CN103704135B - In-vitro rapid propagation method for plantains - Google Patents
In-vitro rapid propagation method for plantains Download PDFInfo
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- CN103704135B CN103704135B CN201310710565.4A CN201310710565A CN103704135B CN 103704135 B CN103704135 B CN 103704135B CN 201310710565 A CN201310710565 A CN 201310710565A CN 103704135 B CN103704135 B CN 103704135B
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Abstract
The invention provides an in-vitro rapid propagation method for plantains. The in-vitro rapid propagation method for the plantains comprises the following steps: (1) callus induction culture, namely cutting off leaf veins and leaf margins of sterilized and disinfected plantains and cutting into small blocks; inoculating the plantains to a callus induction culture medium; and carrying out regular culture to form calluses and adventitious buds; and (2) rooting culture, namely selecting the adventitious buds, with the bud lengths of not more than 2cm-3cm, obtained in the step (1); cutting off the adventitious buds from the calluses and continuing to carry out callus induction culture on the residual calluses and the adventitious buds; and inoculating the cut adventitious buds on a rooting culture medium; and regularly culturing until white roots grow to obtain aseptic plantain germchits. According to the method provided by the invention, the disinfection efficiency of explants is up to 80% and the aseptic seedling rooting rate is 100%; the aseptic plantain germchits can be rapidly propagated; a plurality of problems that the propagation capability of plantain plants is low, the propagation period is long, resources are lacked and a lot of pesticides are remained are solved; the in-vitro rapid propagation method for the plantains has very important meanings in the aspects of protecting the natural resources of the wild plantains, protecting a good ecological environment and the like.
Description
Technical field
The invention belongs to the field of tissue culture of plant, be specifically related to a kind of method of Asiatic plantain Vitro Quick Reproduction.
Background technology
Plantago is in Plantaginaceae plant, and another name be again Niu Tiancao, Che Qiancai, wheel dish, pig Auricled Hedyotis Herb etc., long in a humid environment, as pond, set under, river bank, roadside etc.Asiatic plantain has edible and food therapy value, is rich in multivitamin and protein, containing enriching excellent soluble dietary fiber.
The seed of Asiatic plantain and herb all can be used as medicine, and toxic and side effect is less, are a kind of excellent medicinal plants, containing multiple medicinal component, as ursolic acid, plantagin, cupreol etc., there is inducing diuresis for removing edema, antidiarrheal, heat-clearing sterilization, improving eyesight reduced phlegm, anti-inflammatory, the effect such as hypotensive; β-Biao logaric acid (β-epiloganic acid), daucosterol and galuteolin three kinds of compositions of containing not only can prevent aging by the oxygen radical in purged body, but also there is antidepressant effect, the application of clinicing aspect has: treatment vaginitis, treatment acute mastitis, treatment postpartum retention of urine, treatment latent nephritis, treatment bedsore, treatment CAH, treatment exercise hemoglobinuria etc.; Also there is certain diuresis, blood urine, flu and diarrhoea can be treated, treatment gynaecological disease (as leukorrhea), treatment intractable toothache.Also have with Asiatic plantain is at present (as the ointment, syrup, health care buccal tablets, health drink etc.) that preparation produced by raw material.In addition, the materials such as aliphatic acid, adenine such as medicinal chemistry composition and plantaglucide, Catalpol, choline, arachidic acid and leukotrienes such as ursolic acid B-sitosterol, plantagin Aucubin, benzyl carbinol glycosides, Cobastab, positive hentriacontane are also contained in Asiatic plantain.
Breeding at present for Asiatic plantain only has traditional planting technology.But because long-term over-exploitation causes that wild Asiatic plantain reduces gradually, seed carries virus, environmental condition worsens shortcomings such as causing that the difference of medical value, breeding cycle length, fine quality degeneration and residue of pesticide amount exceed standard, all seriously have impact on the exploitation of Asiatic plantain.Research Asiatic plantain organizes cultured in vitro quick breeding technology to be solve the effective ways that current Asiatic plantain utilizes predicament.
Summary of the invention
For solving during current Asiatic plantain utilizes the problems such as seed takes poison, breeding cycle length, quality deterioration, medical value is uneven, residue of pesticide amount exceeds standard existed, the invention provides a kind of method of Asiatic plantain Vitro Quick Reproduction.
The method of Asiatic plantain Vitro Quick Reproduction provided by the invention, comprises the steps:
1) induction of callus: the Asiatic plantain blade through sterilization is cut vein and leaf margin, is cut into small pieces, be inoculated on callus inducing medium, cellar culture is to forming callus and adventitious bud;
2) culture of rootage: selecting step 1) in bud reach the adventitious bud of 2-3 centimetre, it is cut from callus, all the other callus and adventitious bud continue induction of callus, the adventitious bud cut is inoculated on root media, cellar culture, to the root growing white, obtains aseptic Asiatic plantain seedling.
Wherein, the concrete grammar of described sterilization is: pluck without the Asiatic plantain blade of disease and pest, running water, superclean bench is used alcohol-pickled 30s, then soaks 8min in the mercuric chloride solution proceeding to 0.1%, with aseptic water washing 3-5 time.
Wherein, the fritter of the preferred 0.8mm × 0.8mm of described fritter.
Wherein, described callus inducing medium is with MS culture medium for minimal medium, and contained sucrose concentration is 10 ~ 50g/L, methyl α-naphthyl acetate (NAA) concentration is 0.05 ~ 0.5mg/L, 6-benzyl aminopurine (6-BA) concentration is the culture medium of 1.0 ~ 4.0mg/L.
Preferably, described callus inducing medium is for minimal medium with MS culture medium, contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 1.0mg/L, or with MS culture medium for minimal medium, contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L.
Wherein, described root media is with MS culture medium for minimal medium, and contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L.
Wherein, the condition of culture of described cellar culture is pH value 6.0, cultivation temperature 25 DEG C, periodicity of illumination 12 hours/day, and intensity of illumination is 2000lx.
Wherein, the time of described induction of callus is 30 days.
Wherein, the time of described culture of rootage is 15 days.
The present invention also provides the application of method in Asiatic plantain breeds of described Asiatic plantain Vitro Quick Reproduction.
Beneficial effect of the present invention is:
1) explant disinfection efficiency of the present invention is up to 80%, and aseptic seedling rooting rate reaches 100%, can the aseptic seedling of Fast-propagation Asiatic plantain.
2) fertility that the invention solves Asiatic plantain plant is low, the problems such as breeding cycle length, scarcity of resources, residues of pesticides.
3) the present invention adopts Vitro Quick Reproduction technology to carry out tissue cultures to Asiatic plantain first, not only can meet the need of market, but also can provide useful theoretical reference for factory's large-scale production.
4) the present invention is significant in protection wild Asiatic plantain natural resources and good ecological environment etc.
Detailed description of the invention
Following examples for illustration of the present invention, but are not used for limiting the scope of the invention.Without departing from the spirit and substance of the case in the present invention, the amendment do the inventive method, step or condition or replacement, all belong to scope of the present invention.
If do not specialize, the conventional means that technological means used in embodiment is well known to those skilled in the art.In embodiment, chemical reagent used is all commercially available.
Embodiment 1
The present invention is utilized to carry out Asiatic plantain Vitro Quick Reproduction.
The selection of explant and sterilization: pluck the wild Asiatic plantain blade without disease and pest from field, dust is washed away under flowing water, alcohol-pickled 30s first used by superclean bench, proceed to again in the mercuric chloride solution of 0.1% and soak 8min, with aseptic water washing 3-5 time, last vein and the edge aseptically cutting Asiatic plantain blade, is cut into the fritter of 0.8mm × 0.8mm.
1) induction of callus: 0.8mm × 0.8mm fritter that the Asiatic plantain blade through sterilization is cut into is inoculated on callus inducing medium, cellar culture.Described callus inducing medium is with MS culture medium for minimal medium, and contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 1.0mg/L, and Medium's PH Value is 6.0.The condition of culture of described cellar culture is pH value 6.0, cultivation temperature 25 DEG C, periodicity of illumination 12 hours/day, and intensity of illumination is 2000lx.Cultivate 30 days, define callus and a large amount of adventitious bud, wherein some adventitious bud reaches 2-3 centimetre.
2) culture of rootage: selecting step 1) in bud reach the adventitious bud of 2-3 centimetre, it cut from callus, all the other callus and adventitious bud continue induction of callus, and the adventitious bud cut is inoculated on root media, cellar culture, the same step 1) of condition of culture.Described root media is with MS culture medium for minimal medium, and contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L, and Medium's PH Value is 6.0.Cultivate 15 days, grown white root, obtain aseptic Asiatic plantain seedling.
Result shows, explant disinfection efficiency is up to 80%, and can obtain aseptic adventitious bud fast, rooting rate reaches 100%, a large amount of Asiatic plantain seedlings of the maternal plant merit that can be maintained.
Embodiment 2
The present invention is utilized to carry out Asiatic plantain Vitro Quick Reproduction.
Described callus inducing medium is with MS culture medium for minimal medium, and contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L, and Medium's PH Value is 6.0.Other conditions are with embodiment 1.
Result shows, explant disinfection efficiency is up to 80%, and can obtain aseptic adventitious bud fast, rooting rate reaches 100%, a large amount of Asiatic plantain seedlings of the maternal plant merit that can be maintained.
The above is only the preferred embodiment of the present invention; it should be pointed out that for those skilled in the art, under the prerequisite not departing from the technology of the present invention principle; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (7)
1. a method for Asiatic plantain Vitro Quick Reproduction, comprises the steps:
1) induction of callus: the Asiatic plantain blade through sterilization is cut vein and leaf margin, is cut into small pieces, be inoculated on callus inducing medium, cellar culture is to forming callus and adventitious bud; Described fritter is the fritter of 0.8mm × 0.8mm; Described callus inducing medium is with MS culture medium for minimal medium, and contained sucrose concentration is 10 ~ 50g/L, concentration of NAA is 0.05 ~ 0.5mg/L, 6-benzyl aminopurine concentration is the culture medium of 1.0 ~ 4.0mg/L;
2) culture of rootage: selecting step 1) in bud reach the adventitious bud of 2-3 centimetre, it is cut from callus, all the other callus and adventitious bud continue induction of callus, the adventitious bud cut is inoculated on root media, cellar culture, to the root growing white, obtains aseptic Asiatic plantain seedling; Described root media is with MS culture medium for minimal medium, and contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L.
2. the method for Asiatic plantain Vitro Quick Reproduction as claimed in claim 1, it is characterized in that, the concrete grammar of described sterilization is: pluck the Asiatic plantain blade without disease and pest, running water, superclean bench is used alcohol-pickled 30s, proceed to again in the mercuric chloride solution of 0.1% and soak 8min, with aseptic water washing 3-5 time.
3. the method for Asiatic plantain Vitro Quick Reproduction as claimed in claim 1, it is characterized in that, described callus inducing medium is for minimal medium with MS culture medium, contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 1.0mg/L, or with MS culture medium for minimal medium, contained sucrose concentration is 30g/L, concentration of NAA is 0.1mg/L, 6-benzyl aminopurine concentration is the culture medium of 2.0mg/L.
4. the method for Asiatic plantain Vitro Quick Reproduction as claimed in claim 1, it is characterized in that, the condition of culture of described cellar culture is pH value 6.0, cultivation temperature 25 DEG C, periodicity of illumination 12 hours/day, and intensity of illumination is 2000lx.
5. the method for Asiatic plantain Vitro Quick Reproduction as claimed in claim 1, it is characterized in that, the time of described induction of callus is 28-32 days.
6. the method for Asiatic plantain Vitro Quick Reproduction as claimed in claim 1, it is characterized in that, the time of described culture of rootage is 14-16 days.
7. the application of method in Asiatic plantain breeds of the Asiatic plantain Vitro Quick Reproduction described in any one of claim 1-6.
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| CN108260526B (en) * | 2017-12-05 | 2021-01-12 | 长沙学院 | Tissue culture method of early-falling marsdenia tenacissima |
| CN115868408B (en) * | 2022-10-18 | 2024-05-14 | 中国长江三峡集团有限公司 | Method for efficiently breeding endangered plant plantain |
| CN115474551B (en) * | 2022-10-18 | 2023-03-31 | 中国长江三峡集团有限公司 | Tissue culture and rapid propagation method of national second-level protection plant of Tokyo plantago |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1685811A (en) * | 2005-06-03 | 2005-10-26 | 中国科学院植物研究所 | In vitro breeding method of plantage |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1685811A (en) * | 2005-06-03 | 2005-10-26 | 中国科学院植物研究所 | In vitro breeding method of plantage |
Non-Patent Citations (4)
| Title |
|---|
| MICROPROPAGATION OF PLANTAGO CAMTSCHATICA LINK;EMILIA ANDRZEJEWSKA-GOLEC ETAL;《ACTA SOCIETATIS BOTANICORUM POLONIAE》;20081231;第77卷(第4期);第269-273页 * |
| 大车前体外再生体系的建立和优化;李平等;《生物工程学报》;20051130;第21卷(第6期);第916-921页 * |
| 车前不定芽直接诱导及再生体系的建立;曾建军等;《安徽农业科学》;20081231;第36卷(第6期);第1810- 1811页 * |
| 车前的离体培养和植株再生;涂艺声;《江西科学》;19950930;第13卷(第3期);第149页摘要、第150页第1-2段、第151页第2-3段 * |
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