CN104737911A - Quick cultivation method for rhizoma bletillae tissue culture seedlings - Google Patents
Quick cultivation method for rhizoma bletillae tissue culture seedlings Download PDFInfo
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- CN104737911A CN104737911A CN201510143716.1A CN201510143716A CN104737911A CN 104737911 A CN104737911 A CN 104737911A CN 201510143716 A CN201510143716 A CN 201510143716A CN 104737911 A CN104737911 A CN 104737911A
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Abstract
The invention discloses a quick cultivation method for rhizoma bletillae tissue culture seedlings. The method comprises the following steps: selecting intact non-cracked rhizoma bletillae capsules and disinfecting the capsules by 75% alcohol for 30 seconds; then soaking and disinfecting the capsules by using 0.2% mercury bichloride for 10 minutes; washing 6 times by sterile water; fully and thoroughly washing, airing and uniformly spreading on a culture medium to cultivate; and when the seeds germinate to obtain seedlings which are about 2cm long, inoculating 3-4 seedlings as a cluster to a strong sprout culture medium to be cultured. Compared with the conventional tissue culture seedlings obtained by sterile germination of rhizoma bletillae seeds, the method just needs subculture once, so that the time consumed is short and at least three months are saved.
Description
Technical field
The invention belongs to traditional Chinese medicine planting technology field, be specifically related to a kind of fast breeding method of bletilla plantlet in vitro.
Background technology
Bletilla is perennial herb.Pseudobulb block root shape, white, plump, there is finger-like difference.Spend beautiful greatly, aubergine, can view and admire for cultivation.Pseudobulb is containing gum bletilla, starch, volatilization wet goods composition, and have the effects such as convergence, tonifying lung hemostasis, detumescence, oral administration external application, is applied to the diseases such as hemoptysis haematemesis, traumatism and bleeding, sore swollen toxin, chapped skin, pulmonary tuberculosis hemoptysis, ulcer haemorrhage.Bletilla is a kind of wild plant day by day in imminent danger, and the condition of seed germination is harsh, needs and mycosymbiosis, adds that Seedling Stage is longer, to environment sensitive, is therefore difficult to effectively be bred by seed under natural environment situation.Along with the exhaustion of wild resource, the rising all the way of the Bletilla striata medicinal materials market price, has broken through 400 yuan/kg high point.Under stimulation so at high price, finally remaining wild bletilla resource is estimated to exhaust in the near future.Under the ordering about of interests, plantation family start to seek bletilla artificial cultivation, but due to seedling very deficient, be difficult to scale development, even can not start to walk.Therefore, the technical problem of seedling breeding must be solved, just can break the restriction of bletilla seedling bottleneck.
Summary of the invention
The object of the present invention is to provide a kind of fast breeding method of bletilla plantlet in vitro.The method is simple to operate, and the seed germination time is short, the cost-saving and time.
The present invention realizes especially by following technical scheme:
A fast breeding method for bletilla plantlet in vitro, comprises the following steps:
1) select intact bletilla capsule with 75% alcohol-pickled sterilization 30s;
2) with 0.2% mercuric chloride by capsule soaking disinfection 10min, aseptic water washing 6 times, fully rinses well;
3) will disinfect and the capsule dried cut, seed is sown on medium equably, cultivates according to condition of culture;
4) seedling after seed germination having been grown to 2 centimetres is forwarded in strong seedling culture base by 3 ~ 4 one clump and cultivates.
Further, step 3) described in medium prepare by the following method: adopt C medium, add each mother liquor successively, NAA 1.0mg/L, potato 200g/L (stripping and slicing liquor 0.5h, only get juice), banana 40g/L pulls an oar, sucrose 30g/L, agar 4g/L, constant volume, surveys pH value to 6.0.
Further, step 3) described in condition of culture be: 25 ± 2 DEG C, illuminance be in the culturing room of 1500lux, 12h illumination, 12h dark cultivate three months.
Further, step 4) described in strong seedling culture base prepare by the following method: adopt 1/2MS medium, add each mother liquor successively, NAA 0.5mg/L, 6-BA 0.2mg/L, potato tubers making beating directly adds 10g/L, banana making beating 75g/L, sucrose 20g/L, agar 4g/L, active carbon 0.4g/L, constant volume, surveys pH value to 6.0.
Further, step 4) in condition of culture be: 25 ± 2 DEG C, illuminance is 1500lux, 12h illumination, cultivate in the culturing room of 12h dark, pseudobulb to be grown and root can carry out hardening.
Beneficial effect of the present invention is: adopt seed group training, the seedling pollution rate obtained is low, differentiation rate is high, stabilization characteristics of genetics, active principle content is high, compared with the method obtains plantlet in vitro with traditional bletilla seed asepsis sprouting simultaneously, only needs subculture once, used time is short, has saved at least trimestral time.
Embodiment
Below in conjunction with embodiment, the present invention is described further, the following stated, only to preferred embodiment of the present invention, not do other forms of restriction to the present invention, any those skilled in the art may utilize the technology contents of above-mentioned announcement to be changed to the Equivalent embodiments of equal change.Everyly do not depart from the present invention program's content, any simple modification done following examples according to technical spirit of the present invention or equivalent variations, all drop in protection scope of the present invention.
Embodiment 1
One, bletilla seed asepsis sprouting
1, the preparation of medium
Adopt C medium, add each mother liquor successively, NAA 1.0mg/L, potato 200g/L (stripping and slicing liquor 0.5h, only gets juice), banana 40g/L pulls an oar, sucrose 30g/L, agar 4g/L.Constant volume, about surveying pH value to 6.0.
2, bletilla capsule/seed disinfection
This operates on superclean bench and carries out.If bletilla capsule is intact not splitting, the thorough wiping bletilla of the alcohol with 75% capsule, or soak 30s, pour out alcohol; If capsule ftractures, the pouch parcel seed made with highdensity silk stocking, sterilization method is the same.
Use the mercuric chloride of 0.2% by above-mentioned capsule/seed disinfection 10min again, sterile water fully rinses 6 times.Capsule/seed is dried in the air to parching completely on superclean bench.
3, bletilla seed inoculation
Will disinfect and the capsule dried incision, or directly the seed disinfected is sown on medium equably, density appropriateness.
4, cultivate
The seed inoculated is placed on 25 ± 2 DEG C, illuminance be in the culturing room of 1500lux (12 h light, 12 h dark) cultivate three months.
Two, the strong plantlets and rootage of bletilla plantlet in vitro
1, the preparation of Rooting and hardening-off culture base
Adopt 1/2MS medium, add each mother liquor successively, NAA 0.5mg/L, 6-BA 0.2mg/L, potato tubers making beating directly adds 10g/L, banana making beating 75g/L, sucrose 20g/L, agar 4g/L, active carbon 0.4g/L.Constant volume, about surveying pH value to 6.0.
2, inoculate
Seedling seed germination having been grown to 2 cm is forwarded in strong seedling culture base by 3 ~ 4 one clump.
3, cultivate
The seedling inoculated is placed on 25 ± 2 DEG C, illuminance is cultivate in the culturing room of 1500lux (12 h light, 12 h dark).The bletilla having grown pseudobulb and root after three months can carry out hardening.
Embodiment 2
On December 18th, 2013, the experiment of bletilla seed asepsis sprouting is carried out in laboratory, Yunnan University of Traditional Chinese Medicine Yi Jianji building 6301 ~ 6303.Select 10 capsules, sow 400 bottles altogether.Because bletilla seed is tiny, and maturity between capsule is inconsistent, concrete emergence rate can only rough estimate for being greater than 80%.
After cultivating three wheat harvesting periods, on March 25th, 2014, the bletilla seedling growing to 2 ~ 3 centimetres high is gone to squamous subculture in strong seedling culture base by 3 ~ 4 one clump.
After cultivating three wheat harvesting periods, on July 15th, 2014, the bletilla seedling growing pseudobulb and root is delivered to little Shao planting base, Kunming.After the hardening of ten days, plant in ground, cover pine needle and take shade net.After two months, most seedling grows fine, and statistics survival rate is to 92.36%.In January, 2015, dig out bletilla pseudobulb statistics size after seedling, mean size is 1.78 centimetres.
Facts have proved that the enforcement of this method can shorten the cultivation cycle of bletilla, and good result can be reached.
Claims (5)
1. a fast breeding method for bletilla plantlet in vitro, is characterized in that: comprise the following steps:
1) select intact bletilla capsule with 75% alcohol-pickled sterilization 30s;
2) with 0.2% mercuric chloride by capsule soaking disinfection 10min, aseptic water washing 6 times, fully rinses well;
3) will disinfect and the capsule dried cut, seed is sown on medium equably, cultivates according to condition of culture;
4) seedling after seed germination having been grown to 2 centimetres is forwarded in strong seedling culture base by 3 ~ 4 one clump and cultivates.
2. the fast breeding method of a kind of bletilla plantlet in vitro according to claim 1, it is characterized in that: step 3) described in medium prepare by the following method: adopt C medium, add each mother liquor successively, NAA 1.0mg/L, potato juice 200g/L, banana 40g/L pull an oar, sucrose 30g/L, agar 4g/L, constant volume, surveys pH value to 6.0.
3. the fast breeding method of a kind of bletilla plantlet in vitro according to claim 1, it is characterized in that: step 3) described in condition of culture be: 25 ± 2 DEG C, under illuminance is 1500lux condition, cultivate three months in the culturing room of 12h illumination, 12h dark.
4. the fast breeding method of a kind of bletilla plantlet in vitro according to claim 1, it is characterized in that: step 4) described in strong seedling culture base prepare by the following method: adopt 1/2MS medium, add each mother liquor successively, NAA 0.5mg/L, 6-BA 0.2mg/L, potato tubers making beating directly adds 10g/L, banana making beating 75g/L, sucrose 20g/L, agar 4g/L, active carbon 0.4g/L, constant volume, surveys pH value to 6.0.
5. the fast breeding method of a kind of bletilla plantlet in vitro according to claim 1, it is characterized in that: step 4) in condition of culture be: 25 ± 2 DEG C, under illuminance is 1500lux condition, cultivate in the culturing room of 12h illumination, 12h dark, pseudobulb to be grown and root can carry out hardening.
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105145352A (en) * | 2015-08-03 | 2015-12-16 | 河南科技大学 | Efficient tissue culture and rapid propagation technology for seedlings of bletilla striata |
| CN105145361A (en) * | 2015-09-17 | 2015-12-16 | 康美药业（文山）药材种植管理有限公司 | Method for efficiently breeding tissue culture seedlings of bletilla striata and planting method of bletilla striata |
| CN105165368A (en) * | 2015-10-15 | 2015-12-23 | 向君民 | Method for rapidly breeding seedlings by using bletilla striata tubers |
| CN105210875A (en) * | 2015-10-15 | 2016-01-06 | 铜仁市金农绿色农业科技有限公司 | A kind of bletilla striata quick propagation method |
| CN106416972A (en) * | 2016-08-29 | 2017-02-22 | 浙江金石生物科技有限公司 | Method for quickly cultivating bletilla striata seedlings |
| CN107047310A (en) * | 2017-05-10 | 2017-08-18 | 文山学院 | A kind of cultural method of bletilla striata seeds culture seedling |
| CN109169273A (en) * | 2018-09-03 | 2019-01-11 | 云南中医学院 | A kind of breeding method of bletilla proliferation pseudobulb |
| CN110679421A (en) * | 2019-11-06 | 2020-01-14 | 安徽东方金桥农林科技股份有限公司 | Bletilla striata seedling cultivation method |
| CN116649216A (en) * | 2023-06-25 | 2023-08-29 | 江苏护理职业学院 | Pure bletilla striata three-fork tissue culture domesticated seedling raising method |
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105145352A (en) * | 2015-08-03 | 2015-12-16 | 河南科技大学 | Efficient tissue culture and rapid propagation technology for seedlings of bletilla striata |
| CN105145352B (en) * | 2015-08-03 | 2017-06-06 | 河南科技大学 | A kind of bletilla seed height of seedling imitates group culturation rapid propagating technology |
| CN105145361A (en) * | 2015-09-17 | 2015-12-16 | 康美药业（文山）药材种植管理有限公司 | Method for efficiently breeding tissue culture seedlings of bletilla striata and planting method of bletilla striata |
| CN105165368A (en) * | 2015-10-15 | 2015-12-23 | 向君民 | Method for rapidly breeding seedlings by using bletilla striata tubers |
| CN105210875A (en) * | 2015-10-15 | 2016-01-06 | 铜仁市金农绿色农业科技有限公司 | A kind of bletilla striata quick propagation method |
| CN106416972A (en) * | 2016-08-29 | 2017-02-22 | 浙江金石生物科技有限公司 | Method for quickly cultivating bletilla striata seedlings |
| CN107047310A (en) * | 2017-05-10 | 2017-08-18 | 文山学院 | A kind of cultural method of bletilla striata seeds culture seedling |
| CN109169273A (en) * | 2018-09-03 | 2019-01-11 | 云南中医学院 | A kind of breeding method of bletilla proliferation pseudobulb |
| CN110679421A (en) * | 2019-11-06 | 2020-01-14 | 安徽东方金桥农林科技股份有限公司 | Bletilla striata seedling cultivation method |
| CN116649216A (en) * | 2023-06-25 | 2023-08-29 | 江苏护理职业学院 | Pure bletilla striata three-fork tissue culture domesticated seedling raising method |
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Application publication date: 20150701 |