CN103563741B - A kind of aseptic detoxification cultural method of the dendrobium candidum capsule embryo that ftractures - Google Patents
A kind of aseptic detoxification cultural method of the dendrobium candidum capsule embryo that ftractures Download PDFInfo
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Abstract
A kind of aseptic detoxification cultural method of the dendrobium candidum capsule embryo that ftractures, it is characterized in that first the stem of noble dendrobium outstanding achievement surface of having ftractureed being cleared up, then 10 ~ 15min is irradiated as superclean bench medium ultraviolet, then aseptically embryo is transferred in aseptic cillin bottle, with the alcohol-pickled cleaning 30 ~ 60s of 65 ~ 75%, then by 160 ~ 200 order stainless steel aseptic filter screen, embryo is leached, the aseptic straw of 0.5 ~ 1.5% antiformin is had by embryo with being transferred in aseptic cillin bottle again with suction, after submergence sterilization 10 ~ 15min, embryo is leached to be transferred in aseptic cillin bottle with sterile water and repeatedly rinse 3 ~ 5 times, with aseptic straw, embryo droplet mode is transferred to aseptic seeding in medium to cultivate.Cultural method of the present invention is simple and practical, strong operability, and embryo availability is high, germination rate is high, solves the industry difficult problem that cracking stem of noble dendrobium capsule is difficult to utilize, substantially prolongs the sowing time limit of stem of noble dendrobium embryo.
Description
Technical field
The present invention relates to a kind of cultural method of orchid embryo, particularly relate to a kind of aseptic detoxification cultural method of the dendrobium candidum capsule embryo that ftractures.
Background technology
Dendrobium candidum (DendrobiumofficinaleKimuraetMigo) is the famous and precious rare traditional Chinese medicine of tradition, has reinforcing stomach reg fluid, the unique effects such as nourishing Yin and clearing heat, conventional with the symptom such as consumption of body fluid caused by febrile disease, after being ill abnormal heat etc. are many.Modern pharmacological research show dendrobium candidum treatment diabetes, immunological regulation, antitumor etc. in have good curative effect.Dendrobium candidum medical value, health value are high, but because of dendrobium officinale nature fruit-setting rate extremely low, in addition under its seed natural conditions, germination rate is extremely low, oneself is very rare for its wild resource, the domestic artificial culture for dendrobium candidum obtains certain progress now, and in production, artificial culture carries out increment cultivation mainly through the capsule of dendrobium candidum.
Dendrobium candidum generally starts to bloom in mid or late April, and mid or late May is full-bloom stage.The capsule successfully of pollinating entered growing of 120d, and capsule becomes yellow green, is mature on the whole.Along with the increase of maturity, scattering and disappearing of moisture, the automatic yellow embryo that sheds that ftractures of capsule meeting.The holding time of current capsule can only reach about 30d.Present stage cannot carry out aseptic detoxification sowing for the cracking embryo that is scattered, and the capsule such as, adopted when the patent No. 200310112012.5,200810104220.3,200910036502.9,201110203343.4 etc. carry out the process of aseptic seeding detoxification is uncracked dendrobium candidum capsule.Because dendrobium candidum fruit-setting rate is not high, capsule life cycle is short in addition, and therefore the presence or absence of stem of noble dendrobium capsule has become the deciding factor of present stage restriction dendrobe tissue culture industry value chain.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of aseptic detoxification cultural method of the dendrobium candidum capsule embryo that ftractures, and the method is simple and practical, strong operability, and availability is high, germination rate is high.Solve the industry difficult problem that cracking stem of noble dendrobium capsule is difficult to utilize, substantially prolongs the sowing time limit of stem of noble dendrobium embryo, for the smooth running of dendrobe tissue culture factory provides guarantee.
The present invention solves the problems of the technologies described above adopted technical scheme: a kind of Sterile culture method of the dendrobium candidum capsule embryo that ftractures, is characterized in that comprising the following steps:
1) the dendrobium candidum capsule of cracking is carried out removing surface;
2) cracking of removing surface fruit pod or embryo are carried out detoxification process:
Embryo is placed in superclean bench medium ultraviolet and irradiates 10 ~ 15min, then aseptically embryo is transferred in aseptic cillin bottle, use the alcohol-pickled cleaning 30 ~ 60s of mass percentage concentration 65 ~ 75% afterwards, then with aseptic filter screen, embryo is leached, the aseptic straw of mass percentage concentration 0.5 ~ 1.5% antiformin is had by embryo with being transferred in aseptic cillin bottle again with suction, after submergence sterilization 10 ~ 15min, the aseptic filter screen of embryo is leached, then with sterile water, embryo is transferred in aseptic cillin bottle, repeatedly rinses 3 ~ 5 times with sterile water;
3) finally embryo to be transferred in sowing medium in droplet mode with sterile water and to carry out aseptic seeding cultivation.
As preferably, described step 2) in the concentration of alcohol be mass percentage concentration 70%, need upset mixing shake in alcohol-pickled process.
As preferably, described step 2) described in aseptic filter screen adopt the aseptic filter screen of 160 ~ 200 order stainless steel.
Preferred again, described step 2) in the mass percentage concentration of antiformin be 1%.
Finally, the detailed process of described step 3) is: be transferred in sowing medium in droplet mode by the embryo of detoxification process with sterile water, : 1/3 ~ 1/2MS liquid nutrient medium+methyl α-naphthyl acetate (NAA) 0.5 ~ 1.0mg/L+6-benzyl aminoadenine (BA) 1.0 ~ 2.0mg/L+ active carbon (AC) 0.5 ~ 1.5g/L(art-recognized meanings is that in liquid nutrient medium, large content concentration of element reduces to 1/3 ~ 1/2, and add material below, make concentration of NAA 0.5 ~ 1.0mg/L in liquid nutrient medium, 6-benzyl aminoadenine concentration 1.0 ~ 2.0mg/L, concentration of activated carbon 0.5 ~ 1.5g/L), after planting after light culture 3-5d, be placed in illumination 1000 ~ 1500LUX, temperature 25 ± 1 DEG C, cultivate under the condition of light application time 12-14h/d.
Compared with prior art, the invention has the advantages that: cultural method of the present invention is simple and practical, strong operability, embryo availability is high, germination rate is high, efficiently solve the industry difficult problem that cracking stem of noble dendrobium capsule is difficult to utilize, substantially prolongs the sowing time limit of stem of noble dendrobium embryo, for the smooth running of dendrobe tissue culture factory provides guarantee.
Embodiment
Below in conjunction with embodiment, the present invention is described in further detail.
Embodiment one:
The dendrobium candidum capsule of cracking is carried out removing surface.The cracking of removing surface fruit pod or embryo powder are placed in superclean bench medium ultraviolet and irradiate 10min.Superclean bench art-recognized meanings is that the cleanliness factor of workbench reaches hundred grades (highest levels in current bacterization equipment), and the clump count namely in every cubic metre is hundred, general indoor cleanliness factor be 1,000,000 grades namely every cubic space clump count be 1,000,000.Then aseptically embryo is transferred in aseptic cillin bottle, afterwards with 70% alcohol-pickled cleaning 30s, then by 160 order stainless steel aseptic filter screen, embryo is leached, the aseptic straw of 1% antiformin is had by embryo with being transferred in aseptic cillin bottle again with suction, submergence sterilization 10min, the embryo aseptic filter screen of 160 order stainless steel is leached, is then transferred in aseptic cillin bottle with sterile water, repeatedly rinses 3 times with sterile water.In droplet mode, the embryo of detoxification process is transferred in sowing medium with sterile water, sowing medium is: 1/2MS+NAA0.5mg/L+BA1.0mg/L+AC0.5g/L, after planting by after cultivation 3d, be placed in illumination 1000-1500LUX, temperature 25 ± 1 DEG C, cultivates under the condition of light application time 12h/d, cultivates after 30 days, find that germination rate is 98%, pollution rate is 1.0%.
Embodiment two:
The cracking of removing surface fruit pod or embryo powder are placed in superclean bench medium ultraviolet and irradiate 12min.Then aseptically embryo is transferred in aseptic cillin bottle, afterwards with 70% alcohol-pickled cleaning 45s, then by 180 order stainless steel aseptic filter screen, embryo is leached, the aseptic straw of 1% antiformin is had by embryo with being transferred in aseptic cillin bottle again with suction, submergence sterilization 12min, the embryo aseptic filter screen of 180 order stainless steel is leached, is then transferred in aseptic cillin bottle with sterile water, repeatedly rinses 4 times with sterile water.In droplet mode, the embryo of detoxification process is transferred in sowing medium with sterile water, sowing medium is: 1/2MS+NAA0.8mg/L+BA1.5mg/L+AC1.0g/L, after planting by after cultivation 4d, be placed in illumination 1000-1500LUX, temperature 25 ± 1 DEG C, cultivates under the condition of light application time 13h/d, cultivates after 30 days, find that germination rate is 96%, pollution rate is 0.8%.
Embodiment three:
The cracking of removing surface fruit pod or embryo powder are placed in superclean bench medium ultraviolet and irradiate 15min.Then aseptically embryo is transferred in aseptic cillin bottle, afterwards with 70% alcohol-pickled cleaning 60s, then by 200 order stainless steel aseptic filter screen, embryo is leached, the aseptic straw of 1% antiformin is had by embryo with being transferred in aseptic cillin bottle again with suction, submergence sterilization 15min, the embryo aseptic filter screen of 200 order stainless steel is leached, is then transferred in aseptic cillin bottle with sterile water, repeatedly rinses 5 times with sterile water.In droplet mode, the embryo of detoxification process is transferred in sowing medium with sterile water, sowing medium is: 1/2MS+NAA1.0mg/L+BA2.0mg/L+AC1.5g/L, after planting by after cultivation 5d, be placed in illumination 1000-1500LUX, temperature 25 ± 1 DEG C, cultivates under the condition of light application time 14h/d, cultivates after 30 days, statistics finds that germination rate is 92%, and pollution rate is 0.5%.
Claims (4)
1. ftracture the aseptic detoxification cultural method of dendrobium candidum capsule embryo, it is characterized in that comprising the following steps:
1) the dendrobium candidum capsule of cracking is carried out removing surface;
2) cracking of removing surface fruit pod or embryo are carried out detoxification process:
Embryo is placed in superclean bench medium ultraviolet and irradiates 10 ~ 15min, then aseptically embryo is transferred in aseptic cillin bottle, afterwards with the alcohol-pickled cleaning 30-60s of mass percentage concentration 65 ~ 75%, then with aseptic filter screen, embryo is leached, the aseptic straw of mass percentage concentration 0.5 ~ 1.5% antiformin is had to be transferred in aseptic cillin bottle by embryo with suction again, after submergence sterilization 10 ~ 15min, the aseptic filter screen of embryo is leached, then with sterile water, embryo is transferred in aseptic cillin bottle, repeatedly rinses 3 ~ 5 times with sterile water;
3) finally embryo to be transferred in sowing medium in droplet mode with sterile water and to carry out aseptic seeding cultivation; Step 3) detailed process be: the embryo of detoxification process is transferred in sowing medium with sterile water in droplet mode, sowing medium is: 1/3 ~ 1/2MS liquid nutrient medium+methyl α-naphthyl acetate 0.5 ~ 1.0mg/L+6-benzyl aminoadenine 1.0 ~ 2.0mg/L+ active carbon 0.5 ~ 1.5g/L, after planting after light culture 3 ~ 5d, be placed in illumination 1000-1500LUX, temperature 25 ± 1 DEG C, cultivates under the condition of light application time 12 ~ 14h/d.
2. aseptic detoxification cultural method according to claim 1, is characterized in that described step 2) in the concentration of alcohol be mass percentage concentration 70%, need upset mixing shake in alcohol-pickled process.
3. aseptic detoxification cultural method according to claim 1, is characterized in that described step 2) described in aseptic filter screen adopt the aseptic filter screen of 160-200 order stainless steel.
4. aseptic detoxification cultural method according to claim 1, is characterized in that described step 2) in the mass percentage concentration of antiformin be 1%.
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| CN106171969B (en) * | 2015-05-07 | 2018-07-10 | 泉州正和堂生物科技有限公司 | A kind of clone's culture technique of dendrobium candidum embryo particle |
| CN110651559A (en) * | 2019-11-07 | 2020-01-07 | 安徽祥枫生物科技有限公司 | Preservation method of dendrobium huoshanense capsules for tissue culture seedling |
Citations (2)
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| CN1608424A (en) * | 2004-11-24 | 2005-04-27 | 上海增靓生物科技有限公司 | Large-area cultivation of officinal dendrobium |
| CN101849503A (en) * | 2009-10-26 | 2010-10-06 | 刘宏源 | Method for manufacturing dendrobium officinale artificial seeds |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1608424A (en) * | 2004-11-24 | 2005-04-27 | 上海增靓生物科技有限公司 | Large-area cultivation of officinal dendrobium |
| CN101849503A (en) * | 2009-10-26 | 2010-10-06 | 刘宏源 | Method for manufacturing dendrobium officinale artificial seeds |
Non-Patent Citations (5)
| Title |
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| Micropropagation of an orchid Dendrobium strongylanthum Rchb.f.;Kong, Q. etal;《International Journal of Horticultural Science》;20071231;第13卷(第1期);第61-64页 * |
| Paudel etal.In vitro plant regeneration of Esmeralda clarkei Rchb.f. via protocorm explant.《Full Length Research Paper 》.2012,第11卷(第54期),第11704-11708页. * |
| Shreeti Pradhan etal.In vitro seed germination in Cymbidium elegans Lindl. And Dendrobium densiflorum Lindl. ex Wall. (Orchidaceae).《Botanica Orientalis -Journal of Plant Science》.2009,(第6期),第100页左栏最后一段，右栏第1段. * |
| Witaya Pimda etal.Cryopreservation of Dendrobium heterocarpum.《ELBA Bioflux》.2010,第2卷(第1期),第7-14页. * |
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