves are used as explants.
(3) Explant sterilization
On a clean bench, the explants were soaked in 75% ethanol for 30s, washed with sterile water for 5 times, then soaked in 75% ethanol for 30s, and washed with sterile water for 5 times.
(4) In vitro culture
The sterilized explants were inoculated on MS solid medium, MS medium +30g/L sucrose +5.5g/L agar, pH 6.0. In vitro culture is carried out under the conditions that the temperature is 27 ℃, the illumination intensity is 2000lx and the illumination is 16h/d, and the robust pear germplasm resource in vitro tissue culture seedlings are obtained after 4 weeks.
Comparative example 1
The difference from example 1 is that: the explant collection of step (2) in example 1 (step (1) explant induction was omitted accordingly) was replaced with spring field-grown shoots, and the other steps were the same.
Comparative example 2
The difference from example 1 is that: (2) Replacing the explants of example 1 with shoots taken from the spring field (step (1) explant induced corresponding omission), (3) sterilizing the explants on a clean bench, soaking the explants in 75% ethanol for 30s, rinsing with sterile water 3 times, and then 0.1% HgCl 2 Sterilizing for 8-10 min, and finally washing with sterile water for 3 times. The rest steps are the same.
Comparative example 3
The difference from example 1 is that: (2) Replacing the explant in example 1 with the shoots extracted from the spring field (step (1) explant induction was omitted accordingly), (3) sterilizing the explant on a clean bench, soaking the explant in 75% ethanol for 30s, washing with sterile water for 3 times, soaking in 5% sodium hypochlorite for 8min, and washing with sterile water for 3 times. The rest steps are the same.
Comparative example 4
The difference from example 1 is that: (2) Replacing the explant of example 1 with shoots from spring field (step (1) explant induced corresponding omission), (3) sterilizing the explant on a clean bench, soaking the explant in 5% sodium hypochlorite for 8min, washing with sterile water 3 times, and then using 0.1% HgCl 2 Sterilizing for 10min, and washing with sterile water for 3 times. The rest steps are the same.
Comparative example 5
The difference from example 1 is that: (3) Sterilizing explant on a superclean bench, soaking the explant in 75% ethanol for 30s, washing with sterile water for 3 times, and adding 0.1% HgCl 2 Sterilizing for 8-10 min, and finally washing with sterile water for 3 times. The rest steps are the same.
Comparative example 6
The difference from example 1 is that: (3) The explant is sterilized on a superclean workbench, and is soaked in 75% ethanol for 30s, washed with sterile water for 3 times, then soaked in 5% sodium hypochlorite for 8min, and washed with sterile water for 3 times. The rest steps are the same.
Comparative example 7
The difference from example 1 is that: (3) Sterilizing explant on a clean bench, soaking the explant in 5% sodium hypochlorite for 8min, washing with sterile water for 3 times, and washing with 0.1% HgCl 2 Sterilizing for 10min, and washing with sterile water for 3 times. The rest steps are the same.
The pear germplasm resource in vitro tissue culture seedlings obtained by the culture of the methods of the examples 1-3 and the comparative examples 1-5 are cultured, and the in vitro culture browning condition and the survival rate of the explants are counted through the culture, and the results are shown in table 1.
Survival rate (%) = (in vitro tissue culture seedling of mature pear germplasm resource/total number of explants) × 100%
Browning rate (%) = (number of browned explants/total number of explants) × 100%
Contamination rate (%) = (isolated tissue culture seedling contaminated with fungus or bacteria/total number of explants) × 100%
TABLE 1 statistical results of indexes related to in vitro tissue culture seedling of pear germplasm resources cultured by different methods
From the data, the selection of the explant, the selection of the disinfection reagent and the disinfection mode are directly related to the culture effect of the in vitro tissue culture seedling of the pear germplasm resource. The analysis can be caused by the following factors: during the conventional in vitro culture of pears, most of selected explants are twigs or stem tips which are just germinated in spring or autumn, and plants which usually grow in the natural environment or in the field are easily infected with microorganisms on the outside and inside and carry more germs, so that the in vitro culture is easy to pollute, and the survival rate is not high. Mercuric chloride or sodium hypochlorite is mostly used as a main disinfectant for disinfecting the pear explants, and although the mercuric chloride has a good sterilizing effect, the mercuric chloride is extremely toxic and difficult to remove residual medicaments, so that the environment is polluted, and the death rate of the disinfected cultivated plants is extremely high; the calcium hypochlorite has low toxicity but general bactericidal effect, the isolated culture is easy to brown and pollute, and the survival rate is low. The disinfection time of the pear explants is generally 10-20min, the cleaning is carried out for 6-9 times, the explants which are not disinfected sufficiently are easy to be polluted if the disinfection time is short, and the survival rate of the explants which are easy to brown is low if the disinfection time is long. Tender shoots cultured by the dormant pear branches are explants, and are fresh, so that pathogenic bacteria are few, and pollution sources are greatly reduced. When in disinfection, 75% ethanol is used for disinfection twice, the total disinfection time is 1min, and the highest survival rate of the in vitro culture tissue culture seedlings can reach 100%. And the reduction of the disinfection frequency of the ethanol may cause the reduction of the survival rate due to the existence of a small amount of pathogenic bacteria. In a word, the method disclosed by the invention is simple and easy to implement, simple to operate, fast and efficient, and free of toxicity and residue, can be used for conveniently and efficiently obtaining the in vitro tissue culture seedling of the pear germplasm resource, provides technical support for the establishment of the pear germplasm resource test tube Miao Ku, and lays a foundation for genetic improvement and virus-free seedling breeding of pears.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.
Claims (6)
1. The method for in vitro culture of the high-efficiency nontoxic pear germplasm resources is characterized by comprising the following steps:
taking tender shoots cultured indoors from branches of annual pear trees after dormancy as explants, soaking and disinfecting the explants twice by using 75% ethanol, inoculating MS solid culture medium, and culturing by illumination to obtain the in vitro tissue culture seedlings of pear germplasm resources.
2. The method for in vitro culture of the efficient nontoxic pear germplasm resource according to claim 1, wherein the method for indoor culture of the tender tips of the pear branches comprises the following steps: culturing the pear branches by using a 1/10MS culture solution, replacing the 1/10MS culture solution every 5 days, and inducing and culturing tender tips; the culture conditions were: the illumination time is 12h, the illumination intensity is 1500-2100lx, and the temperature is 25-27 ℃.
3. The method for in vitro culture of a high efficiency nontoxic pear germplasm resource of claim 2, wherein the young plant grows to 5-10cm, 3-5 leaves are unfolded, 2-4 visible internodes are grown, and the leaf on the young plant is removed and used as an explant.
4. The method for in vitro culture of the efficient nontoxic pear germplasm resource of claim 1, wherein the disinfection method comprises the following steps: soaking in 75% ethanol for 30s, washing with sterile water for 3-5 times, soaking in 75% ethanol for 30s, and washing with sterile water for 3-5 times.
5. The method for in vitro culture of the efficient nontoxic pear germplasm resource of claim 1, wherein the MS solid culture medium comprises the following components in concentration: MS culture medium +30g/L sucrose +5.5g/L agar.
6. The method for in vitro culture of the efficient nontoxic pear germplasm resources according to claim 1, wherein the illumination culture conditions are as follows: the temperature is 25-27 ℃, the illumination intensity is 2000lx and the illumination time is 16h/d.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211583526.8A CN115843689A (en) | 2022-12-09 | 2022-12-09 | Efficient nontoxic pear germplasm resource in-vitro culture method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211583526.8A CN115843689A (en) | 2022-12-09 | 2022-12-09 | Efficient nontoxic pear germplasm resource in-vitro culture method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN115843689A true CN115843689A (en) | 2023-03-28 |
Family
ID=85671776
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211583526.8A Pending CN115843689A (en) | 2022-12-09 | 2022-12-09 | Efficient nontoxic pear germplasm resource in-vitro culture method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN115843689A (en) |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104221799A (en) * | 2014-08-27 | 2014-12-24 | 湖北省农业科学院果树茶叶研究所 | Method for breeding virus-free pear tube seedlings |
| CN105815219A (en) * | 2016-03-28 | 2016-08-03 | 安徽农业大学 | Method for acquiring explants on basis of young shoot pinching in June and July to increase Dangshansu pear tissue culture survival rate |
| CN109258462A (en) * | 2018-09-17 | 2019-01-25 | 江苏强农农业技术服务有限公司 | A kind of germplasm resource preservation method of shortcake pears |
| CN113854151A (en) * | 2021-10-29 | 2021-12-31 | 中国热带农业科学院海口实验站 | Tissue culture and rapid propagation method for avocados |
| CN113875585A (en) * | 2021-09-24 | 2022-01-04 | 毕节市林业科学研究所 | Method for in-vitro rapid propagation and seedling raising of roxburgh rose |
| CN114041421A (en) * | 2021-11-25 | 2022-02-15 | 中国热带农业科学院海口实验站 | Tissue rapid propagation method of avocados |
-
2022
- 2022-12-09 CN CN202211583526.8A patent/CN115843689A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104221799A (en) * | 2014-08-27 | 2014-12-24 | 湖北省农业科学院果树茶叶研究所 | Method for breeding virus-free pear tube seedlings |
| CN105815219A (en) * | 2016-03-28 | 2016-08-03 | 安徽农业大学 | Method for acquiring explants on basis of young shoot pinching in June and July to increase Dangshansu pear tissue culture survival rate |
| CN109258462A (en) * | 2018-09-17 | 2019-01-25 | 江苏强农农业技术服务有限公司 | A kind of germplasm resource preservation method of shortcake pears |
| CN113875585A (en) * | 2021-09-24 | 2022-01-04 | 毕节市林业科学研究所 | Method for in-vitro rapid propagation and seedling raising of roxburgh rose |
| CN113854151A (en) * | 2021-10-29 | 2021-12-31 | 中国热带农业科学院海口实验站 | Tissue culture and rapid propagation method for avocados |
| CN114041421A (en) * | 2021-11-25 | 2022-02-15 | 中国热带农业科学院海口实验站 | Tissue rapid propagation method of avocados |
Non-Patent Citations (1)
| Title |
|---|
| 田海青: ""新梨7号梨组培快繁体系建立及微嫁接技术的研究"", 《中国优秀硕士学位论文全文数据库 农业科技辑》, no. 3, pages 048 - 117 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108293878B (en) | Tissue culture seedling raising method for trichosanthes kirilowii Maxim tender leaves | |
| CN107155898B (en) | A kind of dendrobium candidum carries out expanding numerous method using stem section thin slice | |
| CN109258477B (en) | Tissue culture method for pachyrhizua angulatus | |
| CN110192524A (en) | A method of using base of leaf and peduncle kryptoblast as the in vitro fast breeding of the zingiberaceous plant of explant | |
| CN104855292A (en) | Method for tissue culture and rapid propagation of stems of cinnamomum kanehirae hay | |
| CN108935108A (en) | A kind of sweet potato tissue-cultured seedling detoxification tissue culture method | |
| CN111657151A (en) | Rapid seedling method for acer truncatum | |
| CN113331059B (en) | Method for establishing efficient regeneration system by taking bird king tea tree hypocotyls as explants | |
| CN111053032A (en) | Rapid propagation method for promoting shoot elongation and segmented proliferation of purple-bud tea tissue culture seedlings | |
| CN102577960A (en) | Tissue culture method of potatoes | |
| CN107223566B (en) | A kind of Wulian poplar method for tissue culture | |
| Nhut et al. | High frequency shoot formation of yellow passion fruit (Passiflora edulis f. flavicarpa) via thin cell layer (TCL) technology | |
| CN113331052B (en) | Process for cultivating superior blueberry products by using micro-freezing biotechnology | |
| CN105145358A (en) | Tissue culture and rapid propagation method for common fibraurea stem | |
| CN115843689A (en) | Efficient nontoxic pear germplasm resource in-vitro culture method | |
| CN113099866B (en) | Method for rapidly propagating fir excellent family seedlings | |
| CN111316919B (en) | Method for improving regeneration efficiency in cinnamomum camphora tissue culture process | |
| CN114946654A (en) | Phalaenopsis stem tip growth point culture virus removal technology | |
| CN110583489B (en) | Tissue culture rapid propagation method and application of populus euphratica | |
| LU103126B1 (en) | Virus-free seedling breeding method for jinsha pomelo and application | |
| CN114009337B (en) | Method for disinfecting tissue culture explants of roses in axillary bud germination period | |
| CN109122321A (en) | A kind of breeding method of the tissue culture method for obtaining high-quality bletilla pseudobulb and bletilla seedling | |
| Kumari et al. | In vitro Propagation of Medicinally Valuable Traditional Banana Cultivar, Musa acuminata cv. Matti by Shoot Tip Culture | |
| CN109329027B (en) | Efficient propagation method of sweet potatoes | |
| CN110192528B (en) | Method for inducing adventitious buds of cyclocarya paliurus and subculture multiplication culture |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |