CN104381134A - Rapid mongollian thyme herb proliferation method - Google Patents
Rapid mongollian thyme herb proliferation method Download PDFInfo
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- CN104381134A CN104381134A CN201410745993.5A CN201410745993A CN104381134A CN 104381134 A CN104381134 A CN 104381134A CN 201410745993 A CN201410745993 A CN 201410745993A CN 104381134 A CN104381134 A CN 104381134A
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a rapid mongollian thyme herb proliferation method. The method comprises the following steps: carrying out disinfection treatment to anthers in the late uninucleate stage; inoculating in a callus-inducing medium and culturing to obtain a callus; carrying out multiplication culture, transferring the non-contaminated callus into a first subculture medium for bud proliferation, subculturing for 4 times, transferring into a second subculture medium or a blank culture medium without any hormones for subculturing for one time, then transferring into the first subculture medium for further multiplication, and repeating the steps according to the required seedling amount; inoculating in a rooting medium, and screening to obtain an individual plant. The culture results are that the induction rate of the callus is up to 89%, the rooting rate is up to 85%, the transplanting survival rate can reach 80% and the 10-generation aberration rate can be below 6%.
Description
Technical field
The invention belongs to agricultural technology field, relate to tissue culture technique, be specifically related to a kind of method of Mongollian Thyme Herb fast breeding.
Background technology
Mongollian Thyme Herb, has another name called ground Chinese prickly ash (" marine recipe "), thyme (" botany voluminous dictionary "), Chinese pepper (" illustrated handbook planted by middle traditional Chinese medicines "), lindera glauca (" Liaoning economic plants will ").For labiate.Mainly be distributed in the ground such as Inner Mongol, Ningxia, Qinghai, Shanxi.Can expel pathogenic wind from the body surface in application in TCM, promoting qi circulation and relieving pain, for flu, headache, toothache, abdominal distension crymodynia.
Mongollian Thyme Herb complete stool has strong fragrance, adopts several leaves, rubs in hand, just can send light fragrance, and fragrance gentleness can people.Aromatic oil accounts for 3% of the stem tip and leaf, and main component has fragrant camphor tree alcohol, borneol, to a P-cymene, thymol, amine oleyl alcohol, tannin garlic, saponin, bitter substance and various mineral matter, and wherein potassium, calcium content are higher.There is good beauty treatment, protect stiff anticorrosion, sterilization, help digest, relieve the effect of alcohol, diuresis, the effect such as antirheumatic, its special feature is can strengthening immune system.Therefore Mongollian Thyme Herb has very important medical value.The medical value of Mongollian Thyme Herb is just on the books in Compendium of Material Medica.In the medical research of the former Soviet Union, also on the books to the antisepsis and sterilization characteristic of its leaf flower.In recent years, the chemical composition of Thymus plant is used for medical treatment, health care, cosmetic, food, its potential value developed becomes clear day by day.At present, Mongollian Thyme Herb is wild mostly, is underutilized.
Mongollian Thyme Herb can be sowed, cuttage, press strip, offshoot breeding, except summer, be all well suited for breeding.But these propagation method speed are comparatively slow, are difficult to the object reaching fast breeding.For excavating its medical value, it is possible for obtaining a large amount of aseptic seedling in a short time for extracting the prospect of effective medicinal ingredient with tissue culture quick propagation culturing method." tissue cultures of Mongollian Thyme Herb and plant regeneration " (Zhao Qingzhen, Plant Physiology Communications, 38th volume the 4th phase, in August, 2002) disclose in a literary composition and carry out as vegetative material the method that tissue cultures obtains ground pepper plant fast by the tender stem section of band axillalry bud, " Mongollian Thyme Herb tissue cultures and prevent vitrifying from studying " (Xu Meilong, Agriculture of Anhui science, Journal of Anhui Agri.Sci.2009, 37 (1): 114-116) the stem section also disclosing band axillalry bud in a literary composition is carried out tissue cultures as vegetative material and screens its condition of culture, although reproduction rate is also higher, but easily there is variation find continuous squamous subculture 4 generation in long-term operating process after in inventor, therefore, need continuous inoculated and cultured to obtain callus and could breed more plant, unfavorable to Fast-propagation.
Summary of the invention
For current Problems existing, the invention provides a kind of method of Mongollian Thyme Herb fast breeding, Mongollian Thyme Herb can be bred fast and effectively; and solve the problem of its squamous subculture variation, survival rate is higher, not by the impact in season; significantly reduce propagation cost, meet large-scale production needs.
In order to solve the problem, the invention provides following technical scheme:
A method for Mongollian Thyme Herb fast breeding, comprises
1) the flower pesticide disinfection of mid-late uninucleate stage is got;
2) inoculate callus inducing medium, cultivate and obtain callus;
3) shoot proliferation is cultivated, non-staining callus is proceeded in the first subculture medium and makes Shoot propagation, subculture 4 times, in the blank cultures proceeding to the second subculture medium or do not add any hormone, subculture once, and then proceed to continuation propagation in the first subculture medium, repeat above-mentioned steps according to needing seedling amount;
4) inoculate root media, screening obtains monomer plant.
Described disinfecting is that flower pesticide sterile gauze parcel is immersed in the alcohol of 70-75% concentration the 8-16s that sterilizes, and takes out fast to immerse in bleaching powder immediately and soaks 10min, finally use aseptic water washing 4 ~ 6 times.
Described callus inducing medium is: 1/2MS+6-BA 1.2mg/L+NAA0.12mg/L+ sucrose 50g/L; The PH6.6 of medium, cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1700Lx.
The first described subculture medium is 1/2MS+6-BA1.0mg/L+2,4-D0.2mg/L+NAA 0.3 ~ 0.5mg/L+ agar 3g/L+ sucrose 15g/L; Described second subculture medium is 1/2MS+6-BA 2mg/L+NAA 0.1mg/L+ white granulated sugar 35g/L+ agar powder 7.3g/L; Described blank cultures is 1/2MS+ potassium dihydrogen phosphate 150mg/L+ lactoalbumin hydrolysate 500mg/L+ sucrose 30g/L, and in Subculture, the PH6.5 of medium, cultivation temperature is 25 ± 1 DEG C, the photoperiod of 16h/d, about intensity of illumination 1200Lx.
Described root media is: 1/2MS+IAA 0.1 ~ 0.3mg/L+ sucrose 20 ~ 30g/L+ agar 2 ~ 3g/L+ quality of activated carbon is than 0.1% ~ 0.3%; Condition of culture is: the PH6.5 of medium, and cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1600Lx.
Also comprise in incubation, acclimatization and transplants: first blake bottle is moved to greenhouse, natural conditions lower refining seedling 4d, healthy and strong and when having 5-7 sheet true leaf, 6-8cm height until Mongollian Thyme Herb seedling, then sealed membrane is untied, hardening 6d, after hardening completes, taking-up seedling cleans the medium of root attachment, be transplanted in the matrix of sterilization, temperature controls at 24-26 DEG C, and front 7d humid control is about 70%, reduce humidity gradually later, until under transplanted seedling is placed in natural conditions, suitable shading during acclimatization and transplants, makes intensity of illumination be 50% of natural daylight.
Inventor proves validity of the present invention by series of experiments.
(1) selection of explant
In 5, June, the flower pesticide of gathering, the stem section of band axillalry bud, stem apex, petal are contrasted as explant, all the other conditions are identical, and carry out, in table 1 according to step of the present invention, as can be seen from contrast, adopt flower pesticide as explant, not only the inductivity of callus and rooting rate the highest, 80%, 76% can be reached respectively, and through 5 generation squamous subculture aberration rate minimum, the difficult problem easily occurring variation can be solved in Mongollian Thyme Herb reproductive process.
Table 1
(2) selection of squamous subculture medium
Method 1: 50 callus are inoculated into 1/2MS+6-BA1.0mg/L+2, on 4-D0.2mg/L+NAA 0.3 ~ 0.5mg/L+ agar 3g/L+ sucrose 15g/L medium, continues not change medium all the time in Subculture.
Method 2: other 50 callus proceed in the first subculture medium and make Shoot propagation, subculture 4 times, proceed to the second subculture medium subculture once, and then proceed to continuation propagation in the first subculture medium, first subculture medium is 1/2MS+6-BA1.0mg/L+2,4-D0.2mg/L+NAA 0.3 ~ 0.5mg/L+ agar 3g/L+ sucrose 15g/L; Second subculture medium is 1/2MS+6-BA 2mg/L+NAA 0.1mg/L+ white granulated sugar 35g/L+ agar powder 7.3g/L.
Method 3: also have 50 callus to proceed in the first subculture medium and make Shoot propagation, subculture 4 times, to proceed in the blank cultures not adding any hormone subculture once, and then proceeds in the first subculture medium and continue propagation.First subculture medium is 1/2MS+6-BA1.0mg/L+2,4-D 0.2mg/L+NAA 0.3 ~ 0.5mg/L+ agar 3g/L+ sucrose 15g/L; Blank cultures is 1/2MS+ potassium dihydrogen phosphate 150mg/L+ lactoalbumin hydrolysate 500mg/L+ sucrose 30g/L.
After 10 generation squamous subculture, compare its aberration rate, comparing result, in table 2, from comparative result, is starting Transition medium, can greatly reduce its aberration rate, having conclusive effect to Mongollian Thyme Herb fast breeding after 4 cultures.
Table 2
| Contrast | Aberration rate (%) |
| Method 1 | 43 |
| Method 2 | 4％ |
| Method 3 | 6％ |
Beneficial effect:
(1) the application is using flower pesticide as explant, and inventor operates through a large amount of long-term practice, finds the proterties that can it be kept for a long time excellent as explant in flower pesticide, not easily produces variation;
(2) inventor finds, in factorial seedling growth process, subculture, more than about 4 generations, starts to occur variation, finds after deliberation, above-mentioned reason is that hormone accumulation causes, therefore, in subculture process, have employed the method for Transition medium, reduce the accumulation of hormone to the impact of Mongollian Thyme Herb seedling, this is also one of emphasis of the present invention;
(3) in whole incubation, all adopt 1/2MS medium to replace conventional MS medium, avoid plantlet in vitro excessive growth to affect Bud polarization; The potassium dihydrogen phosphate and lactoalbumin hydrolysate that can promote Bud polarization is added in blank cultures; In subculture process, improve light intensity, in medium, decrease inorganic salt content, add potassium dihydrogen phosphate and lactoalbumin hydrolysate, thus inhibit plantlet in vitro excessive growth, facilitate Bud polarization, achieve unexpected effect;
(4) the present invention has pollen germination and becomes that the embryo time is short, doubling etticiency is high, output is large and be easier to obtain the advantages such as good breeding intermediate materials, at aspects such as breeding material purification and rejuvenation, Germplasm enhancement, polyploid breeding and the researchs of pollen development process, there is larger practical value;
(5) incubation speed of the present invention is fast, and regeneration frequency is higher, and aberration rate is lower, and the cycle is relatively short, can keep the genetic stability of Mongollian Thyme Herb preferably.
Embodiment
Embodiment 1: a kind of method of Mongollian Thyme Herb fast breeding, comprises
1, the selection of explant: by the end of June at full-bloom stage, 9:00 ~ 10:00 in the morning) get the flower pesticide of mid-late uninucleate stage.
2, disinfect: flower pesticide sterile gauze parcel is immersed in the alcohol of 70-75% concentration the 8-16s that sterilizes, take out fast to immerse in bleaching powder immediately and soak 10min, finally use aseptic water washing 4 ~ 6 times.
3, inoculated and cultured: by the explant inoculation callus inducing medium after disinfecting, cultivate and obtain callus.Callus inducing medium is: 1/2MS+6-BA1.2mg/L+NAA 0.12mg/L+ sucrose 50g/L; The PH6.6 of medium, cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1700Lx.
4, shoot proliferation is cultivated, and is proceeded in the first subculture medium by non-staining callus and makes Shoot propagation, subculture 4 times, proceed to the second subculture medium subculture once, and then proceeds to continuation propagation in the first subculture medium, repeats above-mentioned steps according to needing seedling amount;
Wherein: the first subculture medium is 1/2MS+6-BA1.0mg/L+2,4-D0.2mg/L+NAA 0.4mg/L+ agar 3g/L+ sucrose 15g/L;
Second subculture medium is 1/2MS+6-BA 2mg/L+NAA 0.1mg/L+ white granulated sugar 35g/L+ agar powder 7.3g/L;
Condition of culture is: the PH6.5 of medium, and cultivation temperature is 25 ± 1 DEG C, the photoperiod of 16h/d, about intensity of illumination 1200Lx.
5, culture of rootage: by the Mongollian Thyme Herb inoculation root media through squamous subculture, screening obtains monomer plant.
Wherein, root media is: 1/2MS+IAA 0.2mg/L+ sucrose 30g/L+ agar 2g/L+ quality of activated carbon is than 0.2%;
Condition of culture is: the PH6.5 of medium, and cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1600Lx.
6, acclimatization and transplants: first blake bottle is moved to greenhouse, natural conditions lower refining seedling 4d, healthy and strong and when having 5-7 sheet true leaf, 6-8cm height until Mongollian Thyme Herb seedling, then sealed membrane is untied, hardening 6d, after hardening completes, taking-up seedling cleans the medium of root attachment, be transplanted in the matrix of sterilization, temperature controls at 24-26 DEG C, front 7d humid control, about 70%, reduces humidity later gradually, until under transplanted seedling is placed in natural conditions, suitable shading during acclimatization and transplants, makes intensity of illumination be 50% of natural daylight.
Embodiment 2: a kind of method of Mongollian Thyme Herb fast breeding, comprises
Except in shoot proliferation incubation, non-staining callus is proceeded in the first subculture medium and makes Shoot propagation, after subculture 4 times, proceed to blank cultures subculture once, and then proceed to continuation propagation in the first subculture medium, wherein blank cultures is that outside 1/2MS+ potassium dihydrogen phosphate 150mg/L+ lactoalbumin hydrolysate 500mg/L+ sucrose 30g/L, all the other are with embodiment 1.
Embodiment 3
Except " root media is: 1/2MS+IAA 0.1mg/L+ sucrose 20g/L+ agar 3g/L+ quality of activated carbon is than 0.3% ", all the other are with embodiment 1.
Any combination of various condition above, all can reach consistent effect.
Cultivation results of the present invention is: the inductivity of callus reaches 89%, and rooting rate reaches 85%, and transplanting survival rate reaches 80%, 10 generation aberration rate below 6%.
Claims (6)
1. a method for Mongollian Thyme Herb fast breeding, is characterized in that, comprises
1) the flower pesticide disinfection of mid-late uninucleate stage is got;
2) inoculate callus inducing medium, cultivate and obtain callus;
3) shoot proliferation is cultivated, non-staining callus is proceeded in the first subculture medium and makes Shoot propagation, subculture 4 times, in the blank cultures proceeding to the second subculture medium or do not add any hormone, subculture once, and then proceed to continuation propagation in the first subculture medium, repeat above-mentioned steps according to needing seedling amount;
4) inoculate root media, screening obtains monomer plant.
2. the method for Mongollian Thyme Herb fast breeding according to claim 1, it is characterized in that, described disinfecting is that flower pesticide sterile gauze parcel is immersed in the alcohol of 70-75% concentration the 8-16s that sterilizes, and takes out fast to immerse in bleaching powder immediately and soaks 10min, finally use aseptic water washing 4 ~ 6 times.
3. the method for Mongollian Thyme Herb fast breeding according to claim 1 and 2, is characterized in that, described callus inducing medium is: 1/2MS+6-BA 1.2mg/L+NAA0.12mg/L+ sucrose 50g/L; The PH of medium is 6.6, and cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1700Lx.
4. the method for Mongollian Thyme Herb fast breeding according to claim 1 and 2, is characterized in that, the first described subculture medium is 1/2MS+6-BA1.0mg/L+2,4-D 0.2mg/L+NAA0.3 ~ 0.5mg/L+ agar 3g/L+ sucrose 15g/L; Described second subculture medium is 1/2MS+6-BA 2mg/L+NAA 0.1mg/L+ white granulated sugar 35g/L+ agar powder 7.3g/L; Described blank cultures is 1/2MS+ potassium dihydrogen phosphate 150mg/L+ lactoalbumin hydrolysate 500mg/L+ sucrose 30g/L, and in Subculture, the PH6.5 of medium, cultivation temperature is 25 ± 1 DEG C, the photoperiod of 16h/d, about intensity of illumination 1200Lx.
5. the method for Mongollian Thyme Herb fast breeding according to claim 1 and 2, is characterized in that, described root media is: 1/2MS+IAA 0.1 ~ 0.3mg/L+ sucrose 20 ~ 30g/L+ agar 2 ~ 3g/L+ quality of activated carbon is than 0.1% ~ 0.3%; Condition of culture is: the PH6.5 of medium, and cultivation temperature is 25 ± 1 DEG C, the photoperiod of 12h/d, about intensity of illumination 1600Lx.
6. the method for Mongollian Thyme Herb fast breeding according to claim 1 and 2, it is characterized in that, also comprise, acclimatization and transplants: first blake bottle is moved to greenhouse, natural conditions lower refining seedling 4d, treat Mongollian Thyme Herb seedling stalwartness and have 5-7 sheet true leaf, during 6-8cm height, then sealed membrane is untied, hardening 6d, after hardening completes, taking-up seedling cleans the medium of root attachment, be transplanted in the matrix of sterilization, temperature controls at 24-26 DEG C, front 7d humid control is about 70%, reduce humidity gradually later, until under transplanted seedling is placed in natural conditions, suitable shading during acclimatization and transplants, intensity of illumination is made to be 50% of natural daylight.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201410745993.5A CN104381134B (en) | 2014-12-08 | 2014-12-08 | A kind of method of Herba Thymi fast breeding |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201410745993.5A CN104381134B (en) | 2014-12-08 | 2014-12-08 | A kind of method of Herba Thymi fast breeding |
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| CN104381134A true CN104381134A (en) | 2015-03-04 |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104813931A (en) * | 2015-03-11 | 2015-08-05 | 常州展华机器人有限公司 | Tissue culture and rapid propagation method for Dendrobium officinale |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102388807A (en) * | 2011-10-27 | 2012-03-28 | 北京市园林学校 | Rapid propagation method for tissue culture of wild Thymus Mongolicus Ronn and application thereof |
| CN102550403A (en) * | 2011-11-04 | 2012-07-11 | 北京市园林学校 | Tissue culture rootage and transplantation methods for wild thymus mongolicus ronn |
| CN103858770A (en) * | 2014-04-03 | 2014-06-18 | 中国科学院植物研究所 | Rapid hosta plantagineu propagation method |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102388807A (en) * | 2011-10-27 | 2012-03-28 | 北京市园林学校 | Rapid propagation method for tissue culture of wild Thymus Mongolicus Ronn and application thereof |
| CN102550403A (en) * | 2011-11-04 | 2012-07-11 | 北京市园林学校 | Tissue culture rootage and transplantation methods for wild thymus mongolicus ronn |
| CN103858770A (en) * | 2014-04-03 | 2014-06-18 | 中国科学院植物研究所 | Rapid hosta plantagineu propagation method |
Non-Patent Citations (1)
| Title |
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| 王玲等: "东北百里香组培再生体系的建立", 《园艺学报》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104813931A (en) * | 2015-03-11 | 2015-08-05 | 常州展华机器人有限公司 | Tissue culture and rapid propagation method for Dendrobium officinale |
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