CN111685038A - Culture medium for improving cell division frequency of dendrobium officinale protoplast - Google Patents
Culture medium for improving cell division frequency of dendrobium officinale protoplast Download PDFInfo
- Publication number
- CN111685038A CN111685038A CN202010476002.3A CN202010476002A CN111685038A CN 111685038 A CN111685038 A CN 111685038A CN 202010476002 A CN202010476002 A CN 202010476002A CN 111685038 A CN111685038 A CN 111685038A
- Authority
- CN
- China
- Prior art keywords
- dendrobium officinale
- protoplast
- culture medium
- cell division
- cell
- 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
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
- A01H4/001—Culture apparatus for tissue culture
Landscapes
- 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)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a culture medium for improving the cell division frequency of a dendrobium officinale protoplast, which is prepared from the following components in parts by weight of inorganic salt: KNO3、CaCl2·2H2O、MgSO4·7H2O, etc., organic components: thiamine hydrochloride, pyridoxine hydrochloride, nicotinic acid, D-biotin, glycine, proline, citric acid, inositol, hydrolyzed casein, glutathione, sugar alcohol components: rhamnose, cellobiose, ribose, sucrose, mannitol, other ingredients: 2,4-D, NAA, 6-BA, sodium selenite, L-carnitine, 5-aminolevulinic acid, sodium butyrate, dendrobium officinale leaching liquor and corn juice. Compared with the prior art, the culture medium can promote the cell wall formation and the cell sustained division growth of the dendrobium officinale, obviously improve the cell division frequency and the cell mass formation frequency of the protoplast of the dendrobium officinale, and lay a foundation for establishing a high-efficiency and stable dendrobium officinale protoplast culture system in the future.
Description
Technical Field
The invention relates to a culture medium for improving the cell division frequency of a dendrobium officinale protoplast, belonging to the field of plant cell engineering.
Background
Dendrobium officinale (academic name:Dendrobium officinalekimura et Migo) is a herbal plant of Dendrobium (Dendrobium) of Orchidaceae (Orchidaceae), naturally grows on cliff or big trees, and is mainly distributed in Anhui, Zhejiang, Hunan, Jiangxi, Fujian, Guangdong, Guangxi, Yunnan and other areas in China, so that wild germplasm resources are wide in distribution range and large in area. The dendrobium officinale is a traditional and rare Chinese medicinal material in China, is listed as the head of the nine-large Chinese mesona in the channel storage, and has the effects of nourishing yin, promoting the production of body fluid, moistening throat, protecting throat, warming stomach, improving eyesight, tonifying kidney, benefiting strength, prolonging life and the like. According to the record of 2010 version pharmacopoeia, dendrobium officinale is sweet in nature and taste, slightly cold, enters stomach and kidney channels, benefits stomach, promotes fluid production, nourishes yin and clears heat. Can be used for treating fever with body fluid deficiency, dry mouth, polydipsia, stomach yin deficiency, poor appetite, vomiting, persistent asthenic fever after illness, yin deficiency, hyperactivity of fire, bone steaming, overstrain, dim and poor vision, and flaccidity of tendons and bones. Preparation of Dendrobium officinaleThe main effective components are dendrobium polysaccharides, alkaloids, phenols, amino acids and the like, and modern medical research shows that the dendrobium officinale has the effects of resisting tumors and aging, enhancing the immunity of the organism, reducing blood sugar, effectively relieving liver injury and the like. With the improvement of living standard and the increase of health care consciousness of people, the medicinal value of the dendrobium officinale is gradually accepted by the public and the market is in short supply. At present, the tissue culture technology is mature day by day, and the dendrobium officinale can realize large-scale propagation by an industrial tissue culture method. However, the dendrobium officinale has a lot of wild resources and good and uneven quality, and the development of the dendrobium officinale industry is slow due to the unclear breeding target, the lagged research on breeding technology and the lack of good and characteristic new varieties of dendrobium officinale in the market.
Plant protoplasts refer to the part of a cell that is left bare or surrounded by a plasma membrane after removal of the plant cell wall. The research on plant protoplasts dates back to 60 th of the 20 th century, and British phytologists Cocking degraded cell walls by an enzymolysis method to obtain tomato root tip protoplasts. In 1970, Nagata and Table report that tobacco mesophyll protoplasts are separated and cultured to obtain regenerated plants for the first time, and then more and more plastid regenerated plants are successful, and the research types are continuously increased, including forest trees, ornamental plants, medicinal plants, fruit trees and crops. Protoplasts have particular advantages because they lose the cell wall barrier. Protoplasts can provide good experimental materials for cell biology, developmental biology, cell physiology, cytogenetics and other biological subjects; the exchange of genetic materials can be solved at a cell level by the biomass culture and cell fusion, and excellent genetic characters of the plant kingdom are widely recombined by the fusion of heterologous protoplasts, so that a new way is opened up for plant breeding; protoplasts can directly take exogenous DNA, organelles, viruses, plasmids and the like, and are ideal receptors for genetic transformation research. Therefore, plant protoplasts have a wide range of applications in modifying plant genetics, improving crop varieties, and basic theory of biology.
Obtaining a large amount of complete and viable protoplasts is the primary condition for the success of protoplast culture. The enzymolysis separation method is a protoplast separation method which is generally adopted at present. The selection of plant material, the pretreatment process, the use of enzyme preparations, etc., all can affect the separation effect of protoplasts. Among dendrobium plants, methods for separating protoplasts of dendrobium officinale, dendrobium huoshanense, dendrobium nobile and the like have been reported, and the protoplasts can be obtained more effectively.
The protoplast after separation and purification can be cultured under a proper culture medium and culture conditions to regenerate a new cell wall, then the cell can be cracked, and further a new cell mass and callus can be formed, and finally the plant can be regenerated from the callus through an organogenesis way or an embryoid way. Protoplast culture is also affected by a number of factors, including the plant material genotype, the initial density of the protoplast culture, the culture medium, the culture method, the concentration of osmostabilizers, hormones, light, temperature, and other environmental factors. At present, the culture result of the dendrobium plant protoplast is generally poor, the best result is that a cell mass is formed only after several divisions, the cell mass forming frequency is extremely low, and callus or embryoid is difficult to further form. Therefore, on the basis of the predecessors, the cultivation research of the dendrobium officinale protoplast is developed, the bottleneck problem encountered by the predecessors is hoped to be solved, the division frequency of the dendrobium officinale protoplast is improved, and a foundation is laid for establishing the researches of fusion of the dendrobium officinale protoplast, genetic transformation and the like.
Disclosure of Invention
In order to solve the problems in the prior art, a culture medium capable of improving the cell division frequency of the dendrobium officinale protoplast is discovered through years of practice, and the culture medium has important significance for establishing a complete dendrobium officinale protoplast culture system.
The invention solves the technical problems through the following technical scheme:
a culture medium for improving the cell division frequency of a dendrobium officinale protoplast comprises the following components:
inorganic salts: KNO31600~1800mg/L，CaCl2·2H2O 700~900mg/L，MgSO4·7H2O 320~380mg/L，NH4Cl 85~105mg/L，KH2PO4130~150mg/L，H3BO31.1~1.5mg/L，KI 0.20~0.24mg/L，Na2MoO4·2H2O 0.13~0.17mg/L，CoCl2·6H2O 0.013~0.017mg/L，MnSO4·4H2O 10.9~11.3mg/L，ZnSO4·7H2O 4.1~4.4mg/L，CuSO4·5H2O 0.012~0.016mg/L， FeSO4·7H2O 13.7~14.1mg/L，Na2-EDTA 18.4~19.0mg/L；
Organic components: thiamine hydrochloride 0.5-1.0 mg/L, pyridoxine hydrochloride 0.5-1.0 mg/L, nicotinic acid 0.5-1.0 mg/L, D-biotin 0.1-0.5 mg/L, glycine 4.0-6.0 mg/L, proline 80-100 mg/L, citric acid 35-45 mg/L, inositol 90-110 mg/L, casein hydrolysate 130-150 mg/L, and glutathione 60-80 mg/L;
sugar alcohols: 150-200 mg/L of rhamnose, 200-250 mg/L of cellobiose, 80-100 mg/L of ribose, 15-20 g/L of sucrose and 0.5-0.7 mol/L of mannitol;
other components: 0.4-0.6 mg/L of 2,4-D, 0.9-1.1 mg/L of NAA, 0.9-1.1 mg/L of 6-BA, 3.0-3.6 mg/L of sodium selenite, 10-14 mg/L of L-carnitine, 0.04-0.06 mg/L of 5-aminolevulinic acid, 1.8-2.2 mmol/L of sodium butyrate, 30-50 mL/L of dendrobium officinale leaching liquor and 20-40 mL/L of corn juice.
Further, the pH of the culture medium is adjusted to 5.5-6.0.
Further, the preparation method of the dendrobium officinale leaching liquor in the culture medium comprises the following steps: cleaning fresh Dendrobium officinale leaves, wiping, weighing, pulping into slurry by using a homogenizer, preparing a leaching solution according to the mass ratio of the Dendrobium officinale to distilled water of 1: 5, standing for 24 h at 4 ℃ in the dark, and filtering by using 4 layers of gauze to obtain the Dendrobium officinale leaching solution mother liquor.
Further, the medium was sterilized by suction filtration through a 0.22 μm filter.
The invention has the beneficial effects that: the invention takes the dendrobium officinale as a research material, and explores a special culture medium suitable for culturing the dendrobium officinale protoplast from several aspects of inorganic salt components, organic components, sugar alcohol components and other components. The culture medium of the invention adjusts the concentration of each inorganic salt component, especially the concentration of calcium salt and ammonium salt, on the basis of the conventional culture medium, thereby improving the stability of protoplast and reducing death; adding proline, citric acid, hydrolyzed casein, glutathione, etc.; the plant growth regulator is optimized by different concentration combinations; in addition, the components of rhamnose, cellobiose, ribose, sodium selenite, L-carnitine, 5-aminolevulinic acid, sodium butyrate, dendrobium officinale leaching liquor and corn juice are added, the components can be used as precursor substances for cell wall synthesis, induction factors for promoting cell division and the like, the culture effect of the dendrobium officinale protoplast is obviously influenced, and the cell division frequency and the cell cluster formation frequency are obviously improved.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention more comprehensible, specific embodiments thereof are described in detail below with reference to examples of the specification.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, but the present invention may be practiced in other ways than those specifically described and will be readily apparent to those of ordinary skill in the art without departing from the spirit of the present invention, and therefore the present invention is not limited to the specific embodiments disclosed below.
Furthermore, reference herein to "one embodiment" or "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one implementation of the invention. The appearances of the phrase "in one embodiment" in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments.
Example 1
The method for separating and culturing the dendrobium officinale protoplast comprises the following steps:
(1) pretreatment: before enzymolysis, the tender leaves of the dendrobium officinale sterile test-tube plantlets are put into a 26% sucrose solution of CPW for pretreatment for 1 h.
(2) Protoplast separation: getCutting 1.0g of the above leaves into strips with a width of about 0.5-1 mm, transferring into sterile enzyme solution containing 10mL (1.0% cellulase, 0.4% pectinase and 10mmol/L CaCl)2·2H2O+0.1%MES+0.7mmol/LKH2PO4+0.5mol/L mannitol, pH 5.8), gently shaking to make the leaf strips and enzyme solution fully contact, then placing on a shaking table with 50r/min rotation speed, and carrying out enzymolysis for 6h at 27 ℃ in the dark.
(3) And (3) purifying protoplasts: and filtering the mixed solution by using a 400-mesh screen after enzymolysis, centrifuging the filtrate at 800r/min for 3-5 min, and removing the supernatant. Protoplasts were settled on the bottom of the tube, the supernatant was discarded and the pellet was resuspended in a solution of CPW in 13% mannitol. And (3) adding a 26% sucrose solution of CPW into another sterile centrifuge tube, adding the protoplast suspension onto the surface of the 26% sucrose solution of CPW, centrifuging at 300r/min for 3min, and gently sucking out the protoplast zone between the two liquid surfaces of sucrose and mannitol.
(4) Protoplast culture, adjusting the density of purified protoplast to 2.0 × 10 with protoplast culture medium52mL of the cells/mL of the cells were aspirated and placed in a 6 cm-diameter petri dish, sealed with Parafilm, and subjected to shallow liquid culture at 23 ℃ under 150lx continuous light, and the frequency of cell division was counted under an inverted microscope at 2 weeks, after which fresh hypotonic protoplast medium (mannitol concentration 0.025 mol/L) was added a little bit a week, and the frequency of cell pellet formation was counted at 4 weeks of culture.
The protoplast culture medium consists of the following components: inorganic salts: KNO31700mg/L，CaCl2·2H2O 800mg/L，MgSO4·7H2O 350mg/L，NH4Cl 95mg/L，KH2PO4140mg/L，H3BO31.3mg/L，KI 0.22mg/L，Na2MoO4·2H2O 0.15mg/L，CoCl2·6H2O 0.015mg/L，MnSO4·4H2O 11.1mg/L，ZnSO4·7H2O4.3mg/L，CuSO4·5H2O 0.014mg/L， FeSO4·7H2O 13.9mg/L，Na2-EDTA 18.7 mg/L; organic components: thiamine hydrochloride 0.75mg/L, pyridoxine hydrochloride 0.75mgL, 0.75mg/L of nicotinic acid, 0.3mg/L of D-biotin, 5.0mg/L of glycine, 90mg/L of proline, 40mg/L of citric acid, 100mg/L of inositol, 140mg/L of hydrolyzed casein and 70mg/L of glutathione; sugar alcohols: 175mg/L of rhamnose, 225mg/L of cellobiose, 90mg/L of ribose, 17.5g/L of sucrose and 0.6mol/L of mannitol; other components: 0.5mg/L of 2,4-D, 1.0mg/L of NAA, 1.0mg/L of 6-BA, 3.3mg/L of sodium selenite, 12mg/L of L-carnitine, 0.05mg/L of 5-aminolevulinic acid, 2.0mmol/L of sodium butyrate, 40mL/L of dendrobium officinale leaching solution, 30mL/L of corn juice, 5.8 of pH value, and carrying out suction filtration and sterilization by a 0.22 mu m filter membrane. The preparation method of the dendrobium officinale leaching liquor in the culture medium comprises the following steps: cleaning fresh Dendrobium officinale leaves, wiping, weighing, pulping into slurry by using a homogenizer, preparing a leaching solution according to the mass ratio of the Dendrobium officinale to distilled water of 1: 5, standing for 24 h at 4 ℃ in the dark, and filtering by using 4 layers of gauze to obtain the Dendrobium officinale leaching solution mother liquor.
Influence of certain components in protoplast culture medium on culturing of dendrobium officinale protoplast
1. Influence of Dendrobium officinale leach liquor concentration on protoplast culture
Under the condition that other components are not changed, the protoplast culture media with the concentrations of the dendrobium officinale leaching liquor being 20, 40, 60 and 80ml/L are respectively prepared, the protoplast culture media without the dendrobium officinale leaching liquor is used as a control group, then the liquid shallow layer static culture is carried out on the dendrobium officinale protoplast according to the method, and the results are shown in table 1.
As can be seen from Table 1, compared with the control group, the addition of the leaching solution of Dendrobium officinale with the tested concentration can increase the cell division frequency and the cell cluster formation frequency of the protoplast of Dendrobium officinale, when the concentration of the leaching solution of Dendrobium officinale is 40ml/L, the cell division frequency reaches 27.64%, the cell cluster formation frequency reaches 5.99%, which are the highest values, and when the concentration of the leaching solution of Dendrobium officinale exceeds 40ml/L, the cell division frequency and the cell cluster formation frequency show a decreasing trend. The addition of the leaching solution of Dendrobium officinale Kimura et Migo can increase the division frequency of the protoplast cells of Dendrobium officinale Kimura et Migo probably because it can provide other natural nutrients required by the growth and division of the protoplast, and some components of the leaching solution can not be provided or are not provided by artificial culture medium.
2. Effect of sodium butyrate concentration on protoplast culture
Under the condition that other components are not changed, protoplast culture media with sodium butyrate concentrations of 0.5, 1.0, 2.0 and 5.0mmol/L are respectively prepared, the protoplast culture media without sodium butyrate are used as a control group, and then the dendrobium officinale protoplast is subjected to liquid shallow layer standing culture according to the method, and the results are shown in table 2.
As can be seen from Table 2, compared with the control group, the addition of the sodium butyrate with the test concentration can promote the cell division of the protoplast of the dendrobium officinale, the culture effect is the best when the concentration of the sodium butyrate is 2.0mmol/L, the cell division frequency can reach 28.40%, and the cell mass formation frequency can reach 6.07%, which are the highest values.
Effect of 3.5-Aminolevulinic acid concentration on protoplast culture
Under the condition that other components are not changed, protoplast culture media with the concentrations of 5-aminolevulinic acid of 0.01, 0.05, 0.1 and 0.5mg/L are respectively prepared, the protoplast culture media without adding 5-aminolevulinic acid are used as a control group, and then the liquid shallow layer standing culture is carried out on the dendrobium officinale protoplast according to the method, and the results are shown in table 3.
As can be seen from Table 3, the effect of 5-aminolevulinic acid on the culture effect of the dendrobium officinale protoplast is remarkably influenced, the culture effect is best when the concentration of 5-aminolevulinic acid is 0.05mg/L, the cell division frequency can reach 28.17%, the cell mass formation frequency can reach 6.10%, and the highest values are obtained. As the concentration of the 5-aminolevulinic acid continues to increase, the cell division frequency and the cell mass formation frequency of the protoplast inversely decrease, and when the concentration of the 5-aminolevulinic acid is 0.5mg/L, the cell division frequency and the cell mass formation frequency are lower than those of a control group, which indicates that the 5-aminolevulinic acid with too high concentration is not beneficial to the growth and division of the dendrobium officinale protoplast.
4. Effect of different media on protoplast culture
The protoplast culture medium of the invention and the protoplast culture medium of the prior art are respectively adopted to carry out liquid shallow layer static culture on the dendrobium officinale protoplast according to the method, thereby comparing the influence of different culture media on the protoplast culture, and the results are shown in table 4.
A protoplast culture medium in the prior art consists of the following components: 1/10MS (1/10 of MS as macroelements and iron salts, unchanged other components), NaH2PO4150mg/L，CaCl2111mg/L，KH2PO4100mg/L, 150mg/L of asparagine, 150mg/L of glutamine, 200mg/L of arginine, 2mg/L of glycine, 250mg/L of hydrolyzed milk protein, 50mg/L of ascorbic acid, 50mg/L of glutathione, 0.10% MES, NAA2mg/L, 6-BA0.5mg/L, 0.06mol/L of sucrose and 0.44mol/L of mannitol. The culture medium is derived from Zhanzhong root, xu process, Zhangming, and the separation and culture research of the dendrobium officinale mesophyll protoplast.
As can be seen from Table 4, after the protoplast culture medium of the invention is adopted for culture, the cell division frequency of the dendrobium officinale protoplast can reach 28.35 percent, which is 2.6 times of that of the prior art, and the cell mass formation frequency can reach 6.12 percent, which is 6.8 times of that of the prior art, and is remarkably improved. The protoplast culture is influenced by various factors such as the type of culture medium, the culture mode, the density of the protoplasts, exogenous growth regulators and the like, and the type of the culture medium is a key factor influencing the culture effect of the protoplasts. Among the components of the medium, inorganic salts are the main constituent, and among them, calcium salts and ammonium salts have a large influence on the culture of protoplasts. The culture medium of the invention adjusts the concentration of each inorganic salt component, especially the concentration of calcium salt and ammonium salt, on the basis of the conventional culture medium, thereby improving the stability of protoplast and reducing death. The combination of 2,4-D, NAA and 6-BA is selected in the aspect of plant growth regulator, and the optimal concentration ratio is screened. In addition, the culture medium is also added with rhamnose, cellobiose, ribose, sodium selenite, L-carnitine, 5-aminolevulinic acid, sodium butyrate, dendrobium officinale leaching liquor, corn juice and the like, which can be used as precursor substances for cell wall synthesis, induction factors for promoting cell division and the like, and the culture effect of the dendrobium officinale protoplast is obviously influenced, so that the cell division frequency and the cell mass formation frequency are obviously improved.
Claims (4)
1. A culture medium for improving the cell division frequency of a dendrobium officinale protoplast is characterized by comprising the following components:
inorganic salts: KNO31600~1800mg/L，CaCl2·2H2O 700~900mg/L，MgSO4·7H2O 320~380mg/L，NH4Cl 85~105mg/L，KH2PO4130~150mg/L，H3BO31.1~1.5mg/L，KI 0.20~0.24mg/L，Na2MoO4·2H2O 0.13~0.17mg/L，CoCl2·6H2O 0.013~0.017mg/L，MnSO4·4H2O 10.9~11.3mg/L，ZnSO4·7H2O 4.1~4.4mg/L，CuSO4·5H2O 0.012~0.016mg/L， FeSO4·7H2O 13.7~14.1mg/L，Na2-EDTA 18.4~19.0mg/L；
Organic components: thiamine hydrochloride 0.5-1.0 mg/L, pyridoxine hydrochloride 0.5-1.0 mg/L, nicotinic acid 0.5-1.0 mg/L, D-biotin 0.1-0.5 mg/L, glycine 4.0-6.0 mg/L, proline 80-100 mg/L, citric acid 35-45 mg/L, inositol 90-110 mg/L, casein hydrolysate 130-150 mg/L, and glutathione 60-80 mg/L;
sugar alcohols: 150-200 mg/L of rhamnose, 200-250 mg/L of cellobiose, 80-100 mg/L of ribose, 15-20 g/L of sucrose and 0.5-0.7 mol/L of mannitol;
other components: 0.4-0.6 mg/L of 2,4-D, 0.9-1.1 mg/L of NAA, 0.9-1.1 mg/L of 6-BA, 3.0-3.6 mg/L of sodium selenite, 10-14 mg/L of L-carnitine, 0.04-0.06 mg/L of 5-aminolevulinic acid, 1.8-2.2 mmol/L of sodium butyrate, 30-50 mL/L of dendrobium officinale leaching liquor and 20-40 mL/L of corn juice.
2. The culture medium for increasing the cell division frequency of the dendrobium officinale protoplast according to claim 1, wherein the pH of the culture medium is adjusted to 5.5-6.0.
3. The culture medium for increasing the cell division frequency of the protoplast of dendrobium officinale according to claim 1, wherein the leaching solution of dendrobium officinale in the culture medium is prepared by the following steps: cleaning fresh Dendrobium officinale leaves, wiping, weighing, pulping into slurry by using a homogenizer, preparing a leaching solution according to the mass ratio of the Dendrobium officinale to distilled water of 1: 5, standing for 24 h at 4 ℃ in the dark, and filtering by using 4 layers of gauze to obtain the Dendrobium officinale leaching solution mother liquor.
4. The culture medium for increasing the cell division frequency of the dendrobium officinale protoplast according to claim 1, wherein the culture medium needs to be sterilized by suction filtration through a 0.22 μm filter membrane.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010476002.3A CN111685038A (en) | 2020-05-29 | 2020-05-29 | Culture medium for improving cell division frequency of dendrobium officinale protoplast |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010476002.3A CN111685038A (en) | 2020-05-29 | 2020-05-29 | Culture medium for improving cell division frequency of dendrobium officinale protoplast |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111685038A true CN111685038A (en) | 2020-09-22 |
Family
ID=72478882
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010476002.3A Pending CN111685038A (en) | 2020-05-29 | 2020-05-29 | Culture medium for improving cell division frequency of dendrobium officinale protoplast |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111685038A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113462633A (en) * | 2021-08-16 | 2021-10-01 | 广西大学 | Method for separating and extracting protoplast of sugarcane young leaf in different development stages |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105794647A (en) * | 2016-04-10 | 2016-07-27 | 沈阳农业大学 | Method for culturing Brassica rapa microspores with sodium butyrate-NaB as histone deacetylase inhibitor |
-
2020
- 2020-05-29 CN CN202010476002.3A patent/CN111685038A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105794647A (en) * | 2016-04-10 | 2016-07-27 | 沈阳农业大学 | Method for culturing Brassica rapa microspores with sodium butyrate-NaB as histone deacetylase inhibitor |
Non-Patent Citations (4)
| Title |
|---|
| SYEDA JABUN NAHAR AND KAZUHIKO SHIMASAKI: "Application of 5-aminolevulinic Acid for the in vitro Micropropagation of Cymbidium as a Potential Novel Plant Regulator", 《ENVIRON. CONTROL BIOL.》 * |
| YUPHIN KHENTRY ET AL.: "Protoplast Isolation and Culture of Dendrobium Sonia "Bom 17"", 《KASETSART J. (NAT. SCI.)》 * |
| 宋士清等: "5-氨基乙酰丙酸的生理作用及其在农业生产中的应用(综述)", 《河北科技师范学院学报》 * |
| 詹忠根等: "铁皮石斛叶肉原生质体的分离与培养研究", 《浙江大学学报（理学版）》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113462633A (en) * | 2021-08-16 | 2021-10-01 | 广西大学 | Method for separating and extracting protoplast of sugarcane young leaf in different development stages |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Shahin et al. | Cassava mesophyll protoplasts: Isolation, proliferation, and shoot formation | |
| CN101695283B (en) | Culture medium for tissue culture of pedicel buds of oncidium hybridum and tissue cultured seedling propagating method | |
| CN110396495B (en) | Himalayan mirabilis jalapa callus, proliferation method and suspension cell propagation method | |
| CN111471640A (en) | Separation and culture method of honeysuckle protoplast and special culture medium | |
| CN110506635B (en) | Marigold pollen induction culture medium and induction culture method | |
| EP0380692B1 (en) | Plant tissue culture process | |
| CN102703422A (en) | Culture method for new pogostemon cablin breed | |
| CN111685038A (en) | Culture medium for improving cell division frequency of dendrobium officinale protoplast | |
| CN113455400B (en) | Inducing method for anther callus of dragon boat | |
| CN109392721B (en) | Method for inducing sweet potato plant regeneration | |
| CN102134561A (en) | Culture medium used for fast breeding tissue-culture root-shaped stems of nervilia fordii | |
| CN102505005A (en) | Culture medium for keeping primary airway epithelial cells in physiological state in vivo | |
| Sutter | Micropropagation of Ixia viridifolia and a Gladiolus× Homoglossum hybrid | |
| CN1240269C (en) | Polygonatum cyrtonema Hua regeneration system establishment method | |
| Bellincampi et al. | Evidence for the presence of a second conditioning factor in plant cell cultures | |
| EP1649743B1 (en) | Method of direct regeneration, Shikonin induction in callus and Agrobacterium rhizogenes-mediated genetic transformation of Arnebia hispidissima | |
| CN103283592B (en) | Method for direct regeneration of adventitious bud from Jatropha curcas petiole explant | |
| CN106929464B (en) | A method of promote sterol effective constituents in the lobate layer bacterium of currant to accumulate using plant source cigarette water | |
| CN108243951B (en) | Tissue culture method of oldenlandia diffusa | |
| Yamakawa et al. | Application of feeder layer method for improved colony formation of grape cells and protoplasts at low cell density | |
| Otani et al. | Mesophyll protoplast culture of sweet potato (Ipomoea batatas L.) | |
| US5300128A (en) | Method of plant tissue culture | |
| CN110278875A (en) | A kind of method of orchardgrass children fringe in vitro culture regeneration plant | |
| CN112106665A (en) | Culture medium for one-step seedling formation of amorphophallus rivieri and application and tissue culture method | |
| CN110604053A (en) | In-vitro culture rapid propagation method of holly |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20200922 |