CN111280066A - Culture method of horseradish flower moss callus capable of effectively inhibiting browning - Google Patents
Culture method of horseradish flower moss callus capable of effectively inhibiting browning Download PDFInfo
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- CN111280066A CN111280066A CN202010281668.3A CN202010281668A CN111280066A CN 111280066 A CN111280066 A CN 111280066A CN 202010281668 A CN202010281668 A CN 202010281668A CN 111280066 A CN111280066 A CN 111280066A
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- NKZMPZCWBSWAOX-IBTYICNHSA-M Sulbactam sodium Chemical compound [Na+].O=S1(=O)C(C)(C)[C@H](C([O-])=O)N2C(=O)C[C@H]21 NKZMPZCWBSWAOX-IBTYICNHSA-M 0.000 claims abstract description 7
- 229960004682 cefoperazone Drugs 0.000 claims abstract description 7
- GCFBRXLSHGKWDP-XCGNWRKASA-N cefoperazone Chemical compound O=C1C(=O)N(CC)CCN1C(=O)N[C@H](C=1C=CC(O)=CC=1)C(=O)N[C@@H]1C(=O)N2C(C(O)=O)=C(CSC=3N(N=NN=3)C)CS[C@@H]21 GCFBRXLSHGKWDP-XCGNWRKASA-N 0.000 claims abstract description 7
- 229960000614 sulbactam sodium Drugs 0.000 claims abstract description 7
- 229960002727 cefotaxime sodium Drugs 0.000 claims abstract description 6
- AZZMGZXNTDTSME-JUZDKLSSSA-M cefotaxime sodium Chemical compound [Na+].N([C@@H]1C(N2C(=C(COC(C)=O)CS[C@@H]21)C([O-])=O)=O)C(=O)\C(=N/OC)C1=CSC(N)=N1 AZZMGZXNTDTSME-JUZDKLSSSA-M 0.000 claims abstract description 6
- 238000000034 method Methods 0.000 claims abstract description 5
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- LKDRXBCSQODPBY-AMVSKUEXSA-N L-(-)-Sorbose Chemical compound OCC1(O)OC[C@H](O)[C@@H](O)[C@@H]1O LKDRXBCSQODPBY-AMVSKUEXSA-N 0.000 claims description 6
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 claims description 6
- 229930006000 Sucrose Natural products 0.000 claims description 6
- 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 6
- 230000001954 sterilising effect Effects 0.000 claims description 6
- 239000005720 sucrose Substances 0.000 claims description 6
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 claims description 6
- 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 claims description 5
- IMQLKJBTEOYOSI-GPIVLXJGSA-N Inositol-hexakisphosphate Chemical compound OP(O)(=O)O[C@H]1[C@H](OP(O)(O)=O)[C@@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@@H]1OP(O)(O)=O IMQLKJBTEOYOSI-GPIVLXJGSA-N 0.000 claims description 5
- IMQLKJBTEOYOSI-UHFFFAOYSA-N Phytic acid Natural products OP(O)(=O)OC1C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C1OP(O)(O)=O IMQLKJBTEOYOSI-UHFFFAOYSA-N 0.000 claims description 5
- 238000005520 cutting process Methods 0.000 claims description 5
- 230000000249 desinfective effect Effects 0.000 claims description 5
- 239000008103 glucose Substances 0.000 claims description 5
- 150000002505 iron Chemical class 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
- 229940068041 phytic acid Drugs 0.000 claims description 5
- 235000002949 phytic acid Nutrition 0.000 claims description 5
- 239000000467 phytic acid Substances 0.000 claims description 5
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Inorganic materials [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 claims description 5
- 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 3
- 238000001035 drying Methods 0.000 claims description 3
- 238000004659 sterilization and disinfection Methods 0.000 claims description 3
- 235000003222 Helianthus annuus Nutrition 0.000 claims description 2
- 244000179886 Moringa oleifera Species 0.000 claims description 2
- 235000011347 Moringa oleifera Nutrition 0.000 claims description 2
- 244000020551 Helianthus annuus Species 0.000 claims 1
- 238000011160 research Methods 0.000 abstract description 4
- 239000000463 material Substances 0.000 abstract description 2
- 230000000052 comparative effect Effects 0.000 description 7
- 244000195452 Wasabia japonica Species 0.000 description 4
- 235000000760 Wasabia japonica Nutrition 0.000 description 4
- -1 isothiocyanate compound Chemical class 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 3
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- 240000008892 Helianthus tuberosus Species 0.000 description 2
- 235000003230 Helianthus tuberosus Nutrition 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 244000026811 Brassica nipposinica Species 0.000 description 1
- 241000219193 Brassicaceae Species 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
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- 241000628997 Flos Species 0.000 description 1
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- 206010028980 Neoplasm Diseases 0.000 description 1
- 208000004880 Polyuria Diseases 0.000 description 1
- 230000003260 anti-sepsis Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 230000035619 diuresis Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 239000012476 oxidizable substance Substances 0.000 description 1
- 239000002352 surface water Substances 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
- A01H4/005—Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques
-
- 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
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- 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 culture method of horseradish flower moss callus capable of effectively inhibiting browning, aiming at the condition that a large amount of death of a culture is caused by serious browning of a horseradish flower moss explant in the callus induction and mass multiplication culture processes, a sterilized material washed and disinfected by sterile water containing cefoperazone sulbactam sodium is innovatively used, the components of a callus induction and multiplication culture medium are improved, and meanwhile, a step of pretreating by using a water solution containing cefotaxime sodium is added before the horseradish callus multiplication culture. The invention effectively solves the problem that the horseradish flower moss explant is easy to generate brown stain in the callus induction and multiplication culture processes, and provides a new way for producing the effective components of the horseradish plant; meanwhile, the invention also lays a certain theoretical foundation for further developing the biotechnology research of the horseradish plants.
Description
Technical Field
The invention relates to a culture method of horseradish flower moss calluses, in particular to a culture method of horseradish flower moss calluses capable of effectively inhibiting browning.
Background
Wasabi (Wasabia japonica Matsum), also known as Wasabia japonica and potherb mustard, is a perennial semipudendum herbaceous plant of Wasabia japonica of Brassicaceae.
The whole plant of horseradish contains a substance of sinapside, when plant cells are ground, the substance is hydrolyzed under the action of sinapside enzyme to form an isothiocyanate compound, and the isothiocyanate compound has multiple medicinal activities of sterilization, antisepsis, diuresis, blood cleaning, infection resistance, cancer resistance and the like. However, the growth and development of the horseradish plants have extremely strict requirements on the environment, the horseradish plants are only suitable for being planted in high mountain areas with the altitude of 1000-2000 meters, the growth temperature is 8-22 ℃, the optimum temperature is 12-15 ℃, the horseradish plants are not suitable for being planted when the temperature exceeds 23 ℃, and soft rot easily occurs when the temperature is 23-30 ℃; is easy to be frozen when the temperature is lower than minus 3 ℃; therefore, the horseradish resource is very rare, and the market demand can not be met at present. And the development of the biotechnology research of the horseradish, mass production of the horseradish callus with high content of the isothiocyanate compounds by a tissue culture mode is an effective way for obtaining the effective components in the horseradish plant.
At present, researchers have conducted researches on callus induction on various organs of different parts of a horseradish plant, but researches on callus induction and propagation culture by selecting a horseradish plant flower moss with high biological yield and high effective component content as an explant are hardly reported. Because the cells of the horseradish tree contain more isothiocyanate compounds and other easily-oxidizable substances, serious browning is often generated in the callus induction and mass multiplication culture processes to cause mass death of culture materials, which is also a big problem in the mass production of effective components of the horseradish by utilizing the biotechnology at present. At present, no relevant solution is reported.
Disclosure of Invention
The invention aims to provide a culture method of horseradish moss callus capable of effectively inhibiting browning, which can effectively inhibit the problem that the horseradish moss callus is easy to brown in induction and multiplication culture.
In order to achieve the above object, the solution adopted by the present invention comprises the following steps:
(1) selecting and disinfecting explants: selecting flower moss with the length of 10-20 cm under the terminal bud of the flower moss of the mountain sunflower as an explant, and sterilizing the flower moss and then drying water on the surface of the flower moss by using sterile filter paper;
(2) callus induction culture: cutting the disinfected horseradish lawn into the length of 1-2 cm, then quickly inoculating the horseradish lawn into a callus induction culture medium of improved MS +6-BA 0.5-1.5 mg/L +2, 4-D2.0-4.0 mg/L + tyrosine 50-100 g/L + glucose 10-20 g/L + sorbose 5-15 g/L + agar 4.5-5.5 g/L, and culturing under the conditions that the culture temperature is 18-22 ℃, the illumination is 4-8 h every day and the illumination intensity is 300-500 lx, wherein the iron salt of the improved MS basic culture medium is changed into 55.6mg/L, KNO3Changing to 316-950 mg/L;
(3) callus proliferation pre-culture: selecting green callus, putting the green callus into cefotaxime sodium sterile water with the concentration of 50-100 mg/L, and pre-culturing for 12-24 hours under the conditions that the rotating speed of a shaking table is 100-200 r/min and no illumination exists;
(4) and (3) callus proliferation culture: inoculating the pre-cultured callus into a callus proliferation culture medium of White +6-BA 1.0-2.0 mg/L +2, 4-D1-3 mg/L + NAA 0.5-1 mg/L + phytic acid 2.0-6.0 ml/L + sucrose 10-20 g/L + EDTA 10-20 g/L + agar 4.0-6.0 g/L, and performing proliferation culture under the conditions of a culture temperature of 14-18 ℃, illumination for 4-6 h every day and illumination intensity of 600-800 lx;
all the above media had a pH of 5.5.
Further, the disinfection in the step (1) is to disinfect with 0.1% mercuric chloride for 4-8 min, and then wash with 40-80 mg/L cefoperazone sulbactam sodium sterile water for 3-6 times.
The invention effectively solves the problem that the horseradish flower moss explant is easy to generate brown stain in the callus induction and multiplication culture processes, and provides a new way for producing effective components of horseradish plants.
Detailed Description
The present invention will be described in further detail with reference to examples.
Example 1
The culture method of the horseradish lawn callus capable of effectively inhibiting browning provided by the embodiment comprises the following steps:
(1) selecting and disinfecting explants: selecting flower moss 15cm in length below terminal bud of flos Helianthi Mollissimae as explant, placing on an ultraclean workbench, sterilizing with 0.1% mercuric chloride for 6min, washing with 60mg/L cefoperazone sulbactam sodium sterile water for 5 times, and drying surface water with sterile filter paper;
(2) callus induction culture: cutting sterilized horseradish flower moss into 1.5cm in length, quickly inoculating into improved MS +6-BA1.0mg/L +2,4-D3.0mg/L + tyrosine 75g/L + glucose 15g/L + sorbose 10g/L + agar 5.0g/L callus induction culture medium, culturing at 20 deg.C under illumination for 6 hr per day and illumination intensity of 400lx, and changing iron salt of the improved MS basic culture medium into 55.6mg/L, KNO3Changed to 633 mg/L; the induction rate of the callus is 87.74% and the browning rate of the callus is 0% after 30 days of culture;
(3) callus proliferation pre-culture: selecting green callus, putting the green callus into cefotaxime sodium sterile water with the concentration of 80mg/L, and pre-culturing for 18 hours at the rotating speed of a shaker of 150r/min under the condition of no illumination;
(4) and (3) callus proliferation culture: inoculating the callus after proliferation pre-culture into a callus proliferation culture medium of White +6-BA1.5mg/L +2,4-D2.0mg/L + NAA 0.75mg/L + phytic acid 4.0ml/L + sucrose 15g/L + EDTA15g/L + agar 5.0g/L, performing proliferation culture at the culture temperature of 16 ℃, under the conditions of illumination for 5h every day and illumination intensity of 700lx, counting the proliferation multiple of the callus to be 2.89 times after 30 days of culture, and counting the browning rate of the proliferation callus to be 0%;
all the above media had a pH of 5.5.
Example 2
The culture method of the horseradish lawn callus capable of effectively inhibiting browning provided by the embodiment comprises the following steps:
(1) selecting and disinfecting explants: selecting flower moss with a length of 10cm under terminal bud of the flower moss of Helianthus tuberosus as an explant, placing the flower moss on an ultraclean workbench, sterilizing with mercuric chloride with a concentration of 0.1% for 4min, washing with sterile water containing 40mg/L cefoperazone sulbactam sodium for 3 times, and then sucking water on the surface with sterile filter paper;
(2) callus induction culture: cutting sterilized horseradish flower moss into 1cm in length, quickly inoculating the horseradish flower moss into a callus induction culture medium of improved MS +6-BA0.5mg/L +2,4-D2.0mg/L + tyrosine 50g/L + glucose 10g/L + sorbose 5g/L + agar 4.5g/L, and culturing under the conditions that the culture temperature is 18 ℃, the illumination is 4h per day and the illumination intensity is 300lx, wherein the iron salt of the improved MS basic culture medium is changed into 55.6mg/L, KNO3Changing to 316 mg/L; the induction rate of the callus is 79.84 percent and the browning rate of the callus is 7.80 percent when the callus is cultured for 30 days;
(3) callus proliferation pre-culture: selecting green callus, putting the green callus into cefotaxime sodium sterile water with the concentration of 50mg/L, and pre-culturing for 12 hours under the condition that the rotating speed of a shaking table is 100r/min and no light is emitted;
(4) and (3) callus proliferation culture: inoculating the callus after proliferation pre-culture into a callus proliferation culture medium of White, 6-BA1.0mg/L +2,4-D1.0 mg/L + NAA 0.5mg/L + phytic acid 2.0ml/L + sucrose 10g/L + EDTA10 g/L + agar 4.0g/L, performing proliferation culture at the culture temperature of 14 ℃, under the conditions of illumination for 4h every day and illumination intensity of 600lx, counting the proliferation multiple of the callus to be 2.34 times after 30 days of culture, and counting the browning rate of the proliferation callus to be 4.79%;
all the above media had a pH of 5.5.
Example 3
The culture method of the horseradish lawn callus capable of effectively inhibiting browning provided by the embodiment comprises the following steps:
(1) selecting and disinfecting explants: selecting flower moss with a length of 20cm under terminal buds of the flower moss of Helianthus tuberosus as an explant, placing the flower moss on an ultraclean workbench, sterilizing for 8min by using mercuric chloride with a concentration of 0.1%, washing for 6 times by using sterile water of cefoperazone sulbactam sodium with a concentration of 80mg/L, and then sucking water on the surface by using sterile filter paper;
(2) callus induction culture: cutting sterilized horseradish flower moss into 2cm in length, quickly inoculating the horseradish flower moss into an improved MS +6-BA1.5mg/L +2,4-D4.0mg/L + tyrosine 100g/L + glucose 20g/L + sorbose 15g/L + agar 5.5g/L callus induction culture medium, and culturing under the conditions that the culture temperature is 22 ℃, the illumination is 8 hours per day and the illumination intensity is 500lx, wherein the iron salt of the improved MS basic culture medium is changed into 55.6mg/L, KNO3Changed to 950 mg/L; the induction rate of the callus is 88.21% and the browning rate of the callus is 8.70% after 30 days of culture;
(3) callus proliferation pre-culture: selecting green callus, putting the green callus into 100mg/L cefotaxime sodium sterile water, and pre-culturing for 24 hours at the rotating speed of a shaker of 200r/min under the condition of no illumination;
(4) and (3) callus proliferation culture: inoculating the callus after proliferation pre-culture into a callus proliferation culture medium of White, 6-BA 2.0mg/L +2,4-D3.0mg/L + NAA 1.0mg/L + phytic acid 6.0ml/L + sucrose 20g/L + EDTA 20g/L + agar 6.0g/L, performing proliferation culture at the culture temperature of 18 ℃, under the conditions of illumination for 6h every day and illumination intensity of 800lx, counting the proliferation multiple of the callus to be 2.54 times after 30 days of culture, and counting the browning rate of the proliferation callus to be 8.55%;
all the above media had a pH of 5.5.
Comparative example 1
In the step (1) of example 1, the sterilized horseradish lawn was washed with sterile water without adding cefoperazone sulbactam sodium, and the other steps were the same as in example 1, and the induction rate of the callus was 61.14% and the browning rate of the callus was 50.97% as counted by 30-day culture.
Comparative example 2
The minimal medium in the callus induction medium in the step (2) of the example 1 is changed from modified MS to MS, and other steps are the same as the example 1; the induction rate of the callus is 33.64% and the browning rate of the callus is 35.21% by statistics of 30 days of culture.
Comparative example 3
Tyrosine and sorbose in the callus induction medium in step (2) of example 1 were removed, and the other steps were the same as in example 1; the statistics of the culture for 30 days show that the induction rate of the callus is 68.64 percent, and the browning rate of the callus is 49.21 percent.
Comparative example 4
The culture conditions in the step (2) of example 1 were changed to culture under the conditions of a culture temperature of 28 ℃, 12 hours of light per day and 1000lx of light intensity, and the induction rate of the callus was 72.14% and the browning rate of the callus was 50.03% after 30 days of culture.
Comparative example 5
The callus proliferation preculture step (3) in example 1 was omitted, and other steps were the same as in example 1, and the callus proliferation was 2.10-fold as counted for 30 days of proliferation culture, and the browning rate of the proliferated callus was 46.3%.
Comparative example 6
The callus proliferation medium in step (4) of example 1 was changed to MS +6-BA3.0mg/L + NAA0.1mg/LL + sucrose 30g/L + agar 7.0g/L, and other steps were the same as in example 1, and the proliferation of the callus was 1.82-fold, and the browning rate of the proliferated callus was 42.37%.
Comparative example 7
The callus proliferation culture conditions in the step (4) of example 1 were changed to 28 ℃ culture temperature, 16 hours of light per day and 1500lx light intensity, and other steps were the same as in example 1, and statistics on 30 days of culture revealed that the callus proliferated 2.82 times and the browning rate of the proliferated callus was 51.14%.
Claims (2)
1. A culture method of horseradish flower moss callus capable of effectively inhibiting browning is characterized by comprising the following steps:
(1) selecting and disinfecting explants: selecting flower moss with the length of 10-20 cm under the terminal bud of the flower moss of the mountain sunflower as an explant, and sterilizing the flower moss and then drying water on the surface of the flower moss by using sterile filter paper;
(2) callus induction culture: cutting the disinfected horseradish lawn into the length of 1-2 cm, then quickly inoculating the horseradish lawn into a callus induction culture medium of improved MS +6-BA 0.5-1.5 mg/L +2, 4-D2.0-4.0 mg/L + tyrosine 50-100 g/L + glucose 10-20 g/L + sorbose 5-15 g/L + agar 4.5-5.5 g/L, and culturing under the conditions that the culture temperature is 18-22 ℃, the illumination is 4-8 h every day and the illumination intensity is 300-500 lx, wherein the iron salt of the improved MS basic culture medium is changed into 55.6mg/L, KNO3Changing to 316-950 mg/L;
(3) callus proliferation pre-culture: selecting green callus, putting the green callus into cefotaxime sodium sterile water with the concentration of 50-100 mg/L, and pre-culturing for 12-24 hours under the conditions that the rotating speed of a shaking table is 100-200 r/min and no illumination exists;
(4) and (3) callus proliferation culture: inoculating the pre-cultured callus into a callus proliferation culture medium of White +6-BA 1.0-2.0 mg/L +2, 4-D1.0-3.0 mg/L + NAA 0.5-1 mg/L + phytic acid 2.0-6.0 ml/L + sucrose 10-20 g/L + EDTA 10-20 g/L + agar 4.0-6.0 g/L, and performing proliferation culture under the conditions of a culture temperature of 14-18 ℃, illumination for 4-6 h every day and illumination intensity of 600-800 lx;
all the above media had a pH of 5.5.
2. The method for culturing callus of horseradish tree flower moss capable of effectively inhibiting browning according to claim 1, wherein the culture method comprises the following steps: the disinfection in the step (1) is to disinfect for 4-8 min by using 0.1% mercuric chloride, and then wash with 40-80 mg/L cefoperazone sulbactam sodium sterile water for 3-6 times.
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| CN103704137A (en) * | 2013-12-24 | 2014-04-09 | 成都大学 | Culturing method capable of effectively reducing reverse mutation rate of horseradish polyploid plants |
| CN104140978A (en) * | 2014-07-23 | 2014-11-12 | 成都大学 | Genetic transformation method using horseradish seed leaves as explant |
| WO2015168733A1 (en) * | 2014-05-05 | 2015-11-12 | Queensland University Of Technology | Decoy molecules |
-
2020
- 2020-04-10 CN CN202010281668.3A patent/CN111280066B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5386333A (en) * | 1977-01-07 | 1978-07-29 | Ajinomoto Kk | Increasing method for nursery horseeradish |
| CN102823491A (en) * | 2012-08-06 | 2012-12-19 | 成都大学 | Method for culturing horseradish callus through taking horseradish root as explant |
| CN103704137A (en) * | 2013-12-24 | 2014-04-09 | 成都大学 | Culturing method capable of effectively reducing reverse mutation rate of horseradish polyploid plants |
| WO2015168733A1 (en) * | 2014-05-05 | 2015-11-12 | Queensland University Of Technology | Decoy molecules |
| CN104140978A (en) * | 2014-07-23 | 2014-11-12 | 成都大学 | Genetic transformation method using horseradish seed leaves as explant |
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