CN104255463A - Rapid propagation method of suspension cells of Epilobium angustifolium Linn. - Google Patents
Rapid propagation method of suspension cells of Epilobium angustifolium Linn. Download PDFInfo
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- CN104255463A CN104255463A CN201410463061.1A CN201410463061A CN104255463A CN 104255463 A CN104255463 A CN 104255463A CN 201410463061 A CN201410463061 A CN 201410463061A CN 104255463 A CN104255463 A CN 104255463A
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- callus
- willow herb
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- 239000000725 suspension Substances 0.000 title claims abstract description 18
- 238000000034 method Methods 0.000 title claims abstract description 14
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- 244000046536 Chamerion angustifolium subsp angustifolium Species 0.000 title 1
- 206010020649 Hyperkeratosis Diseases 0.000 claims abstract description 24
- 235000008302 Chamaenerion angustifolium Nutrition 0.000 claims description 20
- 239000002609 medium Substances 0.000 claims description 15
- 230000009514 concussion Effects 0.000 claims description 10
- 238000005286 illumination Methods 0.000 claims description 10
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- 238000005406 washing Methods 0.000 claims description 10
- 238000009395 breeding Methods 0.000 claims description 9
- 230000001488 breeding effect Effects 0.000 claims description 8
- 102000004190 Enzymes Human genes 0.000 claims description 5
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- 241000031003 Monascus ruber Species 0.000 claims description 5
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- 230000006698 induction Effects 0.000 claims description 5
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- 229960002523 mercuric chloride Drugs 0.000 claims description 5
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- 235000015097 nutrients Nutrition 0.000 claims description 5
- 238000005070 sampling Methods 0.000 claims description 5
- 239000008399 tap water Substances 0.000 claims description 5
- 235000020679 tap water Nutrition 0.000 claims description 5
- 239000007077 tomato juice medium Substances 0.000 claims description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- 241000721098 Epilobium Species 0.000 claims 7
- 241000646858 Salix arbusculoides Species 0.000 claims 1
- 239000006194 liquid suspension Substances 0.000 claims 1
- 235000015193 tomato juice Nutrition 0.000 claims 1
- 244000103926 Chamaenerion angustifolium Species 0.000 abstract description 17
- 238000004519 manufacturing process Methods 0.000 abstract description 6
- 230000009286 beneficial effect Effects 0.000 abstract 1
- 239000006285 cell suspension Substances 0.000 abstract 1
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- 238000005516 engineering process Methods 0.000 description 3
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- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 2
- 240000009089 Quercus robur Species 0.000 description 2
- 235000011471 Quercus robur Nutrition 0.000 description 2
- 229960002246 beta-d-glucopyranose Drugs 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- LNTHITQWFMADLM-UHFFFAOYSA-N gallic acid Chemical compound OC(=O)C1=CC(O)=C(O)C(O)=C1 LNTHITQWFMADLM-UHFFFAOYSA-N 0.000 description 2
- 229920001864 tannin Polymers 0.000 description 2
- 235000018553 tannin Nutrition 0.000 description 2
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- PZIRUHCJZBGLDY-UHFFFAOYSA-N Caffeoylquinic acid Natural products CC(CCC(=O)C(C)C1C(=O)CC2C3CC(O)C4CC(O)CCC4(C)C3CCC12C)C(=O)O PZIRUHCJZBGLDY-UHFFFAOYSA-N 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 description 1
- ATJXMQHAMYVHRX-CPCISQLKSA-N Ellagic acid Natural products OC1=C(O)[C@H]2OC(=O)c3cc(O)c(O)c4OC(=O)C(=C1)[C@H]2c34 ATJXMQHAMYVHRX-CPCISQLKSA-N 0.000 description 1
- 229920002079 Ellagic acid Polymers 0.000 description 1
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- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 1
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- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- WURBFLDFSFBTLW-UHFFFAOYSA-N benzil Chemical group C=1C=CC=CC=1C(=O)C(=O)C1=CC=CC=C1 WURBFLDFSFBTLW-UHFFFAOYSA-N 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
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- 229940074393 chlorogenic acid Drugs 0.000 description 1
- CWVRJTMFETXNAD-JUHZACGLSA-N chlorogenic acid Chemical compound O[C@@H]1[C@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-JUHZACGLSA-N 0.000 description 1
- FFQSDFBBSXGVKF-KHSQJDLVSA-N chlorogenic acid Natural products O[C@@H]1C[C@](O)(C[C@@H](CC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O FFQSDFBBSXGVKF-KHSQJDLVSA-N 0.000 description 1
- 235000001368 chlorogenic acid Nutrition 0.000 description 1
- BMRSEYFENKXDIS-KLZCAUPSSA-N cis-3-O-p-coumaroylquinic acid Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)cc2)[C@@H]1O)C(=O)O BMRSEYFENKXDIS-KLZCAUPSSA-N 0.000 description 1
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- FAARLWTXUUQFSN-UHFFFAOYSA-N methylellagic acid Natural products O1C(=O)C2=CC(O)=C(O)C3=C2C2=C1C(OC)=C(O)C=C2C(=O)O3 FAARLWTXUUQFSN-UHFFFAOYSA-N 0.000 description 1
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to a rapid propagation method for culture of suspension cells of Epilobium angustifolium Linn. The rapid propagation method comprises the steps of inducing callus from Epilobium angustifolium Linn., subculturing the callus, culturing suspension cells, optimizing a cell suspension system, etc. The suspension cells of Epilobium angustifolium Linn. prepared by the method have the advantages of being simple in technological process and short in production cycle, and being not influenced by natural environment factors such as terrain, seasons and climate, thus being beneficial to large-scale industrialized production of the suspension cells of Epilobium angustifolium Linn.
Description
Technical field
The present invention relates to the quick-breeding method of willow herb suspension cell, belong to plant technology field.
Background technology
Willow herb,
epilobium angustifolium Linn., Oenotheraceae, perennial herb, height about 1-1.3 rice.Stem is upright, usually not branch.Leaf alternate, lanceolar, raceme top is raw, and extend, rhachis is by pubescence, and bract is linear, long 1-2 centimetre, Hua great, both sexes, reddish violet.Root, stem, leaf, flower, fruit, all containing tannin, are distributed in Southwestern China, northwest, North China to northeast; North temperate zone blazons, and the higher border of height above sea level, woodland, patana, riverbank thick grass and baked wheaten cake or cutting blank, to Japan, are born in naturally in North America, Europe.Cold-resistant.Happiness is nice and cool, humid climate and moistening, fertile, well-drained soil.Slightly resistance to the moon.The environment that fear is hot, arid.9 tannin classes and other phenoloids is got in willow herb herb 70% acetone extract; be accredited as 3-oxygen-galloyl-D-Glucose, 1 respectively; Ma Su, shrimp-roe florigen, chlorogenic acid, gallic acid in 6-bis--oxygen-galloyl-β-D-glucopyranose, 1-oxygen-galloyl-4,6-hexahydroxy dibenzoyl base-β-D-glucopyranose, throatroot D prime, English oak (Quercus robur) ellagic acid, spy.Root-like stock or all herbal medicine, slightly poisonous, energy regulating menstruation and activating blood, swelling and pain relieving, cure mainly irregular menstruation, fracture, joint sprain.Willow herb breeding adopts seminal propagation, also can plant division and cottage propagation, and suspension cell culture is unmanned research also, and suspension cell has simple to operate controlled, and production cost is low, reproduction rate advantages of higher.
Summary of the invention
Technical problem to be solved by this invention is to provide the method for quickly breeding of willow herb suspension cell culture, and simple to operate controlled, production cost is low, reproduction rate advantages of higher.
For solving the problems of the technologies described above, the present invention adopts following technical proposal:
Select the seed of willow herb, 30min is soaked in suds, tap water 1h, with the mercuric chloride sterilization 15min of 0.2% on superclean bench, aseptic water washing 5 times is to noresidue solution, explant after sterile-processed, remove exosper to be seeded in 1/2WPM+NAA0.1-0.2mg/L+6-BA0.5-1mg/L medium and to carry out callus induction, the condition of evoked callus is that half-light shines, illumination 10h/d, temperature 21 DEG C, after 30 days, by the callus access medium WPM+1-2mg/L6-BA+0.3-0.5mg/L2 derived, squamous subculture is carried out in 4-D+150mg/L tomato juice medium, squamous subculture is after 40 days, select light green loose callus, put into the cultivation liquid nutrient medium that addition of 150mg/L Monascus ruber enzyme, the bead added after sterilization jolts, be placed in concussion incubator and carry out level concussion cultivation, amplitude 2-4cm, vibration frequency 90r/min, temperature 23 DEG C, illumination 2500lx, photophase 12h/d, 7-8 days subcultures once, sampling, washing, dry.
Adopt willow herb suspension cell prepared by the present invention, technological process is simple, with short production cycle, and not by region, in season, the advantages such as factor of natural environment impact such as weather, are conducive to the large-scale industrialized production of setting up willow herb suspension cell.
Below in conjunction with embodiment, the present invention is further elaborated, but the scope of protection of present invention is not limited to following embodiments.
Embodiment
Embodiment 1
Select the seed of willow herb, 30min is soaked in suds, tap water 1h, with the mercuric chloride sterilization 15min of 0.2% on superclean bench, aseptic water washing 5 times is to noresidue solution, explant after sterile-processed, remove exosper to be seeded in 1/2WPM+NAA0.1mg/L+6-BA0.5mg/L medium and to carry out callus induction, the condition of evoked callus is that half-light shines, illumination 10h/d, temperature 21 DEG C, after 30 days, by the callus access medium WPM+1mg/L6-BA+0.3mg/L2 derived, squamous subculture is carried out in 4-D+150mg/L tomato juice medium, squamous subculture is after 40 days, select light green loose callus, put into the cultivation liquid nutrient medium that addition of 150mg/L Monascus ruber enzyme, the bead added after sterilization jolts, be placed in concussion incubator and carry out level concussion cultivation, amplitude 2-4cm, vibration frequency 90r/min, temperature 23 DEG C, illumination 2500lx, photophase 12h/d, 7-8 days subcultures once, sampling, washing, dry, survival rate is 90%.
Embodiment 2
Select the seed of willow herb, 30min is soaked in suds, tap water 1h, with the mercuric chloride sterilization 15min of 0.2% on superclean bench, aseptic water washing 5 times is to noresidue solution, explant after sterile-processed, remove exosper to be seeded in 1/2WPM+NAA0.2mg/L+6-BA1mg/L medium and to carry out callus induction, the condition of evoked callus is that half-light shines, illumination 10h/d, temperature 21 DEG C, after 30 days, by the callus access medium WPM+2mg/L6-BA+0.5mg/L2 derived, squamous subculture is carried out in 4-D+150mg/L tomato juice medium, squamous subculture is after 40 days, select light green loose callus, put into the cultivation liquid nutrient medium that addition of 150mg/L Monascus ruber enzyme, the bead added after sterilization jolts, be placed in concussion incubator and carry out level concussion cultivation, amplitude 2-4cm, vibration frequency 90r/min, temperature 23 DEG C, illumination 2500lx, photophase 12h/d, 7-8 days subcultures once, sampling, washing, dry, survival rate is 91%.
Embodiment 3
Select the seed of willow herb, 30min is soaked in suds, tap water 1h, with the mercuric chloride sterilization 15min of 0.2% on superclean bench, aseptic water washing 5 times is to noresidue solution, explant after sterile-processed, remove exosper to be seeded in 1/2WPM+NAA0.2mg/L+6-BA1mg/L medium and to carry out callus induction, the condition of evoked callus is that half-light shines, illumination 10h/d, temperature 21 DEG C, after 30 days, by the callus access medium WPM+2mg/L6-BA+0.3mg/L2 derived, squamous subculture is carried out in 4-D+150mg/L tomato juice medium, squamous subculture is after 40 days, select light green loose callus, put into the cultivation liquid nutrient medium that addition of 150mg/L Monascus ruber enzyme, the bead added after sterilization jolts, be placed in concussion incubator and carry out level concussion cultivation, amplitude 2-4cm, vibration frequency 90r/min, temperature 23 DEG C, illumination 2500lx, photophase 12h/d, 7-8 days subcultures once, sampling, washing, dry, survival rate is 92%.
Claims (5)
1. the method for quickly breeding of a willow herb suspension cell, it is characterized in that: the seed of willow herb is sterile-processed, remove exosper to be seeded in 1/2WPM+NAA0.1-0.2mg/L+6-BA0.5-1mg/L medium and to carry out callus induction, after 30 days, by the callus access medium WPM+1-2mg/L6-BA+0.3-0.5mg/L2 derived, squamous subculture is carried out in 4-D+150mg/L tomato juice medium, squamous subculture accesses and addition of the cultivation carrying out liquid suspension cell in 150mg/L Monascus ruber enzyme for 40 days afterwards, 7-8 days subcultures once, sampling, washing, dry.
2. the method for quickly breeding of a kind of willow herb suspension cell according to claim 1, it is characterized in that: described disinfect as the seed selecting willow herb, 30min is soaked in suds, tap water 1h, with the mercuric chloride sterilization 15min of 0.2% on superclean bench, aseptic water washing 5 times is to noresidue solution.
3. the method for quickly breeding of a kind of willow herb suspension cell according to claim 1, is characterized in that: the condition of evoked callus is half-light photograph, illumination 10h/d, temperature 21 DEG C.
4. the method for quickly breeding of a kind of willow herb suspension cell according to claim 1, is characterized in that: with the addition of tomato juice in willow herb callus subculture, can promote the merisis of cell.
5. the method for quickly breeding of a kind of willow herb suspension cell according to claim 1, it is characterized in that: the cultural method of suspension cell is that callus is after squamous subculture, select light green loose callus, put into liquid nutrient medium, the bead added after sterilization jolts, be placed in concussion incubator and carry out level concussion cultivation, amplitude 2-4cm, vibration frequency 90r/min, temperature 23 DEG C, illumination 2500lx, photophase 12h/d.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410463061.1A CN104255463A (en) | 2014-09-12 | 2014-09-12 | Rapid propagation method of suspension cells of Epilobium angustifolium Linn. |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201410463061.1A CN104255463A (en) | 2014-09-12 | 2014-09-12 | Rapid propagation method of suspension cells of Epilobium angustifolium Linn. |
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| CN104255463A true CN104255463A (en) | 2015-01-07 |
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| CN201410463061.1A Pending CN104255463A (en) | 2014-09-12 | 2014-09-12 | Rapid propagation method of suspension cells of Epilobium angustifolium Linn. |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104719163A (en) * | 2015-03-25 | 2015-06-24 | 温州科技职业学院 | Method for increasing acclimatization planting percent of blueberry tissue culture test-tube seedlings |
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2014
- 2014-09-12 CN CN201410463061.1A patent/CN104255463A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104719163A (en) * | 2015-03-25 | 2015-06-24 | 温州科技职业学院 | Method for increasing acclimatization planting percent of blueberry tissue culture test-tube seedlings |
| CN104719163B (en) * | 2015-03-25 | 2017-04-05 | 温州科技职业学院 | A kind of method for improving blueberry tissue culture test tube seedling seedling exercising planting percent |
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| PB01 | Publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20150107 |
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| WD01 | Invention patent application deemed withdrawn after publication |