CN107494260A - STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution - Google Patents
STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution Download PDFInfo
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- CN107494260A CN107494260A CN201710537325.7A CN201710537325A CN107494260A CN 107494260 A CN107494260 A CN 107494260A CN 201710537325 A CN201710537325 A CN 201710537325A CN 107494260 A CN107494260 A CN 107494260A
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- stevia rebaudiana
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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
- 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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- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution, concretely comprise the following steps：Urea, potassium nitrate, ammonium nitrate, potassium dihydrogen phosphate, zinc chloride, calcium chloride, potassium sulfate, trace element, sucrose, glucose, bactericide, growth regulator, vitamin, amino acid, water by weight, it is well mixed, adjust pH, sterilizing, preserved after cooling, produce STEVIA REBAUDIANA bud differentiation nutrient solution.Have the beneficial effect that：The culture liquid energy of the present invention meets the nutritional need in each stage in STEVIA REBAUDIANA bud differentiation incubation, effectively accelerates bud fission into cell mass, and then is divided into embryoid, forms callus, then is divided into plant, improves differentiation culture efficiency；Inorganic nutrients are sufficient for the needs of STEVIA REBAUDIANA sprout cell on each Stage Nutrition and physiologically in nutrient solution；The culture liquid energy dissipation pathogenic microorganisms, increase the disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud.
Description
Technical field
The present invention relates to nutrient solution technical field, more particularly to the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution.
Background technology
" Herba Hedyotis cantonensis ", South America Paraguay is originated in, drunk always by local resident as sweet drink for centuries, be
A kind of small-sized perennial composite family (Compositae) Si Taiwei Subgenus, perennial root, fibrous root type, perennial, short-day, draft
Plant.Japan in 1970 introduces STEVIA REBAUDIANA from Brazil, starts to tame, cultivates, prepares glucosides, while carries out toxicity, food inspection
The experiment such as survey, and STEVIA REBAUDIANA product-stevioside is developed first；China started by Nanjing Zhong Shan plants in 1976
The R＆D institutions such as garden, the Chinese Academy of Agricultural Sciences successively introduce STEVIA REBAUDIANA from Japan and plant experimentally success.It is present Jiangsu, Fujian, Shandong, new
There are a large amount of plantations on the ground such as boundary, Henan, Anhui, and for the gross area up to more than 1,000,000 mu, China turns into plants STEVIA REBAUDIANA face in the world
The most country of product, and stevioside maximum producing country and exported country in the world.Due to the stevioside contained by STEVIA REBAUDIANA
Glycosides has blood glucose processed, reduces blood pressure, enhances metabolism and treat diabetes, obesity, regulation hydrochloric acid in gastric juice, recovers neural fatigue
Effect, and physics, chemical property are stable, and no Fermented feature, its product has the long shelf-life, particularly suitable as sugar
Sweetener, products and health products additive agent of patient's food such as disease, hypertension etc. are urinated, there are wide market prospects and application.
For STEVIA REBAUDIANA to be selfed not affine cross-pollinatd plant, genetic stability is poor, is unfavorable for homozygosis and the anti-miscellaneous guarantor of kind of cenospecies
Pure, moreover, Stevia seed is tiny, germination percentage is low, only can not meet the needs of in the market is to STEVIA REBAUDIANA by seminal propagation.Using
Tissue culture technique can not only keep the good characteristic of kind, can not also be influenceed to breed throughout the year by external condition.20th century
The seventies, domestic many research units have just carried out the research of STEVIA REBAUDIANA tissue cultures.But all it is with fast numerous for mesh mostly
Experiment, using the stem apex with bud point, lateral bud etc., it expands, and numerous coefficient is limited, and growth coefficient is low.
Prior art such as Authorization Notice No. is the B of CN 103766222 Chinese invention patent, discloses the training of STEVIA REBAUDIANA tissue
The method of supporting and its culture medium, thickening inducing culture includes MS+ benzyladenine 6-BA 0.1~1.0mg/L+ methyl α-naphthyl acetates NAA
0.1~0.5mg/L+0.7% agar, budding seedling inducing culture include：MS+ methyl α-naphthyl acetate NAA 0.4~1.0mg/L+ benzyls
Adenine 6-BA 1.0~2.5mg/L+0.7% agar；Rooting induction culture medium include：1/4MS+ indolebutyric acids IBA 0.05
~0.2mg/L+ methyl α-naphthyl acetate NAA 0.05~0.2mg/L+0.5% agar.The culture medium can ensure that blade can reach in each stage
To expected training objective, but each stage culture medium is all different, uses cumbersome.
The content of the invention
It is nutritious comprehensively it is an object of the invention to provide one kind, STEVIA REBAUDIANA cell differentiation can be accelerated, improve differentiation training
Efficiency is supported, increases the disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud, easy to use, the STEVIA REBAUDIANA bud differentiation training of preparation method simple possible
The preparation method of nutrient solution.
The present invention is directed to the problem of being mentioned in above-mentioned technology, and the technical scheme taken is：STEVIA REBAUDIANA bud differentiation nutrient solution
Preparation method, concretely comprise the following steps：
Urea 0.8-1.3 parts, potassium nitrate 0.6-1 parts, ammonium nitrate 0.6-0.8 parts, potassium dihydrogen phosphate 3-5 parts, chlorination by weight
Zinc 0.05-0.08 parts, calcium chloride 0.06-0.09 parts, potassium sulfate 0.06-0.09 parts, micro- 0.003-0.004 parts, sucrose
8-12 parts, glucose 11-14 parts, bactericide 0.001-0.002 parts, growth regulator 0.0003-0.0005 parts, vitamin
0.002-0.003 parts, amino acid 0.001-0.002 parts, water 100-110 parts, it is well mixed, regulation pH is 5.5-6.8, Ran Hou
Sterilize 40-50min at 130-150 DEG C, is put into 2-5 DEG C of environment and preserves after cooling, produces STEVIA REBAUDIANA bud differentiation nutrient solution, should
Contain substantial amounts of nutritional ingredient and trace element in nutrient solution, the battalion in each stage in STEVIA REBAUDIANA bud differentiation incubation can be met
Supporting needs, and effectively accelerates callus differentiation and bud formation again, improves differentiation culture efficiency；In nutrient solution the quantity of inorganic nutrients and
Ratio is suitable, it is sufficient to meets the needs of STEVIA REBAUDIANA cell on each Stage Nutrition and physiologically.
Preferably, bactericide is that Amici reaches and active peptides mixture, its mass ratio are 1:0.05-0.08, activity are more
The amino acid sequence of peptide is HSHACASYCLRCRVLHPGCVNCYRCSR, and it is harmful that the active peptides can destroy mould germ etc.
The cell membrane material of microorganism, change its osmotic pressure, suppress its growth and breeding, or even kill, be advantageous to prevent STEVIA REBAUDIANA bud point
Change the rotten bud phenomenon in incubation；The bactericide wide sterilization spectrum, can not only dissipation pathogenic microorganisms, prevent STEVIA REBAUDIANA bud break up
Infection of the putrefactivebacteria to bud causes rotten bud phenomenon in incubation, moreover it is possible to increase the disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud, and
The bactericide highly effective and safe.
Preferably, growth regulator is 6-BA and KT, and its weight ratio is 1:0.4-0.6, the growth regulator is to induced bud
Differentiation effect is preferable, can effectively accelerate STEVIA REBAUDIANA bud fission into cell mass, and then is divided into embryoid, forms callus, by
Callus is divided into plant again, and inductivity is high, and budding is neat.
Preferably, amino acid is that glycine or proline or alanine or lysine, amino acid are easier to thin by STEVIA REBAUDIANA
Born of the same parents are absorbed, and certain nutritional ingredient is provided for STEVIA REBAUDIANA, promotes it quickly to divide, and are substantially shorter differentiation cultivation cycle.
Compared with prior art, the advantage of the invention is that：Contain substantial amounts of nutritional ingredient in the nutrient solution of the present invention
And trace element, the nutritional need in each stage in STEVIA REBAUDIANA bud differentiation incubation can be met, effectively accelerate callus
Differentiation and bud formation, differentiation culture efficiency is improved, inductivity is high, and budding is neat；The quantity of inorganic nutrients and ratio are suitable in nutrient solution,
It is sufficient for the needs of STEVIA REBAUDIANA cell on each Stage Nutrition and physiologically；The nutrient solution also has bactericidal property, not only
Energy dissipation pathogenic microorganisms, prevent that infection of the putrefactivebacteria to bud causes rotten bud phenomenon in STEVIA REBAUDIANA bud differentiation incubation, also
The disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud can be increased；The culture liquid and preparation method thereof simple possible, is easy to largely produce.
Specific embodiment
The present invention program is described further below by embodiment：
Embodiment 1：
STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution, concretely comprises the following steps：
1 part of urea, 0.8 part of potassium nitrate, 0.7 part of ammonium nitrate, 4 parts of potassium dihydrogen phosphate, 0.06 part of zinc chloride, calcium chloride by weight
0.08 part, 0.07 part of potassium sulfate, trace element 0.0035 part, 10 parts of sucrose, 12 parts of glucose, 0.0015 part of bactericide, growth
0.0004 part of conditioning agent, 0.0025 part of vitamin, 0.0015 part of amino acid, 105 parts of water, it is well mixed, regulation pH is 6, then
The nutrient solution configured is placed in high-pressure steam sterilizing pan, sterilized 45min at 140 DEG C of vapour phase, and 3 DEG C of environment is put into after cooling
Middle preservation, STEVIA REBAUDIANA bud differentiation nutrient solution is produced, contains substantial amounts of nutritional ingredient and trace element in the nutrient solution, sweet tea can be met
The nutritional need in each stage, effectively accelerates bud fission into cell mass, and then be divided into embryo in Ye Juya differentiation incubations
Shape body, callus is formed, plant is divided into by callus again, improve differentiation culture efficiency；Inorganic nutrients in nutrient solution
Quantity and ratio are suitable, it is sufficient to meet the needs of STEVIA REBAUDIANA sprout cell on each Stage Nutrition and physiologically.
Above-mentioned bactericide is that Amici reaches and active peptides mixture, its mass ratio are 1:0.06, the amino of active peptides
Acid sequence is HSHACASYCLRCRVLHPGCVNCYRCSR, and the active peptides can destroy the harmful microbes such as mould germ
Cell membrane material, change its osmotic pressure, suppress its growth and breeding, or even kill, be advantageous to prevent the differentiation of STEVIA REBAUDIANA bud from cultivating
Rotten bud phenomenon in journey；The bactericide wide sterilization spectrum, can not only dissipation pathogenic microorganisms, prevent STEVIA REBAUDIANA bud break up incubation
Infection of the middle putrefactivebacteria to bud causes rotten bud phenomenon, moreover it is possible to increases the disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud, and the bactericide
Highly effective and safe.
Above-mentioned growth regulator is 6-BA and KT, and its weight ratio is 1:0.5, the growth regulator is to induced bud differentiation effect
Preferably, it can effectively accelerate STEVIA REBAUDIANA bud fission into cell mass, and then be divided into embryoid, callus be formed, by callus
Plant is divided into again, and inductivity is high, and budding is neat.
Above-mentioned amino acid is glycine, proline, alanine, the mixture of lysine, and its weight ratio is 1:1:1:1, ammonia
Base acid is easier to be absorbed by STEVIA REBAUDIANA bud, and certain nutritional ingredient is provided for STEVIA REBAUDIANA bud, promotes it quickly to divide, can be big
It is big to shorten differentiation cultivation cycle.
Embodiment 2：
STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution, concretely comprises the following steps：0.9 part of urea, 0.7 part of potassium nitrate, nitre by weight
Sour 0.8 part of ammonium, 3.8 parts of potassium dihydrogen phosphate, 0.08 part of zinc chloride, 0.07 part of calcium chloride, 0.09 part of potassium sulfate, trace element
0.0032 part, 11 parts of sucrose, 12 parts of glucose, 0.0015 part of bactericide, 0.00035 part of growth regulator, vitamin 0.0022
Part, 0.0018 part of amino acid, 100 parts of water, it is well mixed, regulation pH is 5.6, and then sterilize 48min at 132 DEG C, after cooling
It is put into 3 DEG C of environment and preserves, produce STEVIA REBAUDIANA bud differentiation nutrient solution, substantial amounts of nutritional ingredient and micro is contained in the nutrient solution
Element, the nutritional need in each stage in STEVIA REBAUDIANA bud differentiation incubation can be met, effectively accelerate bud fission into cell mass,
And then embryoid is divided into, callus is formed, plant is divided into by callus again, improves differentiation culture efficiency；Nutrient solution
The quantity and ratio of middle inorganic nutrients are suitable, it is sufficient to meet the need of STEVIA REBAUDIANA sprout cell on each Stage Nutrition and physiologically
Will.
Above-mentioned bactericide is that Amici reaches and active peptides mixture, its mass ratio are 1:0.08, the amino of active peptides
Acid sequence is HSHACASYCLRCRVLHPGCVNCYRCSR, and the active peptides can destroy the harmful microbes such as mould germ
Cell membrane material, change its osmotic pressure, suppress its growth and breeding, or even kill, be advantageous to prevent the differentiation of STEVIA REBAUDIANA bud from cultivating
Rotten bud phenomenon in journey；The bactericide wide sterilization spectrum, can not only dissipation pathogenic microorganisms, prevent STEVIA REBAUDIANA bud break up incubation
Infection of the middle putrefactivebacteria to bud causes rotten bud phenomenon, moreover it is possible to increases the disease resistance and resistance against disadvantage of STEVIA REBAUDIANA bud, and the bactericide
Highly effective and safe.
Above-mentioned growth regulator is 6-BA and KT, and its weight ratio is 1:0.4, the growth regulator is to induced bud differentiation effect
Preferably, it can effectively accelerate STEVIA REBAUDIANA bud fission into cell mass, and then be divided into embryoid, callus be formed, by callus
Plant is divided into again, and inductivity is high, and budding is neat.
Above-mentioned amino acid is proline, and amino acid is easier to be absorbed by STEVIA REBAUDIANA bud, provided necessarily for STEVIA REBAUDIANA bud
Nutritional ingredient, promote it quickly to divide, be substantially shorter differentiation cultivation cycle.
Embodiment 3：
STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution, concretely comprises the following steps：1.2 parts of urea, 0.7 part of potassium nitrate, nitre by weight
Sour 0.75 part of ammonium, 4.7 parts of potassium dihydrogen phosphate, 0.058 part of zinc chloride, 0.08 part of calcium chloride, 0.06 part of potassium sulfate, trace element
0.004 part, 9 parts of sucrose, 13 parts of glucose, 0.0017 part of bactericide, 0.00043 part of growth regulator, 0.0024 part of vitamin,
0.0016 part of amino acid, 110 parts of water, bactericide are that mass ratio is 1:0.08 Amici reaches and active peptides mixture, growth
Conditioning agent is that weight ratio is 1:0.4 6-BA and KT mixtures, amino acid is glycine, proline, the mixture of lysine, respectively
It is 6.5 to adjust pH after raw material is well mixed, and then sterilize 40min at 150 DEG C, is put into 5 DEG C of environment and preserves after cooling, i.e.,
Obtain STEVIA REBAUDIANA bud differentiation nutrient solution.
Routine operation in the operating procedure of the present invention is well known to those skilled in the art, herein without repeating.
Technical scheme is described in detail embodiment described above, it should be understood that it is described above only
For the specific embodiment of the present invention, it is not intended to limit the invention, all any modifications made in the spirit of the present invention,
Supplement or similar fashion replacement etc., should be included in the scope of the protection.
SEQUENCE LISTING
<110>Close big vast gardening research and development Co., Ltd in Pujiang County
<120>STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution
<130> 1
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 27
<212> PRT
<213>It is artificial synthesized
<400> 1
His Ser His Ala Cys Ala Ser Tyr Cys Leu Arg Cys Arg Val Leu His
1 5 10 15
Pro Gly Cys Val Asn Cys Tyr Arg Cys Ser Arg
20 25
Claims (9)
1. STEVIA REBAUDIANA bud breaks up the preparation method of nutrient solution, it is characterised in that：The preparation method concretely comprises the following steps：By culture
Formula measures urea, potassium nitrate, ammonium nitrate, potassium dihydrogen phosphate, zinc chloride, calcium chloride, potassium sulfate, trace element, sucrose, grape
Sugar, bactericide, growth regulator, vitamin, amino acid, water, it is well mixed, adjusts pH, sterilize, preserves, produce STEVIA REBAUDIANA bud
Break up nutrient solution.
2. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The nutrient solution
Composition and its parts by weight are：Urea 0.8-1.3 parts, potassium nitrate 0.6-1 parts, ammonium nitrate 0.6-0.8 parts, potassium dihydrogen phosphate 3-5 parts,
Zinc chloride 0.05-0.08 parts, calcium chloride 0.06-0.09 parts, potassium sulfate 0.06-0.09 parts, micro- 0.003-0.004 parts,
Sucrose 8-12 parts, glucose 11-14 parts, bactericide 0.001-0.002 parts, growth regulator 0.0003-0.0005 parts, vitamin
0.002-0.003 parts, amino acid 0.001-0.002 parts, water 100-110 parts.
3. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The bactericide is
Amici reaches and active peptides mixture, and its mass ratio is 1:0.05-0.08.
4. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 3, it is characterised in that：In the bactericide
The amino acid sequence of active peptides is HSHACASYCLRCRVLHPGCVNCYRCSR.
5. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The growth regulating
Agent is that growth regulator is 6-BA and KT, and its weight ratio is 1:0.4-0.6.
6. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The amino acid is
Glycine or proline or alanine or lysine.
7. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The nutrient solution
PH is 5.5-6.8.
8. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：It is described sterilizing be
Sterilize 40-50min at 130-150 DEG C.
9. the preparation method of STEVIA REBAUDIANA bud differentiation nutrient solution according to claim 1, it is characterised in that：The preservation condition
For 2-5 DEG C.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103436483A (en) * | 2013-08-01 | 2013-12-11 | 南京年吉冷冻食品有限公司 | Anther culture solution |
| CN103858757A (en) * | 2012-12-11 | 2014-06-18 | 丰益(上海)生物技术研发中心有限公司 | Stevia rebaudiana bertoni tissue culture method adopting leaf as explant, and special culture medium thereof |
-
2017
- 2017-07-04 CN CN201710537325.7A patent/CN107494260A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103858757A (en) * | 2012-12-11 | 2014-06-18 | 丰益(上海)生物技术研发中心有限公司 | Stevia rebaudiana bertoni tissue culture method adopting leaf as explant, and special culture medium thereof |
| CN103436483A (en) * | 2013-08-01 | 2013-12-11 | 南京年吉冷冻食品有限公司 | Anther culture solution |
Non-Patent Citations (2)
| Title |
|---|
| 夏树让等: "《枣树无公害病虫草害综防技术问答》", 31 December 2007, 中国农业出版社 * |
| 黄大年等: "《水稻转基因研究及其育种》", 31 December 2006, 中国农业科学技术出版社 * |
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Application publication date: 20171222 |