CN105660393A - Ginseng leafbud induction medium and culture method - Google Patents
Ginseng leafbud induction medium and culture method Download PDFInfo
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- CN105660393A CN105660393A CN201610011309.XA CN201610011309A CN105660393A CN 105660393 A CN105660393 A CN 105660393A CN 201610011309 A CN201610011309 A CN 201610011309A CN 105660393 A CN105660393 A CN 105660393A
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- culture medium
- bud inducement
- folium ginseng
- outer implant
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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
- A01H4/008—Methods for regeneration to complete plants
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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
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- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
Belonging to the technical field of ginseng tissue culture, the invention provides a ginseng leafbud induction medium and a culture method. The medium takes an MS medium as the basic medium, and is added with 2ip (isopentennyladenine), NAA (naphthalene acetic acid), PVP (polyvinylpyrrolidone), agar powder and white granulated sugar. The culture method comprises four steps: explant treatment, inoculation, dark culture and conventional culture. The ginseng leafbud induction medium and the culture method provided by the invention have the advantages of high induction rate and low browning rate.
Description
Technical field
The invention belongs to ginseng tissue culture technique field, particularly relate to a kind of Folium Ginseng bud inducement culture medium, further relate to a kind of Folium Ginseng bud inducement cultural method.
Background technology
Radix Ginseng is Angiospermae Dicotyledoneae Archichlamydeae Araliaceae, herbaceos perennial, it is grown on thick forest, it is distributed in Heilungkiang, Jilin, Liaoning and northern Hebei remote mountains more, containing more than 10 kinds of ginsenosides, and the fast alcohol of Radix Ginseng, several amino acids and vitamin, it is famous and precious Chinese medicinal herbs.
Fast-propagation many employings tissue culture technique of Radix Ginseng, but the outer implant inductivity of existing tissue culture technique is low, can not meet the requirement of daily production, scientific research and species conservation.
Summary of the invention
There are the problems referred to above based on prior art, the present invention provides a kind of Folium Ginseng bud inducement culture medium, it is culture medium based on MS culture medium, and be added with 2ip(isopentennyladenine), NAA (naphthalene acetic acid), PVP(polyvinylpyrrolidone), agar powder, white sugar, also provide for a kind of Folium Ginseng bud inducement and cultivate method for inducing and cultivating, it includes the process of outer implant, inoculation, light culture and four steps of cellar culture, Folium Ginseng bud inducement culture medium provided by the invention and the advantage that cultural method has inductivity height, melting brown rate is low.
A kind of Folium Ginseng bud inducement culture medium, it is culture medium based on MS culture medium, and be added with 2ip(isopentennyladenine), NAA (naphthalene acetic acid), PVP(polyvinylpyrrolidone), agar powder, white sugar, 2ip(isopentennyladenine) can promote that cell division breaks up, coordinate NAA (naphthalene acetic acid) that leaf bud can be induced better to form callus, PVP(polyvinylpyrrolidone) it is the specificity adsorbent of aldehydes matter, the aldehydes matter of absorption Radix Ginseng leaf bud secretion, it is possible to Browning control.
2ip(isopentennyladenine) the concentration that concentration is 2-8mg/L, NAA (naphthalene acetic acid) be 0.6-1.0mg/L, PVP(polyvinylpyrrolidone) concentration be 0.6-1.5mg/L, agar powder concentration be 8-12g/L, white sugar concentration be 25-35g/L.
2ip(isopentennyladenine) the concentration that concentration is 4-6mg/L, NAA (naphthalene acetic acid) be 0.7-0.9mg/L, PVP(polyvinylpyrrolidone) concentration be 0.9-1.2mg/L, agar powder concentration be 9-11g/L, white sugar concentration be 28-32g/L.
The pH value of Folium Ginseng bud inducement culture medium is 5.5-76.5.
The pH value of Folium Ginseng bud inducement culture medium is 5.8, and slightly acidic culture environment is more suitable for Folium Ginseng bud inducement, it is also possible to suppressing aldehydes matter generation oxidation reaction, composition ginseng callus brownization is dead.
A kind of Folium Ginseng bud inducement cultural method, it comprises the following steps:
The outer implant of S10 processes: gathers the leaf bud of normal growth from the Radix Ginseng maternal plant of healthy growth as outer implant, carries out outer implant sterilization, stripping and slicing;
S20 inoculates: be inoculated in Folium Ginseng bud inducement culture medium by the outer implant block obtained in step S10, spaced 1-1.5 centimetre of every piece of outer implant;
S30 light culture: the outer implant block of Radix Ginseng complete for step S20 inoculation is placed in dark surrounds and is cultivated 2-3 days, and cultivation temperature is 23-27 DEG C;
S40 cellar culture: outer for Radix Ginseng implant block is placed in growth cabinet after terminating and cultivates by light culture, and condition of culture is as follows,
Light application time is 12/12 hour (illumination/dark), and intensity of illumination is 1800-2200lux, and cultivation temperature is 23-27 DEG C.
Step S10 also includes the following sub-step of step:
Step S11 sterilizes: by gathering, the bud tap water returned is clean, it is 74-76% alcohol-pickled 30 seconds by concentration, take out, with aseptic water washing 3-4 time, re-use the mercuric chloride immersion that concentration is 0.1-0.12% to steep 5-10 minute, constantly jiggle during immersion, with aseptic water washing 3-4 time;
The outer implant stripping and slicing of step S12: the wound part excision that will contact with mercuric chloride liquid, and remaining part is cut into the outer implant block that 3-5 millimeter is square.
Detailed description of the invention
Below in conjunction with specific embodiment, the invention will be further described.
Embodiment one: preparation Folium Ginseng bud inducement culture medium
MS culture medium prescription according to table 1 and the Folium Ginseng bud inducement culture medium prescription shown in table 2, prepare Folium Ginseng bud inducement culture medium of the present invention.
Weigh each composition in MS culture medium and 2ip, NAA, PVP, agar powder, white sugar with balance respectively, be placed in triangular flask, add appropriate purified water, stirring and dissolving, be settled to 1000mL, and regulate pH value to 5.8. Triangular flask is sealed, is placed in high-pressure sterilizing pot, in 121 DEG C of sterilizing 21min, then in superclean bench, by Folium Ginseng bud inducement culture medium subpackage to tissue culture flasks, natural cooling, by tissue culture's bottle closure after its solidification, standby.
The composition of table 1MS culture medium and consumption thereof.
The composition of table 2 Folium Ginseng bud inducement culture medium and consumption thereof.
Embodiment two: the test of Folium Ginseng bud inducement culture medium of the present invention
MS culture medium prescription according to table 1 and the Folium Ginseng bud inducement culture medium prescription shown in table 3, with reference to the medium preparation method described in embodiment one, prepare the Folium Ginseng bud inducement culture medium of each test group, matched group respectively.
Table 3 Folium Ginseng bud inducement culture medium.
The leaf bud of normal growth is gathered as outer implant from the Radix Ginseng maternal plant of healthy growth, by gathering, the bud tap water returned is clean, it is placed in superclean bench, is 75% alcohol-pickled 30 seconds by concentration, take out, with aseptic water washing 4 times, re-use mercuric chloride immersion that concentration is 0.1% to steep 8 minutes, constantly jiggle during immersion, then with aseptic water washing 4 times, the wound part excision that will contact with mercuric chloride liquid, and remaining part is cut into outer implant block 5 millimeters square.
Outer implant block is inoculated into above-mentioned prepare each test group, matched group Folium Ginseng bud inducement culture medium in, spaced 1.2 centimetres of every piece of outer implant, after inoculation, outer for Radix Ginseng implant block is placed in dark surrounds and cultivates 2 days, cultivation temperature is 25 DEG C.
Outer for Radix Ginseng implant block is placed in growth cabinet after terminating and cultivates by light culture, condition of culture is as follows: light application time is 12/12 hour (illumination/dark), and intensity of illumination is 2000lux, and cultivation temperature is 25 DEG C, adding up inductivity and melting brown rate after cultivating 25 days, statistical result is as shown in table 4.
Table 4 Folium Ginseng bud inducement culture medium test result.
Note: comprehensive grading y=inductivity b*0.7-melting brown rate a*0.3 in table.
By the result of the test of table 4 it can be seen that adopt Folium Ginseng bud inducement culture medium of the present invention, it is possible to significantly improve Folium Ginseng bud induction rate, and reduce its melting brown rate. Contrast from matched group 3 and test group 5 and can be seen that Folium Ginseng bud inducement tissue culture is had the effect of certain Browning control by PVP, it is possible to melting brown rate is reduced to less than 15%; Compare from matched group 1,2 and test group 5 and can be seen that 2ip and NAA has better facilitation effect with the use of more any than being used alone; The consumption of PVP will not increase along with consumption more than Browning control effect after 0.1mg/L and increase as can be seen from the test results simultaneously; The consumption of 2ip and NAA does not only result in too much and better induces facilitation effect, affects promotion inducing action because consumption is too many on the contrary.
Claims (7)
1. a Folium Ginseng bud inducement culture medium, it is characterised in that it is culture medium based on MS culture medium, and is added with 2ip(isopentennyladenine), NAA (naphthalene acetic acid), PVP(polyvinylpyrrolidone), agar powder, white sugar.
2. a kind of Folium Ginseng bud inducement culture medium according to claim 1, it is characterized in that, 2ip(isopentennyladenine) the concentration that concentration is 2-8mg/L, NAA (naphthalene acetic acid) be 0.6-1.0mg/L, PVP(polyvinylpyrrolidone) concentration be 0.6-1.5mg/L, agar powder concentration be 8-12g/L, white sugar concentration be 25-35g/L.
3. a kind of Folium Ginseng bud inducement culture medium according to claim 1, it is characterized in that, 2ip(isopentennyladenine) the concentration that concentration is 4-6mg/L, NAA (naphthalene acetic acid) be 0.7-0.9mg/L, PVP(polyvinylpyrrolidone) concentration be 0.9-1.2mg/L, agar powder concentration be 9-11g/L, white sugar concentration be 28-32g/L.
4. according to one of them described a kind of Folium Ginseng bud inducement culture medium of claims 1 to 3, it is characterised in that the pH value of described Folium Ginseng bud inducement culture medium is 5.5-6.5.
5. a kind of Folium Ginseng bud inducement culture medium according to claim 4, it is characterised in that the pH value of described Folium Ginseng bud inducement culture medium is 5.8.
6. a Folium Ginseng bud inducement cultural method, it is characterised in that it comprises the following steps:
The outer implant of S10 processes: gathers the leaf bud of normal growth from the Radix Ginseng maternal plant of healthy growth as outer implant, carries out outer implant sterilization, stripping and slicing;
S20 inoculates: be inoculated in Folium Ginseng bud inducement culture medium by the outer implant block obtained in step S10, spaced 1-1.5 centimetre of every piece of outer implant;
S30 light culture: the outer implant block of Radix Ginseng complete for step S20 inoculation is placed in dark surrounds and is cultivated 2-3 days, and cultivation temperature is 23-27 DEG C;
S40 cellar culture: outer for Radix Ginseng implant block is placed in growth cabinet after terminating and cultivates by light culture, and condition of culture is as follows,
Light application time is 12/12 hour (illumination/dark), and intensity of illumination is 1800-2200lux, and cultivation temperature is 23-27 DEG C.
7. a kind of Folium Ginseng bud inducement cultural method according to claim 6, it is characterised in that described step S10 also includes following sub-step:
Step S11 sterilizes: by gathering, the bud tap water returned is clean, it is 74-76% alcohol-pickled 30 seconds by concentration, take out, with aseptic water washing 3-4 time, re-use the mercuric chloride immersion that concentration is 0.1-0.12% to steep 5-10 minute, constantly jiggle during immersion, with aseptic water washing 3-4 time;
The outer implant stripping and slicing of step S12: the wound part excision that will contact with mercuric chloride liquid, and remaining part is cut into the outer implant block that 3-5 millimeter is square.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106069770A (en) * | 2016-07-08 | 2016-11-09 | 徐伟明 | A kind of promote the tissue culture medium (TCM) of saponin content in Radix Ginseng |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101156533A (en) * | 2007-11-21 | 2008-04-09 | 中国科学院西北高原生物研究所 | Method for planting India swertiamarin artificially in warmhouse |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101156533A (en) * | 2007-11-21 | 2008-04-09 | 中国科学院西北高原生物研究所 | Method for planting India swertiamarin artificially in warmhouse |
Non-Patent Citations (5)
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| 于丽杰: "《植物组织培养教程》", 30 April 2015, 华中科技大学出版社 * |
| 安永辉等: "人参的组织培养及快繁体系的建立", 《安徽农业科学》 * |
| 曹孜义: "《果树花卉无毒苗快繁技术》", 30 October 1998, 甘肃科学技术出版社 * |
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| 郑彩霞: "《植物生理学 第3版》", 31 July 2013, 中国林业出版社 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106069770A (en) * | 2016-07-08 | 2016-11-09 | 徐伟明 | A kind of promote the tissue culture medium (TCM) of saponin content in Radix Ginseng |
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