CN107347642A - Culture medium for bletilla pseudobulb cutting tissue culture seedling and tissue culture seedling method thereof - Google Patents
Culture medium for bletilla pseudobulb cutting tissue culture seedling and tissue culture seedling method thereof Download PDFInfo
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- CN107347642A CN107347642A CN201710619723.3A CN201710619723A CN107347642A CN 107347642 A CN107347642 A CN 107347642A CN 201710619723 A CN201710619723 A CN 201710619723A CN 107347642 A CN107347642 A CN 107347642A
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- China
- Prior art keywords
- pseudobulb
- bletilla
- culture
- seedling
- illumination
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Links
- 241001313855 Bletilla Species 0.000 title claims abstract description 119
- 238000000034 method Methods 0.000 title claims abstract description 19
- 239000001963 growth medium Substances 0.000 title claims abstract description 18
- 238000005520 cutting process Methods 0.000 title claims abstract description 16
- 230000004069 differentiation Effects 0.000 claims description 47
- 238000005286 illumination Methods 0.000 claims description 39
- SQGYOTSLMSWVJD-UHFFFAOYSA-N silver(1+) nitrate Chemical compound [Ag+].[O-]N(=O)=O SQGYOTSLMSWVJD-UHFFFAOYSA-N 0.000 claims description 26
- 239000002609 medium Substances 0.000 claims description 23
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 22
- PVNIIMVLHYAWGP-UHFFFAOYSA-N Niacin Chemical compound OC(=O)C1=CC=CN=C1 PVNIIMVLHYAWGP-UHFFFAOYSA-N 0.000 claims description 22
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 22
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 claims description 22
- 238000004519 manufacturing process Methods 0.000 claims description 20
- 239000011734 sodium Substances 0.000 claims description 18
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 17
- 230000032459 dedifferentiation Effects 0.000 claims description 17
- KWIUHFFTVRNATP-UHFFFAOYSA-N glycine betaine Chemical compound C[N+](C)(C)CC([O-])=O KWIUHFFTVRNATP-UHFFFAOYSA-N 0.000 claims description 17
- 230000006698 induction Effects 0.000 claims description 17
- 229910052708 sodium Inorganic materials 0.000 claims description 17
- 239000001763 2-hydroxyethyl(trimethyl)azanium Substances 0.000 claims description 16
- 235000019743 Choline chloride Nutrition 0.000 claims description 16
- 229960003237 betaine Drugs 0.000 claims description 16
- SGMZJAMFUVOLNK-UHFFFAOYSA-M choline chloride Chemical compound [Cl-].C[N+](C)(C)CCO SGMZJAMFUVOLNK-UHFFFAOYSA-M 0.000 claims description 16
- 229960003178 choline chloride Drugs 0.000 claims description 16
- JZRWCGZRTZMZEH-UHFFFAOYSA-N Thiamine Natural products CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 claims description 14
- 239000000835 fiber Substances 0.000 claims description 14
- 235000019157 thiamine Nutrition 0.000 claims description 14
- KYMBYSLLVAOCFI-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SCN1CC1=CN=C(C)N=C1N KYMBYSLLVAOCFI-UHFFFAOYSA-N 0.000 claims description 14
- 229960003495 thiamine Drugs 0.000 claims description 14
- 239000011721 thiamine Substances 0.000 claims description 14
- KBRKFTKQRMYINW-UHFFFAOYSA-M sodium;2-methoxy-5-nitrophenolate Chemical compound [Na+].COC1=CC=C([N+]([O-])=O)C=C1[O-] KBRKFTKQRMYINW-UHFFFAOYSA-M 0.000 claims description 12
- 238000002054 transplantation Methods 0.000 claims description 12
- XMKMDCQARLSZNY-UHFFFAOYSA-N 4,5-bis(hydroxymethyl)-2-methylpyridin-3-ol;sulfuric acid Chemical compound OS(O)(=O)=O.CC1=NC=C(CO)C(CO)=C1O XMKMDCQARLSZNY-UHFFFAOYSA-N 0.000 claims description 11
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 claims description 11
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 11
- SQUHHTBVTRBESD-UHFFFAOYSA-N Hexa-Ac-myo-Inositol Natural products CC(=O)OC1C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C1OC(C)=O SQUHHTBVTRBESD-UHFFFAOYSA-N 0.000 claims description 11
- 229930006000 Sucrose Natural products 0.000 claims description 11
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 11
- 239000000654 additive Substances 0.000 claims description 11
- 230000000996 additive effect Effects 0.000 claims description 11
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 claims description 11
- 229910052921 ammonium sulfate Inorganic materials 0.000 claims description 11
- 235000011130 ammonium sulphate Nutrition 0.000 claims description 11
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 claims description 11
- 239000004327 boric acid Substances 0.000 claims description 11
- 239000001110 calcium chloride Substances 0.000 claims description 11
- 229910001628 calcium chloride Inorganic materials 0.000 claims description 11
- GVPFVAHMJGGAJG-UHFFFAOYSA-L cobalt dichloride Chemical compound [Cl-].[Cl-].[Co+2] GVPFVAHMJGGAJG-UHFFFAOYSA-L 0.000 claims description 11
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 claims description 11
- 239000011790 ferrous sulphate Substances 0.000 claims description 11
- 235000003891 ferrous sulphate Nutrition 0.000 claims description 11
- 229960000367 inositol Drugs 0.000 claims description 11
- CDAISMWEOUEBRE-GPIVLXJGSA-N inositol Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](O)[C@@H]1O CDAISMWEOUEBRE-GPIVLXJGSA-N 0.000 claims description 11
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims description 11
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 claims description 11
- 229910000359 iron(II) sulfate Inorganic materials 0.000 claims description 11
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 11
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 11
- 229940099596 manganese sulfate Drugs 0.000 claims description 11
- 239000011702 manganese sulphate Substances 0.000 claims description 11
- 235000007079 manganese sulphate Nutrition 0.000 claims description 11
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 claims description 11
- 239000000463 material Substances 0.000 claims description 11
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 11
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 11
- 229960003512 nicotinic acid Drugs 0.000 claims description 11
- 235000001968 nicotinic acid Nutrition 0.000 claims description 11
- 239000011664 nicotinic acid Substances 0.000 claims description 11
- 239000005416 organic matter Substances 0.000 claims description 11
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 11
- 239000004323 potassium nitrate Substances 0.000 claims description 11
- 235000010333 potassium nitrate Nutrition 0.000 claims description 11
- CDAISMWEOUEBRE-UHFFFAOYSA-N scyllo-inosotol Natural products OC1C(O)C(O)C(O)C(O)C1O CDAISMWEOUEBRE-UHFFFAOYSA-N 0.000 claims description 11
- CURNJKLCYZZBNJ-UHFFFAOYSA-M sodium;4-nitrophenolate Chemical compound [Na+].[O-]C1=CC=C([N+]([O-])=O)C=C1 CURNJKLCYZZBNJ-UHFFFAOYSA-M 0.000 claims description 11
- 239000005720 sucrose Substances 0.000 claims description 11
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 claims description 11
- 229960001763 zinc sulfate Drugs 0.000 claims description 11
- 229910000368 zinc sulfate Inorganic materials 0.000 claims description 11
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims description 10
- 235000013399 edible fruits Nutrition 0.000 claims description 10
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 claims description 8
- 230000001939 inductive effect Effects 0.000 claims description 8
- 230000001902 propagating effect Effects 0.000 claims description 8
- 238000009331 sowing Methods 0.000 claims description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- 230000007613 environmental effect Effects 0.000 claims description 6
- 239000005556 hormone Substances 0.000 claims description 6
- 229940088597 hormone Drugs 0.000 claims description 6
- 239000002775 capsule Substances 0.000 claims description 5
- 239000001569 carbon dioxide Substances 0.000 claims description 5
- 229910002092 carbon dioxide Inorganic materials 0.000 claims description 5
- 230000035800 maturation Effects 0.000 claims description 5
- 229960002523 mercuric chloride Drugs 0.000 claims description 5
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims description 5
- 239000007787 solid Substances 0.000 claims description 5
- 230000001954 sterilising effect Effects 0.000 claims description 5
- 238000005406 washing Methods 0.000 claims description 5
- ZOKXTWBITQBERF-UHFFFAOYSA-N Molybdenum Chemical compound [Mo] ZOKXTWBITQBERF-UHFFFAOYSA-N 0.000 claims description 4
- 229910021529 ammonia Inorganic materials 0.000 claims description 4
- 235000009508 confectionery Nutrition 0.000 claims description 4
- 229910052750 molybdenum Inorganic materials 0.000 claims description 4
- 239000011733 molybdenum Substances 0.000 claims description 4
- 238000012549 training Methods 0.000 claims description 4
- 239000011573 trace mineral Substances 0.000 claims 2
- 235000013619 trace mineral Nutrition 0.000 claims 2
- 230000012010 growth Effects 0.000 abstract description 10
- 230000004083 survival effect Effects 0.000 abstract description 9
- 239000000126 substance Substances 0.000 abstract description 8
- 241001313857 Bletilla striata Species 0.000 abstract description 2
- 239000003112 inhibitor Substances 0.000 abstract description 2
- 241000196324 Embryophyta Species 0.000 description 21
- 210000001519 tissue Anatomy 0.000 description 17
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 14
- 235000001674 Agaricus brunnescens Nutrition 0.000 description 7
- 239000004471 Glycine Substances 0.000 description 7
- 230000008859 change Effects 0.000 description 7
- 239000011684 sodium molybdate Substances 0.000 description 7
- 235000015393 sodium molybdate Nutrition 0.000 description 7
- TVXXNOYZHKPKGW-UHFFFAOYSA-N sodium molybdate (anhydrous) Chemical compound [Na+].[Na+].[O-][Mo]([O-])(=O)=O TVXXNOYZHKPKGW-UHFFFAOYSA-N 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 230000001976 improved effect Effects 0.000 description 5
- 238000011160 research Methods 0.000 description 5
- 231100000241 scar Toxicity 0.000 description 5
- 230000004060 metabolic process Effects 0.000 description 4
- 239000003292 glue Substances 0.000 description 3
- 210000001161 mammalian embryo Anatomy 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- 101710134784 Agnoprotein Proteins 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 229930192334 Auxin Natural products 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- FAIXYKHYOGVFKA-UHFFFAOYSA-N Kinetin Natural products N=1C=NC=2N=CNC=2C=1N(C)C1=CC=CO1 FAIXYKHYOGVFKA-UHFFFAOYSA-N 0.000 description 2
- 241000233855 Orchidaceae Species 0.000 description 2
- 230000032683 aging Effects 0.000 description 2
- 239000002363 auxin Substances 0.000 description 2
- 230000008033 biological extinction Effects 0.000 description 2
- 230000000740 bleeding effect Effects 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 238000009395 breeding Methods 0.000 description 2
- 230000001488 breeding effect Effects 0.000 description 2
- -1 compound sodium nitrophenolate Chemical class 0.000 description 2
- 238000012136 culture method Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- SEOVTRFCIGRIMH-UHFFFAOYSA-N indole-3-acetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CNC2=C1 SEOVTRFCIGRIMH-UHFFFAOYSA-N 0.000 description 2
- QANMHLXAZMSUEX-UHFFFAOYSA-N kinetin Chemical compound N=1C=NC=2N=CNC=2C=1NCC1=CC=CO1 QANMHLXAZMSUEX-UHFFFAOYSA-N 0.000 description 2
- 229960001669 kinetin Drugs 0.000 description 2
- 230000029553 photosynthesis Effects 0.000 description 2
- 238000010672 photosynthesis Methods 0.000 description 2
- 230000008635 plant growth Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- YGSDEFSMJLZEOE-UHFFFAOYSA-N salicylic acid Chemical compound OC(=O)C1=CC=CC=C1O YGSDEFSMJLZEOE-UHFFFAOYSA-N 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- LUEWUZLMQUOBSB-FSKGGBMCSA-N (2s,3s,4s,5s,6r)-2-[(2r,3s,4r,5r,6s)-6-[(2r,3s,4r,5s,6s)-4,5-dihydroxy-2-(hydroxymethyl)-6-[(2r,4r,5s,6r)-4,5,6-trihydroxy-2-(hydroxymethyl)oxan-3-yl]oxyoxan-3-yl]oxy-4,5-dihydroxy-2-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol Chemical compound O[C@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@@H](O[C@@H]2[C@H](O[C@@H](OC3[C@H](O[C@@H](O)[C@@H](O)[C@H]3O)CO)[C@@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O LUEWUZLMQUOBSB-FSKGGBMCSA-N 0.000 description 1
- KIZQNNOULOCVDM-UHFFFAOYSA-M 2-hydroxyethyl(trimethyl)azanium;hydroxide Chemical compound [OH-].C[N+](C)(C)CCO KIZQNNOULOCVDM-UHFFFAOYSA-M 0.000 description 1
- BTJIUGUIPKRLHP-UHFFFAOYSA-N 4-nitrophenol Chemical compound OC1=CC=C([N+]([O-])=O)C=C1 BTJIUGUIPKRLHP-UHFFFAOYSA-N 0.000 description 1
- 208000002874 Acne Vulgaris Diseases 0.000 description 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 1
- 229920000856 Amylose Polymers 0.000 description 1
- 241000416162 Astragalus gummifer Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- 244000146553 Ceiba pentandra Species 0.000 description 1
- 244000247747 Coptis groenlandica Species 0.000 description 1
- 235000002991 Coptis groenlandica Nutrition 0.000 description 1
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 1
- 229920002581 Glucomannan Polymers 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 206010023126 Jaundice Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000867725 Mucilago Species 0.000 description 1
- 244000063675 Orchis mascula Species 0.000 description 1
- 241001250598 Pleione bulbocodioides Species 0.000 description 1
- 206010040849 Skin fissures Diseases 0.000 description 1
- 241000191940 Staphylococcus Species 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 241000194017 Streptococcus Species 0.000 description 1
- 206010053615 Thermal burn Diseases 0.000 description 1
- 241000130764 Tinea Species 0.000 description 1
- 208000002474 Tinea Diseases 0.000 description 1
- 229920001615 Tragacanth Polymers 0.000 description 1
- 208000025865 Ulcer Diseases 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 206010000496 acne Diseases 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 230000001355 anti-mycobacterial effect Effects 0.000 description 1
- 230000000118 anti-neoplastic effect Effects 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000001109 blastomere Anatomy 0.000 description 1
- 230000003833 cell viability Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 230000002950 deficient Effects 0.000 description 1
- 230000002183 duodenal effect Effects 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000014509 gene expression Effects 0.000 description 1
- 230000035784 germination Effects 0.000 description 1
- 229940046240 glucomannan Drugs 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 238000005469 granulation Methods 0.000 description 1
- 230000003179 granulation Effects 0.000 description 1
- 239000003630 growth substance Substances 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 230000003859 lipid peroxidation Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 230000004899 motility Effects 0.000 description 1
- 210000004877 mucosa Anatomy 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 238000010899 nucleation Methods 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- FJKROLUGYXJWQN-UHFFFAOYSA-N papa-hydroxy-benzoic acid Natural products OC(=O)C1=CC=C(O)C=C1 FJKROLUGYXJWQN-UHFFFAOYSA-N 0.000 description 1
- 230000032696 parturition Effects 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 231100000572 poisoning Toxicity 0.000 description 1
- 230000000607 poisoning effect Effects 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- 238000002407 reforming Methods 0.000 description 1
- 230000029058 respiratory gaseous exchange Effects 0.000 description 1
- 230000010496 root system development Effects 0.000 description 1
- 229960004889 salicylic acid Drugs 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000011218 seed culture Methods 0.000 description 1
- 230000021217 seedling development Effects 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- 230000000392 somatic effect Effects 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 230000014793 stomatal movement Effects 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
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- 239000000375 suspending agent Substances 0.000 description 1
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- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 239000000196 tragacanth Substances 0.000 description 1
- 229940116362 tragacanth Drugs 0.000 description 1
- 235000010487 tragacanth Nutrition 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 201000008827 tuberculosis Diseases 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
- 235000019156 vitamin B Nutrition 0.000 description 1
- 239000011720 vitamin B Substances 0.000 description 1
- 229940046001 vitamin b complex Drugs 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
- 230000037303 wrinkles Effects 0.000 description 1
Classifications
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a bletilla pseudobulb cutting tissue culture seedling method and a culture medium for tissue culture seedling, which effectively break through the bottleneck of producing a large number of small seedlings. The culture medium effectively improves the quality of the emergence of seedlings of protocorms by adding substances such as organic substances and inhibitors of harmful substances required by the bletilla striata tissue culture seedlings in each development period, also improves the growth efficiency of differentiated seedlings, prevents the death phenomenon after the cut seedlings are transferred, greatly improves the quality of grown seedlings, and further improves the survival rate of the tissue culture seedlings after the tissue culture seedlings are out of bottles and transplanted.
Description
Technical field
The present invention relates to bletilla Cultivating techniques field, and in particular to bletilla pseudobulb cut tissue culture culture medium and its
Tissue culture method.
Background technology
Bletilla Bletilla striata (Thunb. ex A.Murray) Reichb.f. is that orchid family bletilla belongs to perennial
Terrestrial orchid class plant, also known as：Company and grass, Bai Gen, pleionebulbocodioides rolfe, YANGJIAOQI.Underground stem tuber is used as medicine.《Chinese Pharmacopoeia》Record：Have
Astringing to arrest bleeding, detumescence and promoting granulation.For spitting blood, spit blood, traumatism and bleeding, sore swollen toxin, chapped skin.Pharmacological research, bletilla have
Resist stomach and duodenal mucosa ulcer and perforation, anti-mycobacterium tuberculosis, staphylococcus, streptococcus and antineoplastic action；It is modern
Research is had shown that, the compositions such as abundant mucilaginous substance (bletilla glue), starch, volatile oil, glucose are contained in the stem tuber of bletilla.Bletilla glues
Liquid matter belongs to plant roots and stems class natural macromolecule amylose, can replace tragacanth powder with gummi arabicum pulveratum as emulsifying agent, suspension
Agent is used for food and chemical field.It can be additionally used in silk cotton yarn and the slurry of smart table binding and layout；Bletilla purifies out white by science
And glucomannan, can promote and directly participation damaged tissues and cell reparation and metabolic process, applied in cosmetic industry
Acted on always with preferable removing wrinkle and resisting aging.Bletilla glue external application is embrocated, and can eliminate the vestige under acne scar on the face, allows the smooth nothing of skin
Trace.Plastics is made with bletilla, is applied to the surface of a wound, for treating scald；With mucilago salep compatibility coptis medicinal extract, salicylic acid etc.
Ointment is made, tinea of feet and hands can be treated.In addition, the medicine-food two-purpose that bletilla or the Ministry of Public Health specify, the former material available for health food
Material, stem tuber can be used for making wine.Bletilla is also known as purple blue, purple a species of orchid, and not only plant shape is graceful, and pattern is gorgeous, so being afforestation again
Rare rare Flower Resources.
Bletilla is widely used, and comprehensive utilization and industrial value increasingly highlight, and for a long time, is arranged due to lacking effectively protection
Apply, excessive xcessive digging, cause wild bletilla resource to be faced with the danger of extinction.In order to protect the wild resource of bletilla,《In
State's plant Red Data Book --- rare extinction plants》（1st）With《Endangered species of wild fauna and flora international trade pact》(CITES),
Bletilla is included in the list of the endangered animal and plant of protection.Wild resource is deficient, and supply falls short of demand and price continuous rise in market,
Produced through significantly affecting to China's pharmaceuticals industry, food service industry, daily-use chemical industry etc..But because the seed of bletilla is extremely small
And without endosperm, therefore it is difficult to which growth seedling, the cultivation of seedling turn into bottleneck under natural situation；Traditional cultivation is mainly by underground
Stem tuber is bred, and not only breeding coefficient is low, and consumption kind amount is big, and production cost is high, it is difficult to meet the needs of large area production.
Therefore, carry out the protection of bletilla wild resource and the kind research of wild change man, accelerate the seedling of bletilla by tissue cultures
Breed, have become and solve bletilla in imminent danger, resource scarcity, market the important means and approach that supply falls short of demand.
At present, the work with seedling is sprouted on induction bletilla seed, the country there are many scholars to do research, but seed is sprouted
Used program is but had nothing in common with each other after hair, and traditional method has：1）" point of protocorm → induction Protocorm Multiplication → induced bud
Change → root induction ", breed mainly using inducing Protocorm Multiplication to complete, squamous subculture also uses protocorm；2) " protocorm →
Induce the differentiation of adventitious buds proliferation → induced bud and take root ", squamous subculture uses Multiple Buds；3) " point of protocorm → induced bud
Change → root induction ", 4) using protocorm directly " Growth and Differentiation → root induction " etc., although these methods can be taken root
Seedling, but it is widely different in effect and input ratio, and laboratory stage is remained in mostly, really it is used for the very few of production, and
And there is many unfavorable factors：1）Need the process that carries out more；2）The cycle time of seedling is longer；3）The quality of seedling is very
Difference, it is unfavorable for transplant survival, survival rate is low, is not easy to wide popularization and application.
The content of the invention
The present invention breaks through the bottleneck for producing a large amount of seedlings, there is provided a kind of bletilla by reforming the method cultivated and culture medium
The method that pseudobulb cuts tissue culture, using " protocorm differentiation → strong sprout → pseudobulb cutting → differentiation culture → production
Seedling " or the culture program of " pseudobulb cutting → differentiation is cultivated → produces seedling ".Simultaneously, there is provided suitable for the training of above-mentioned method for culturing seedlings
Support base, shorten the cycle so as to effectively realize, improve seedling amount and seedling quality, and make to transplant under bletilla seedling domestication into
Motility rate significantly improves.
In order to solve the above technical problems, the present invention is adopted the following technical scheme that to realize：
A kind of bletilla protocorm induction medium, its prescription include consisting of point（Without special instruction, unit mg/L）：
1st, a great number of elements：Potassium nitrate 2700-2900, ammonium sulfate 461-465, potassium dihydrogen phosphate 390-415, magnesium sulfate 180-190,
Calcium chloride 164-166；
2nd, it is micro-：Manganese sulfate 22.0-22.5, zinc sulfate 8.2-8.8, boric acid 6.0-6.4, KI 0.80-0.83, molybdenum
Sour sodium 0.22-0.26, copper sulphate 0.021-0.025, cobalt chloride 0.023-0.025；
3rd, molysite：Ferrous sulfate 27.2-27.8, EDTA 37.0-37.43；
4th, organic matter：Inositol 100-105, sulfuric acid thiamine element 50-55, sulfuric acid pyridoxol 0.8-1.2, nicotinic acid 0.3-0.5, sweet ammonia
Sour 1.8-2.1, glycine betaine 500-520, Choline Chloride 280-315,5- nitroguaiacol sodium salt 0.9-1.2, Sodium p-nitrophenoxide
1.8- 2.2, sodium onitrophenol 1.8- 2.2；
5th, additive： AgNO3 2.85-3.10, sucrose 28-33g/L.
A kind of bletilla breaks up strong seedling culture base, and its prescription includes consisting of point（Without special instruction, unit mg/L）：
1st, a great number of elements：Potassium nitrate 2700-2900, ammonium sulfate 461-465, potassium dihydrogen phosphate 390-415, magnesium sulfate 180-190,
Calcium chloride 164-166；
2nd, it is micro-：Manganese sulfate 22.0-22.5, zinc sulfate 8.2-8.8, boric acid 6.0-6.4, KI 0.80-0.83, molybdenum
Sour sodium 0.22-0.26, copper sulphate 0.021-0.025, cobalt chloride 0.023-0.025；
3rd, molysite：Ferrous sulfate 27.2-27.8, EDTA 37.0-37.43；
4th, organic matter：Inositol 100-105, sulfuric acid thiamine element 50-55, sulfuric acid pyridoxol 0.8-1.2, nicotinic acid 0.3-0.5, sweet ammonia
Sour 1.8-2.1, glycine betaine 500-520, Choline Chloride 280-315,5- nitroguaiacol sodium salt 0.9-1.2, Sodium p-nitrophenoxide
1.8- 2.2, sodium onitrophenol 1.8- 2.2；
5th, hormone NAA 0.15-0.22
6th, additive：AgNO3 2.85-3.10, sucrose 18-22g/L.
The pH value of above-mentioned culture medium controls： 5.5—5.8.
A kind of method of bletilla pseudobulb cutting tissue culture, comprises the following steps：
1）Bletilla protocorm Fiber differentiation seedling：Take the bletilla capsule of maturation to be scrubbed in bromogeramine solution and remove dirt, be placed in
On superclean bench, 15-20 min are sterilized with 0.2% mercuric chloride, then with aseptic water washing 3-5 time, the filter paper suction by it with sterilizing
After solid carbon dioxide point, cut at the top of fruit, then longitudinally splitted from middle part of fruit, seed is uniformly broadcasted sowing to the bletilla protocorm prepared
On stem inducing culture；Bletilla protocorm induction medium after planting be placed in 28 DEG C, intensity of illumination 1500-2000lux, daily
Culture 50-65 days is carried out under the light application time environmental condition of 12 hours, obtains the healthy and strong seedling that base portion expands pseudobulb spherical in shape；
Pay attention to sowing gap during sowing, using light seeding mode, not broadcast too much, reserve the space for allowing seedling to grow, broadcast too
Close, seedling growth is excessively slim and frahile, is unfavorable for later stage culture；When bletilla seed is cultivated 1 day under above-mentioned suitable condition of culture, embryo
Globulate is expanded in water suction, and blastocyte starts to divide within second day, and globular embryo progressively greening, seed just starts to sprout after 5 days, and seed is sprouted
Small protocorm is formed during hair, chloroplaset occurs into the cell in protocorm；When continuing to develop 15-20 days, the side of apical meristem
Differentiate a piece of cotyledon, and differentiate some very thin rhizoids protocorm the lower portion of the stem surrounding is radial, in son after 20 days
The relative one side of leaf initially forms rough leaf, is cultivated 50-65 days on inducing culture, can be to grow 3-5 pieces of leaflet
Piece, highly reach 5-7cm, base portion expands the healthy and strong seedling of pseudobulb spherical in shape.
2）Strong seedling culture：By step 1）The bletilla seedling of Fiber differentiation is timely transferred on differentiation strong seedling culture base,
Under conditions of 28 DEG C of room temperature, intensity of illumination 2000-2500lux, daily illumination 12 hours, to cultivate 50-55 days, pseudobulb is ripe,
The bletilla plantlet of pseudobulb must be carried；Production transplantation of seedlings domestication can be used as or carry out pseudobulb increasing as propagation propagating materials
Grow, dedifferentiation culture；
Strong seedling culture after 3 days visible bletilla seedling start to grow, 7 days rear blades and stem mushroom out, and after 15 days, pseudobulb is obvious
Increase, 30-40 days pseudobulbs stop increase, and plant body progressively tends to ripe, and small part blade starts to turn to be yellow withered after 50-55 days
Wither, tissue-cultured seedling now can be as transplanting domestication or propagating materials progress pseudobulb increasing as Multiplying culture under production seedling
Grow, dedifferentiation culture.
3）Pseudobulb Multiplying culture：Ripe bletilla plant is peelled off into cauline leaf on sterile platform, cuts coring, vacation
Bulb is transferred on fresh differentiation strong seedling culture base, small in 28 DEG C of room temperature, intensity of illumination 2000-2500lux, daily illumination 12
When under conditions of cultivate 20-25 days；
During Multiplying culture, pseudobulb dissolves several sturdy adventitious buds in leaf scar punishment, and mushrooms out, 20-25 days, point
The seedling of change can grow, blade 4-5 piece high to 3-5cm.
4）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band
One fritter pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C of room temperature, intensity of illumination 2000-
After 2500lux, daily illumination are cultivated 45-50 days under conditions of 12 hours, pseudobulb is ripe, obtains the small plant of bletilla with pseudobulb
Strain, production transplantation of seedlings domestication can be used as or carry out pseudobulb propagation, dedifferentiation culture as propagation propagating materials.
The application breaks up tissue culture using the cutting of bletilla pseudobulb, and drastically increase tissue culture breeds efficiency,
By splitting to the pseudobulb with several adventitious buds, breed coefficient and effectively improve several times, meanwhile, the later stage can directly adopt
Propagation dedifferentiation culture is carried out with ripe pseudobulb, growing-seedling period is greatly shortened, and is effectively broken through and is produced a large amount of bletilla seedlings
Bottleneck, artificial breeding cost is greatly reduced, realize bletilla it is extensive extensively plantation.
The culture medium of the application, by adding the organic substance required for each developmental stage of bletilla tissue-cultured seedling and being unfavorable for giving birth to
Long hair educates the inhibitor of harmful substance, has effectively promoted ammonium ion more rapidly to be absorbed by plant, and alleviates training
It is too high to toxic action caused by bletilla tissue-cultured seedling to support nitrogen concentration in base.Applicant uses conventional medium before, and bletilla divides again
When changing culture, the seedling after cutting is transferred in new culture medium, and blade turns yellow quickly, and applicant passes through by researching and analysing
A certain amount of AgNO is added in the medium3, so as to effectively change the problem of blade flavescence aging, and effectively facilitate organ
Occur with somatic embryo.Appropriate AgNO is separately added into protocorm induction medium, differentiation strong seedling culture base3Can be effective
Caused ethene during suppression Protocorm Multiplication and strong seedling culture, the quality of protocorm emergence is improved and improved, is also improved
Differentiation seedling growth efficiency, preventing from cutting dead phenomenon after seedling transfer, seedling quality is also greatly improved, while still further
Ground improves the survival rate of tissue-cultured seedling bottle outlet transplanting.
Choline Chloride in the application（Scientific name：2- ethoxys-trimethyl bursine）, one kind of vitamin B complex, it is one
The growth regulator of kind plant photosynthesis promoter, more wide spectrum.Choline Chloride can absorb via the stem, leaf, root of plant,
Then the position worked is transmitted to faster, and its physiological action can promote the photosynthesis of plant, promote root system development, can make
Photosynthate is as much as possible to be run up in stem tuber, root tuber and bulb.In addition, it is mixed with kinetin, class auxin, can add
Its fast movement, more effectively plays the effect of kinetin, class auxin.The effect stability under Different climate, ecological environmental condition,
There is the obvious effect of expanding to root tuber, stem tuber, bulb, pseudobulb.
The Choline Chloride of the proportioning is added when inducing bletilla protocorm, drastically increases the vacation of seedling base portion spheroidal
Bulb expands the speed of growth；During strong seedling culture, appropriate Choline Chloride is added, after cultivating 15 days, pseudobulb significantly increases,
30-40 days pseudobulbs progressively tend to ripe, and Choline Chloride effectively makes pseudobulb in bletilla tissue cultures increase rapidly, increase weight.
Thiamine is one of material extremely sensitive during bletilla tissue-cultured seedling development growth, is found in research, wild shape
Bletilla root under state plays the role of self synthesis thiamine, and the corresponding content for improving thiamine in culture medium is advantageous to bletilla tissue culture
The growth of seedling.
Glycine betaine in the application culture medium is a kind of alkaloid, its molecular structure, application effect and natural glycine betaine without
Significant difference, belong to the natural goods equivalent of chemical synthesis（Scientific name：Betaine）.Glycine betaine effectively can maintain cell to ooze
Pressure thoroughly, avoid the murder by poisoning of cytoplasm high concentration inorganic ion-pair enzyme and metabolism；Lipid peroxidation metabolism degree is effectively reduced, is significantly reduced
Na water ratio in root system, by ca2+Root system is trapped in, the ability of root system resistance salt damage can be effectively improved, meanwhile, glycine betaine is to inverse
There is certain regulating and controlling effect under the conditions of border to stomatal movement, respiration and related gene expression.
Pass through a certain amount of sodium onitrophenol in the application（Molecular formula：C6H4NO3Na, relative molecular weight：161）, it is right
P-nitrophenol sodium（Molecular formula：C6H4NO3Na, relative molecular weight：161）With 5- nitroguaiacol sodium salts（Molecular formula：
C7H6NO4Na, relative molecular weight：191）Organic complex compound sodium nitrophenolate is formed, promotes plant growth regulating, it is a kind of strength
Cell-activating agent, by increasing capacitance it is possible to increase the permeability of plasmalemma, with can be penetrated into after plant contact in plant rapidly, promote cell
Plasm flows, and improves cell viability, accelerates the new of plant and deposits metabolism, strengthens the activity of ATP enzyme, produce a large amount of ATP energy
Quantum, supply Plant To Nutrient element receive energy, the ATP energy of consumption required for supplement plant can actively receive, promote to plant
Thing blade broadens, becomes big, thickening, increase blade face to the contact surface of nutrient, improve blade to nutrient can yield.It is logical
The compound sodium nitrophenolate for adding the weight in the medium is crossed, after the bletilla seedling transfer in each period, can mushroom out, shorten
The time of slow seedling, promote the quick robust growth of plant.
Experiment confirms：Sprouting and the quick shape of seedling growth and pseudobulb of the culture medium to bletilla seed by transformation
Into formed with good facilitation, the germination percentage of seed and the speed of growth of embryoid and seedling pseudobulb and quality all
Apparently higher than other conventional mediums.
Brief description of the drawings
The present invention is described further below in conjunction with the accompanying drawings：
Fig. 1 is the culture process schematic flow sheet of the present invention.
Embodiment
Referring to Fig. 1, technical solution of the present invention is further elaborated on below in conjunction with accompanying drawing.
Embodiment one
The bletilla protocorm induction medium of the present embodiment, its prescription include consisting of point（Without special instruction, unit mg/
L）：
1st, a great number of elements：Potassium nitrate 2800, ammonium sulfate 463, potassium dihydrogen phosphate 400, magnesium sulfate 185, calcium chloride 166；
2nd, it is micro-：Manganese sulfate 22.3, zinc sulfate 8.6, boric acid 6.2, KI 0.83, sodium molybdate 0.25, copper sulphate
0.025th, cobalt chloride 0.025；
3rd, molysite：Ferrous sulfate 27.8, EDTA 37.3；
4th, organic matter：Inositol 100, sulfuric acid thiamine element 50, sulfuric acid pyridoxol 1.0, nicotinic acid 0.5, glycine 2.0, glycine betaine 500,
Choline Chloride 300,5- nitroguaiacol sodium salts 1.0, Sodium p-nitrophenoxide 2.0, sodium onitrophenol 2.0；
5th, additive： AgNO33.0th, the g/L of sucrose 30.
The bletilla differentiation strong seedling culture base of the present embodiment, its prescription include consisting of point（Without special instruction, unit is
mg/L）：
1st, a great number of elements：Potassium nitrate 2800, ammonium sulfate 463, potassium dihydrogen phosphate 400, magnesium sulfate 185, calcium chloride 166；
2nd, it is micro-：Manganese sulfate 22.3, zinc sulfate 8.6, boric acid 6.2, KI 0.83, sodium molybdate 0.25, copper sulphate
0.025th, cobalt chloride 0.025；
3rd, molysite：Ferrous sulfate 27.8, EDTA 37.3；
4th, organic matter：Inositol 100, sulfuric acid thiamine element 50, sulfuric acid pyridoxol 1.0, nicotinic acid 0.5, glycine 2.0, glycine betaine 500,
Choline Chloride 300,5- nitroguaiacol sodium salts 1.0, Sodium p-nitrophenoxide 2.0, sodium onitrophenol 2.0；
5th, hormone NAA 0.2；
6th, additive：AgNO33.0, sucrose 20g/L.
The pH value of above two culture medium is 5.5.
The method of the bletilla pseudobulb cutting tissue culture of the present embodiment, comprises the following steps：
1）Bletilla protocorm Fiber differentiation seedling：Take the bletilla capsule of maturation to be scrubbed in bromogeramine solution and remove dirt, be placed in
On superclean bench, 15-20 min are sterilized with 0.2% mercuric chloride, then with aseptic water washing 3-5 time, the filter paper suction by it with sterilizing
After solid carbon dioxide point, cut at the top of fruit, then longitudinally splitted from middle part of fruit, seed is uniformly broadcasted sowing to the bletilla protocorm prepared
On stem inducing culture；Bletilla protocorm induction medium after planting be placed in 28 DEG C, intensity of illumination 1800lux, daily illumination
Culture 55 days is carried out under the environmental condition of 12 hours time, obtains the healthy and strong seedling that base portion expands pseudobulb spherical in shape；
Bletilla seed culture after 5 days seed start to sprout, form small protocorm, chloroplaset occurs into the cell in protocorm；Continue to send out
When educating 15 days, top differentiates cotyledon, and protocorm the lower portion of the stem surrounding differentiates some rhizoids, in the one side shape that cotyledon is relative after 20 days
Into rough leaf, during culture 55 days, grow 3-5 pieces of vanelets, highly reach 5-7cm, base portion expands pseudobulb spherical in shape
Healthy and strong seedling.
2）Strong seedling culture：By step 1）The bletilla seedling of Fiber differentiation is timely transferred on differentiation strong seedling culture base,
Under conditions of 28 DEG C of room temperature, intensity of illumination 2200lux, daily illumination 12 hours, cultivate 50 days, pseudobulb is ripe, obtains with vacation
The bletilla plantlet of bulb；
Strong seedling culture after 3 days bletilla seedling start to grow, 7 days rear blades and stem mushroom out, and after 15 days, pseudobulb significantly increases
36 days pseudobulbs stop increase, and plant body progressively tends to ripe, and it is withered to start jaundice for small part blade after 50 days.
3）Pseudobulb Multiplying culture：Ripe bletilla plant obtained by step 2 is peelled off into cauline leaf on sterile platform, cuts coring,
Pseudobulb is transferred on fresh differentiation strong seedling culture base, it is small in 28 DEG C of room temperature, intensity of illumination 2200lux, daily illumination 12
When under the conditions of cultivate 20 days, pseudobulb dissolves the sturdy adventitious buds that 4-8 do not wait not in leaf scar punishment, and mushrooms out, at 20 days,
The adventitious bud length of differentiation is to seedling high 3-5cm, blade 4-5 pieces.
4）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band
Part pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C, intensity of illumination 2200lux of room temperature, every daylight
After being cultivated 50 days under the conditions of 12 hours, pseudobulb is ripe, obtains the bletilla plantlet with pseudobulb, now, as production seedling
Transplanting domestication.
The present embodiment, to culture to 105 days bletilla plantlet used times with pseudobulb, Multiplying culture and divides again from seed
Change 70 days used times of culture, tame used time 175 days altogether eventually as production transplantation of seedlings, the seedling of Fiber differentiation is compared with conventional medium
The seedling of cultivation averagely weighs 1.4 times, and production seedling averagely weighs 1.5 times compared with the production seedling that conventional medium is cultivated, and transplanting domestication survives
Rate is up to 98.2%.
Embodiment two
The bletilla protocorm induction medium of the present embodiment, its prescription include consisting of point（Without special instruction, unit mg/
L）：
1st, a great number of elements：Potassium nitrate 2700, ammonium sulfate 461, potassium dihydrogen phosphate 390, magnesium sulfate 180, calcium chloride 164；
2nd, it is micro-：Manganese sulfate 22.0, zinc sulfate 8.2, boric acid 6.0, KI 0.80, sodium molybdate 0.22, copper sulphate
0.021st, cobalt chloride 0.023；
3rd, molysite：Ferrous sulfate 27.2, EDTA 37.0；
4th, organic matter：Inositol 100, sulfuric acid thiamine element 50, sulfuric acid pyridoxol 0.80, nicotinic acid 0.3, glycine 1.8, glycine betaine
500th, Choline Chloride 280,5- nitroguaiacol sodium salts 0.9, Sodium p-nitrophenoxide 1.8, sodium onitrophenol 1.8；
5th, additive： AgNO32.85th, the g/L of sucrose 28.
The bletilla differentiation strong seedling culture base of the present embodiment, its prescription include consisting of point（Without special instruction, unit is
mg/L）：
1st, a great number of elements：Potassium nitrate 2700, ammonium sulfate 461, potassium dihydrogen phosphate 390, magnesium sulfate 180, calcium chloride 164；
2nd, it is micro-：Manganese sulfate 22.0, zinc sulfate 8.2, boric acid 6.0, KI 0.80, sodium molybdate 0.22, copper sulphate
0.021st, cobalt chloride 0.023；
3rd, molysite：Ferrous sulfate 27.2, EDTA 37.0；
4th, organic matter：Inositol 100, sulfuric acid thiamine element 50, sulfuric acid pyridoxol 0.80, nicotinic acid 0.3, glycine 1.8, glycine betaine
500th, Choline Chloride 280,5- nitroguaiacol sodium salts 0.9, Sodium p-nitrophenoxide 1.8, sodium onitrophenol 1.8；
5th, hormone NAA 0.15；
6th, additive：AgNO33.0th, sucrose 20g/L.
The pH value of above two culture medium is 5.8.
The method of the bletilla pseudobulb cutting tissue culture of the present embodiment, comprises the following steps：
1）Bletilla protocorm Fiber differentiation seedling：Take the bletilla capsule of maturation to be scrubbed in bromogeramine solution and remove dirt, be placed in
On superclean bench, 15-20 min are sterilized with 0.2% mercuric chloride, then with aseptic water washing 3-5 time, the filter paper suction by it with sterilizing
After solid carbon dioxide point, cut at the top of fruit, then longitudinally splitted from middle part of fruit, seed is uniformly broadcasted sowing to the bletilla protocorm prepared
On stem inducing culture；Bletilla protocorm induction medium after planting be placed in 28 DEG C, intensity of illumination 1500lux, daily illumination
Culture 65 days is carried out under the environmental condition of 12 hours time, obtains the healthy and strong seedling that base portion expands pseudobulb spherical in shape, blade 3-5
Piece, highly reach 5-7cm.
2）Strong seedling culture：By step 1）The bletilla seedling of Fiber differentiation is timely transferred on differentiation strong seedling culture base,
Under conditions of 28 DEG C of room temperature, intensity of illumination 2000lux, daily illumination 12 hours, cultivate 45 days, pseudobulb is ripe, obtains with vacation
The bletilla plantlet of bulb, now, as production transplantation of seedlings domestication.
The present embodiment is planted as the production transplantation of seedlings domestication strain used time 110 from seed to culture to the bletilla with pseudobulb is small
My god, the seedling of Fiber differentiation averagely weighs 1.1 times compared with the seedling that conventional medium is cultivated, transplanting domestication survival rate 97.9%.
Embodiment three
The bletilla differentiation strong seedling culture base of the present embodiment, its prescription include consisting of point（Without special instruction, unit mg/L）：
1st, a great number of elements：Potassium nitrate 2900, ammonium sulfate 465, potassium dihydrogen phosphate 415, magnesium sulfate 190, calcium chloride 166；
2nd, it is micro-：Manganese sulfate 22.5, zinc sulfate 8.8, boric acid 6.4, KI 0.83, sodium molybdate 0.26, copper sulphate
0.025th, cobalt chloride 0.025；
3rd, molysite：Ferrous sulfate 27.8, EDTA 37.43；
4th, organic matter：Inositol 105, sulfuric acid thiamine element 55, sulfuric acid pyridoxol 1.2, nicotinic acid 0.5, glycine 2.1, glycine betaine 520,
Choline Chloride 315,5- nitroguaiacol sodium salts 1.2, Sodium p-nitrophenoxide 2.2, sodium onitrophenol 2.2；
5th, hormone NAA 0.22；
6th, additive：AgNO33.1st, sucrose 22g/L.
The pH value of above two culture medium is 5.7.
The method of the bletilla pseudobulb cutting tissue culture of the present embodiment, comprises the following steps：
1）Pseudobulb Multiplying culture：Ripe bletilla plant is peelled off into cauline leaf on sterile platform, coring is cut, pseudobulb
It is transferred on fresh differentiation strong seedling culture base, in 28 DEG C of room temperature, intensity of illumination 2000lux, under the conditions of daily illumination 12 hours
Culture 25 days, pseudobulb dissolves the sturdy adventitious buds that 4-8 do not wait not in leaf scar punishment, and mushrooms out, and at 25 days, differentiation is not
Normal bud is grown to seedling high 3-5cm, blade 4-5 pieces；
2）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band one is small
Block pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C of room temperature, intensity of illumination 2000lux, daily illumination
After being cultivated 50 days under the conditions of 12 hours time, pseudobulb is ripe, obtains the bletilla plantlet with pseudobulb, as production transplantation of seedlings
Domestication.
The present embodiment, by Multiplying culture and dedifferentiation culture, obtains conduct using the pseudobulb of ripe bletilla plant
Transplantation of seedlings domestication bletilla seedling, 75 days used times are produced, production seedling averagely weighs 1.3 times compared with the production seedling that conventional medium is cultivated, transplanting
Tame survival rate 98.3%.
Example IV
The bletilla protocorm induction medium of the present embodiment, its prescription include consisting of point（Without special instruction, unit mg/
L）：
1st, a great number of elements：Potassium nitrate 2850, ammonium sulfate 462, potassium dihydrogen phosphate 405, magnesium sulfate 188, calcium chloride 165；
2nd, it is micro-：Manganese sulfate 22.2, zinc sulfate 8.5, boric acid 6.3, KI 0.82, sodium molybdate 0.24, copper sulphate
0.024th, cobalt chloride 0.024；
3rd, molysite：Ferrous sulfate 27.4, EDTA 37.26；
4th, organic matter：Inositol 102, sulfuric acid thiamine element 53, sulfuric acid pyridoxol 1.1, nicotinic acid 0.4, glycine 2.0, glycine betaine 510,
Choline Chloride 305,5- nitroguaiacol sodium salts 1.1, Sodium p-nitrophenoxide 2.2, sodium onitrophenol 2.2；
5th, additive： AgNO32.95th, the g/L of sucrose 31.
The bletilla differentiation strong seedling culture base of the present embodiment, its prescription include consisting of point（Without special instruction, unit is
mg/L）：
1st, a great number of elements：Potassium nitrate 2850, ammonium sulfate 462, potassium dihydrogen phosphate 405, magnesium sulfate 188, calcium chloride 165；
2nd, it is micro-：Manganese sulfate 22.2, zinc sulfate 8.5, boric acid 6.3, KI 0.82, sodium molybdate 0.24, copper sulphate
0.024th, cobalt chloride 0.024；
3rd, molysite：Ferrous sulfate 27.4, EDTA 37.26；
4th, organic matter：Inositol 102, sulfuric acid thiamine element 53, sulfuric acid pyridoxol 1.1, nicotinic acid 0.4, glycine 2.0, glycine betaine 510,
Choline Chloride 305,5- nitroguaiacol sodium salts 1.1, Sodium p-nitrophenoxide 2.2, sodium onitrophenol 2.2；
5th, hormone NAA 0.17；
6th, additive：AgNO32.85th, sucrose 18g/L.
The pH value of above two culture medium is 5.6.
The method of the bletilla pseudobulb cutting tissue culture of the present embodiment, comprises the following steps：
1）Bletilla protocorm Fiber differentiation seedling：Take the bletilla capsule of maturation to be scrubbed in bromogeramine solution and remove dirt, be placed in
On superclean bench, 15-20 min are sterilized with 0.2% mercuric chloride, then with aseptic water washing 3-5 time, the filter paper suction by it with sterilizing
After solid carbon dioxide point, cut at the top of fruit, then longitudinally splitted from middle part of fruit, seed is uniformly broadcasted sowing to the bletilla protocorm prepared
On stem inducing culture；Bletilla protocorm induction medium after planting be placed in 28 DEG C, intensity of illumination 2000lux, daily illumination
Culture 58 days is carried out under the environmental condition of 12 hours time, obtains the healthy and strong seedling that base portion expands pseudobulb spherical in shape；Bletilla seed
Seed starts to sprout after cultivating 5 days, forms small protocorm, chloroplaset occurs into the cell in protocorm；When continuing to develop 15 days, top
Cotyledon is differentiated, protocorm the lower portion of the stem surrounding differentiates some rhizoids, and rough leaf is formed in the relative one side of cotyledon after 20 days,
During culture 58 days, grow up to 3-5 pieces of vanelets, highly reach 5-7cm, base portion expands the healthy and strong seedling of pseudobulb spherical in shape.
2）Strong seedling culture：By step 1）The bletilla seedling of Fiber differentiation is timely transferred on differentiation strong seedling culture base,
Under conditions of 28 DEG C of room temperature, intensity of illumination 2500lux, daily illumination 12 hours, cultivate 55 days, pseudobulb is ripe, obtains with vacation
The bletilla plantlet of bulb, strong seedling culture after 3 days bletilla seedling start to grow, 7 days rear blades and stem mushroom out, after 15 days,
Pseudobulb significantly increases 36 days pseudobulbs and stops increase, and plant body progressively tends to ripe, and small part blade starts to turn to be yellow after 50 days
Withered, half bletilla plantlet is used to produce transplantation of seedlings domestication, and second half carries out pseudobulb as the propagating materials of Multiplying culture
Propagation, dedifferentiation culture.
3）Pseudobulb Multiplying culture：The ripe bletilla plant of the propagating materials of Multiplying culture will be used as obtained by step 2 in nothing
Cauline leaf is peelled off on bacterium platform, cuts coring, pseudobulb is transferred on fresh differentiation strong seedling culture base, in 28 DEG C of room temperature, illumination
Intensity 2500lux, daily light application time are cultivated 22 days under the conditions of 12 hours, and pseudobulb dissolves the individual not grades of 4-8 in leaf scar punishment
Sturdy adventitious bud, and mushrooming out, at 22 days, the adventitious bud length of differentiation to seedling high 3-5cm, blade 4-5 pieces.
4）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band
Part pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C, intensity of illumination 2500lux of room temperature, every daylight
After being cultivated 45 days under the conditions of 12 hours time, pseudobulb is ripe, the bletilla plantlet with pseudobulb is obtained, now, as life
Produce transplantation of seedlings domestication.
The present embodiment, to culture to 108 days bletilla plantlet used times with pseudobulb, Multiplying culture and divides again from seed
Change 67 days used times of culture, the seedling of Fiber differentiation averagely weighs 1.2 times compared with the seedling that conventional medium is cultivated, produces the more traditional training of seedling
Support the production seedling that base is cultivated and averagely weigh 1.5 times, the bletilla plantlet survival rate 98.5% of domestication is transplanted after strong seedling culture, is passed through
The bletilla plantlet survival rate 98.6% of domestication is transplanted after Multiplying culture and dedifferentiation culture.
The method seedling cycle of the bletilla pseudobulb cutting tissue culture of the present invention is short, from seed to culture to false squama
The bletilla plantlet used time of stem relatively shortens 10-20 days using conventional medium, and Multiplying culture and dedifferentiation culture are relatively using tradition
Culture medium shortens 15-25 days, if directly carrying out Multiplying culture and dedifferentiation culture using ripe bletilla pseudobulb, can greatly shorten
Growing-seedling period.Seedling quality is high, and robust plant, the seedling of Fiber differentiation averagely weighs 1.2 times compared with the seedling that conventional medium is cultivated,
Production seedling averagely weighs 1.4 times compared with the production seedling that conventional medium is cultivated, and transplants the survival rate of domestication lowerly generally in 98-98.5%.
Claims (6)
- A kind of 1. bletilla protocorm induction medium, it is characterised in that：Its prescription includes consisting of point（It is single without special instruction Position is mg/L）：1）A great number of elements：Potassium nitrate 2700-2900, ammonium sulfate 461-465, potassium dihydrogen phosphate 390-415, magnesium sulfate 180-190, Calcium chloride 164-166；2）Trace element：Manganese sulfate 22.0-22.5, zinc sulfate 8.2-8.8, boric acid 6.0-6.4, KI 0.80-0.83, molybdenum Sour sodium 0.22-0.26, copper sulphate 0.021-0.025, cobalt chloride 0.023-0.025；3）Molysite：Ferrous sulfate 27.2-27.8, EDTA 37.0-37.43；4）Organic matter：Inositol 100-105, sulfuric acid thiamine element 50-55, sulfuric acid pyridoxol 0.8-1.2, nicotinic acid 0.3-0.5, sweet ammonia Sour 1.8-2.1, glycine betaine 500-520, Choline Chloride 280-315,5- nitroguaiacol sodium salt 0.9-1.2, Sodium p-nitrophenoxide 1.8- 2.2, sodium onitrophenol 1.8- 2.2；5）Additive： AgNO3 2.85-3.10, sucrose 28-33g/L.
- 2. a kind of bletilla breaks up strong seedling culture base, it is characterised in that：Its prescription includes consisting of point（Without special instruction, unit For mg/L）：1）A great number of elements：Potassium nitrate 2700-2900, ammonium sulfate 461-465, potassium dihydrogen phosphate 390-415, magnesium sulfate 180-190, Calcium chloride 164-166；2）Trace element：Manganese sulfate 22.0-22.5, zinc sulfate 8.2-8.8, boric acid 6.0-6.4, KI 0.80-0.83, molybdenum Sour sodium 0.22-0.26, copper sulphate 0.021-0.025, cobalt chloride 0.023-0.025；3）Molysite：Ferrous sulfate 27.2-27.8, EDTA 37.0-37.43；4）Organic matter：Inositol 100-105, sulfuric acid thiamine element 50-55, sulfuric acid pyridoxol 0.8-1.2, nicotinic acid 0.3-0.5, sweet ammonia Sour 1.8-2.1, glycine betaine 500-520, Choline Chloride 280-315,5- nitroguaiacol sodium salt 0.9-1.2, Sodium p-nitrophenoxide 1.8- 2.2, sodium onitrophenol 1.8- 2.2；5）Hormone NAA 0.15-0.226）Additive：AgNO3 2.85-3.10, sucrose 18-22g/L.
- 3. bletilla protocorm induction medium as claimed in claim 1 or bletilla as claimed in claim 2 differentiation strong sprout training Support base, it is characterised in that：The pH value of the culture medium controls：5.5—5.8.
- A kind of 4. method of bletilla pseudobulb cutting tissue culture, it is characterised in that：Comprise the following steps：1）Bletilla protocorm Fiber differentiation seedling：Take the bletilla capsule of maturation to be scrubbed in bromogeramine solution and remove dirt, be placed in On superclean bench, 15-20 min are sterilized with 0.2% mercuric chloride, then with aseptic water washing 3-5 time, the filter paper suction by it with sterilizing After solid carbon dioxide point, cut at the top of fruit, then longitudinally splitted from middle part of fruit, seed is uniformly broadcasted sowing to the bletilla protocorm prepared On stem inducing culture；Bletilla protocorm induction medium after planting be placed in 28 DEG C, intensity of illumination 1500-2000lux, daily Culture 50-65 days is carried out under the light application time environmental condition of 12 hours, obtains the healthy and strong seedling that base portion expands pseudobulb spherical in shape；2）Strong seedling culture：By step 1）The bletilla seedling of Fiber differentiation is timely transferred on differentiation strong seedling culture base, in room temperature 28 DEG C, intensity of illumination 2000-2500lux, under conditions of daily illumination 12 hours, cultivate 50-55 days, pseudobulb is ripe, obtains band There is the bletilla plantlet of pseudobulb；Production transplantation of seedlings domestication can be used as or carry out pseudobulb propagation, again as propagation propagating materials Differentiation culture.
- It is 5. a kind of such as the method for claim 4 bletilla pseudobulb cutting tissue culture, it is characterised in that：It is further comprising the steps of：3）Pseudobulb Multiplying culture：Ripe bletilla plant is peelled off into cauline leaf on sterile platform, coring is cut, pseudobulb It is transferred on fresh differentiation strong seedling culture base, in 28 DEG C of room temperature, intensity of illumination 2000-2500lux, daily illumination 12 hours Under the conditions of cultivate 20-25 days；4）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band one is small Block pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C, intensity of illumination 2000-2500lux of room temperature, often After its illumination is cultivated 45-50 days under conditions of 12 hours, pseudobulb is ripe, obtains the bletilla plantlet with pseudobulb, can conduct Produce transplantation of seedlings domestication or carry out pseudobulb propagation, dedifferentiation culture as propagation propagating materials.
- A kind of 6. method of bletilla pseudobulb cutting tissue culture, it is characterised in that：Comprise the following steps：1）Pseudobulb Multiplying culture：Ripe bletilla plant is peelled off into cauline leaf on sterile platform, coring is cut, pseudobulb It is transferred on fresh differentiation strong seedling culture base, in 28 DEG C of room temperature, intensity of illumination 2000-2500lux, daily illumination 12 hours Under the conditions of cultivate 20-25 days；2）Dedifferentiation culture：The pseudobulb with several adventitious buds is split on sterile platform, each adventitious bud band one is small Block pseudobulb mother tuber, then it is implanted on new differentiation strong seedling culture base, in 28 DEG C, intensity of illumination 2000-2500lux of room temperature, often After its illumination is cultivated 45-50 days under conditions of 12 hours, pseudobulb is ripe, obtains the bletilla plantlet with pseudobulb, can conduct Produce transplantation of seedlings domestication or carry out pseudobulb propagation, dedifferentiation culture as propagation propagating materials.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109526957A (en) * | 2018-11-08 | 2019-03-29 | 河南云帮农业科技有限公司 | A method of promoting bletilla striata pseudobulb fast-growth |
| CN110447540A (en) * | 2019-09-11 | 2019-11-15 | 遵义医科大学 | A kind of tissue cultivating method of bletilla seedling |
| CN110574686A (en) * | 2019-10-24 | 2019-12-17 | 永州职业技术学院 | In-vitro cultivation method of Dendrobium Nanyue |
| CN114342807A (en) * | 2022-01-05 | 2022-04-15 | 广西壮族自治区药用植物园 | In-vitro preservation method of rhizoma bletillae germplasm resources |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1836499A (en) * | 2005-03-24 | 2006-09-27 | 中国科学院遗传与发育生物学研究所 | Cultivation method for highly effective plant tissue differentiation and regeneration |
| CN103651121B (en) * | 2013-11-19 | 2016-01-27 | 安徽振扬农林生态开发有限公司 | A kind of bletilla differentiation, strong seedling culture base |
| CN103651122B (en) * | 2013-11-19 | 2016-01-27 | 安徽振扬农林生态开发有限公司 | A kind of bletilla protocorm induction medium |
-
2017
- 2017-07-26 CN CN201710619723.3A patent/CN107347642A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1836499A (en) * | 2005-03-24 | 2006-09-27 | 中国科学院遗传与发育生物学研究所 | Cultivation method for highly effective plant tissue differentiation and regeneration |
| CN103651121B (en) * | 2013-11-19 | 2016-01-27 | 安徽振扬农林生态开发有限公司 | A kind of bletilla differentiation, strong seedling culture base |
| CN103651122B (en) * | 2013-11-19 | 2016-01-27 | 安徽振扬农林生态开发有限公司 | A kind of bletilla protocorm induction medium |
Non-Patent Citations (1)
| Title |
|---|
| 郑先福: "《植物生长调节剂应用技术》", 30 September 2009, 中国农业大学出版社 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109526957A (en) * | 2018-11-08 | 2019-03-29 | 河南云帮农业科技有限公司 | A method of promoting bletilla striata pseudobulb fast-growth |
| CN110447540A (en) * | 2019-09-11 | 2019-11-15 | 遵义医科大学 | A kind of tissue cultivating method of bletilla seedling |
| CN110574686A (en) * | 2019-10-24 | 2019-12-17 | 永州职业技术学院 | In-vitro cultivation method of Dendrobium Nanyue |
| CN114342807A (en) * | 2022-01-05 | 2022-04-15 | 广西壮族自治区药用植物园 | In-vitro preservation method of rhizoma bletillae germplasm resources |
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