CN109673514A - A kind of method of south yampi type Chinese yam tissue-culturing quick-propagation - Google Patents
A kind of method of south yampi type Chinese yam tissue-culturing quick-propagation Download PDFInfo
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Classifications
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- 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
- A01H4/008—Methods for regeneration to complete plants
-
- 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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- Developmental Biology & Embryology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a kind of methods of southern yampi type Chinese yam tissue-culturing quick-propagation, comprising the following steps: (1) explant is handled: clip Chinese yam plant shoot carries out disinfection shoot；(2) shoot after disinfection bud inducement cultivation primary: is cut into stem section and stem apex as explant, stem section cuts blade, retain a leaf segment, then stem section and stem apex are inoculated in bud inducement cultivation base and cultivated 40-60 days, induction differentiates Chinese yam aseptic seedling；(3) Multiplying culture: taking Chinese yam aseptic seedling, cuts the band base of leaf dissection of Chinese yam aseptic seedling single-unit, and morphology lower end is inoculated in proliferated culture medium, cultivates 40-60 days, Multiplying culture goes out a large amount of Chinese yam aseptic seedlings；(4) Rooting and hardening-off culture；(5) practice seedling and transplanting.Method of the invention has the advantages that breeding coefficient is high, reproductive efficiency is high and seedling genetic stability is good and is not subject to seasonal restrictions.Chinese yam tissue-cultured seedling transplanting survival rate is high, is 95% or more.
Description
Technical field
The present invention relates to Chinese yam field of planting, in particular to a kind of side of southern yampi type Chinese yam tissue-culturing quick-propagation
Method.
Background technique
The monocotyledon of Chinese yam category Dioscoreaceae (Dioscoreaceae) Dioscorea (Dioscorea L.), medicine food is simultaneous excellent,
Medicinal part is stem tuber.The regional more appellations of Chinese yam are Chinese yam in south China, and ginseng potato (D.alata) is that these areas are generally cultivated
Chinese yam type, they are suitable for growing under Perenniporia martius weather conditions.Climate in Guangxi is superior, and Chinese yam cultivated area is big,
It is the big producing region of South China's Chinese yam first.The yam variety of osmanthus Huaihe River series such as osmanthus Huaihe River 5, Guihuai No.6, Gui Huai 7, Gui Huai 8
It is in long cylinder yampi type Chinese yam type etc. a kind of mutation for being ginseng potato, between long 70~120cm, is rich in starch, crude protein, ammonia
Base acid, mucopolysaccharide, saponin, microelement such as zinc, iron etc., in recent years, these kinds are in Guangxi by commerial growing.In production,
Chinese yam mainly uses stem tuber to breed, and there are sowing quantities greatly, production cost is higher, kind sexual involution (especially disease resistance and resistance
Decline is serious) a series of problems, such as, these problems are not to seldom knot bulbil or for tying the southern yampi type kind of bulbil
It is more prominent.On the other hand, as the improvement of people's living standards, Chinese yam is edible and medical value is increasingly taken seriously,
Demand constantly increases, and the phenomenon that supply falls short of demand seriously occurs.Therefore, in order to meet the market demand, yam cultivation area day
Benefit expands.
Vitro Quick Reproduction based on tissue culture technique is a kind of most widely used and most effective seedling-raising technique,
It has the advantages that breeding coefficient is high, reproduction speed is fast, is not subject to seasonal restrictions.The research of Chinese yam tissue cultures has report both at home and abroad
Road, but mostly using evoked callus such as blade, stem section, bulbils as research contents, induction tissue-cultured seedling time length, low efficiency, and
It is unfavorable for keeping kind stabilization characteristics of genetics；In addition, also there is a few studies report to directly obtain group using inductions such as stem section, stem apexs
Seedling is trained, but the quality of tissue-cultured seedling and induced efficiency etc. are to be improved.In addition, plant difference its method for tissue culture is not yet
Together, even same plant, because its different tissue culture method of kind is also not quite similar.In other words, same endophytic every
Nutrition and environmental condition required for one kind has its respective growth-development law and suitable growth to develop.Therefore,
Suitable southern yampi type Chinese yam tissue culture and rapid proliferation, optimal medium and external environmental condition, with other plant or
Kind is also different.
Summary of the invention
The purpose of the present invention is to provide a kind of methods of southern yampi type Chinese yam tissue-culturing quick-propagation, to overcome
Chinese yam breeds the disadvantage that sowing quantity is big, induction time is long, planting percent is low, keeps kind heredity difference.
To achieve the above object, the present invention provides a kind of method of southern yampi type Chinese yam tissue-culturing quick-propagation,
The following steps are included:
(1) explant is handled: clip Chinese yam plant shoot carries out disinfection shoot；
(2) bud inducement cultivation primary: the shoot after disinfection is cut into stem section and stem apex as explant, stem section cuts leaf
Piece retains a leaf segment, and then stem section and stem apex are inoculated in bud inducement cultivation base and cultivated 40-60 days, condition of culture are as follows: temperature
20-30 DEG C, light intensity 1500-2000lux are spent, periodicity of illumination is illumination 8-10hd-1, dark 14-16hd-1, induction differentiates
Chinese yam aseptic seedling；
(3) Multiplying culture: taking Chinese yam aseptic seedling, cuts the band base of leaf dissection of Chinese yam aseptic seedling single-unit, the inoculation of morphology lower end
In proliferated culture medium, cultivate 40-60 days, condition of culture are as follows: 20-30 DEG C of temperature, light intensity 1500-2000lux, periodicity of illumination are
Illumination 8-10hd-1, dark 14-16hd-1, Multiplying culture goes out Chinese yam aseptic seedling；
(4) Rooting and hardening-off culture: taking the aseptic seedling of Multiplying culture to be inoculated in Rooting and hardening-off culture base, cultivates 25-30 days,
Condition of culture are as follows: 20-30 DEG C of temperature, light intensity 1500-2000lux, periodicity of illumination are illumination 8-10hd-1, dark 14-16h
d-1, obtain tissue-cultured seedling of taking root；
(5) practice seedling and transplanting: the tissue-cultured seedling that will take root carries out being transplanted to crop field after practicing seedling.
Preferably, in above-mentioned technical proposal, step (1) disinfection includes: the fracture end for cutting away shoot, is rushed with water
It washes, is placed in after impregnating 30-45s in alcoholic solution, take out shoot and be placed in 0.1% mercuric chloride added with 1-3 drop Tween-20
(HgCl2) soaking disinfection 6-8min, after removing mercuric chloride solution, with aseptic water washing 4-6 times, water is blotted on sterile blotting paper
Point.
Preferably, in above-mentioned technical proposal, clip Chinese yam plant 8-10cm shoot in step (1), then cut away shoot
Fracture end, stay the tender tip of 6-8cm.
Preferably, in above-mentioned technical proposal, the raw material of bud inducement cultivation base includes: MS culture medium, excitement in step (2)
Element, methyl α-naphthyl acetate, active carbon, sucrose and agar；Wherein, the concentration of kinetin is 2.0-3.0mg/L, and the concentration of methyl α-naphthyl acetate is 0.2-
0.3mg/L, the concentration of active carbon are 1.5-2.0g/L, and the concentration of sucrose is 20-35g/L, and the concentration of agar is 8-9g/L,
pH5.8-5.9。
Preferably, in above-mentioned technical proposal, each culture bottle is inoculated with 5-6 explant in step (2).
Preferably, in above-mentioned technical proposal, in step (3) raw material of proliferated culture medium include: MS culture medium, kinetin,
Methyl α-naphthyl acetate, active carbon, sucrose and agar；Wherein, the concentration of kinetin is 1.0-2.0mg/L, and the concentration of methyl α-naphthyl acetate is 0.1-
0.2mg/L, the concentration of active carbon are 0.3-0.5g/L, and the concentration of sucrose is 30-35g/L, and the concentration of agar is 8-9g/L,
pH5.8-5.9。
Preferably, in above-mentioned technical proposal, in step (4) raw material of Rooting and hardening-off culture base include: 1/2MS culture medium,
Methyl α-naphthyl acetate, active carbon, sucrose and agar；Wherein, the concentration of methyl α-naphthyl acetate is 0.1-0.2mg/L, and the concentration of active carbon is 0.1-
0.3g/L, the concentration of sucrose are 30-35g/L, and the concentration of agar is 8-9g/L, pH5.8-5.9.
Preferably, in above-mentioned technical proposal, step (5) practices seedling and is, the tissue-cultured seedling that will first take root is placed in outdoor sunshade and practices seedling 2-3
It, after open culture bottle cap, sunshade is practiced seedling 4-5 days, takes out seedling, clean root culture medium, being placed on mass concentration is
0.15%KMnO41~2min is impregnated in solution, is then transplanted in sterilized matrix and is cultivated, the small arch of covered plastic film
Canopy, gradually taking off film and leaking informaton reduces humidity, takes off canopy, regular fertilizer and water management after 15-20d completely, and hardening is transplanted to big after 1.5-2 months
Field.
Preferably, in above-mentioned technical proposal, the matrix includes detritus soil and perlite, the body of detritus soil and perlite
Product is than being 3-4: 1.
Compared with prior art, the invention has the following beneficial effects:
(1) method of the southern yampi type Chinese yam tissue-culturing quick-propagation of the present invention, has breeding coefficient height, reproductive efficiency
High and seedling genetic stability is good and the advantages of not being subject to seasonal restrictions.It can solve a series of the asking of existing Chinese yam potato wedge breeding
The topic and callus induction tissue-cultured seedling time is long, seedling low efficiency and is unfavorable for the problem of keeping kind stabilization characteristics of genetics.
And its pollution rate is low, no microbial contamination is the foundation stone that tissue culturing system is successfully established, and experiment results proved: uses the application
In explant processing and sterilization method, pollution rate is down to 7.14%.
(2) Fiber differentiation primary is high-efficient, and explant is easy to get, and Initial culture directly induces budding, without more
Injured tissue cultivation stage, conducive to the stabilization for keeping kind inhereditary feature.
(3) proliferation rate is high, and the culture medium prescription and cultural method that this method provides can make tissue-cultured seedling fast breeding, cultivates
After 50d, induces the proper healthy and strong aseptic seedling quantity of form and reach as high as 6.5 times, 4.5 times of average out to.
(4) it takes root quickly, rooting rate is high, and radical amount is more, it is especially advantageous for taking root using prescription of rooting medium of the invention,
After cultivating one week, root long 1cm or so, rooting rate 100%, and radical is more, every plant of 4-10 item root, be conducive to transplanting at
It is living.Transplanting survival rate is high.Using method of the invention, Chinese yam tissue-cultured seedling transplanting survival rate is high, is 95% or more.
Detailed description of the invention
Fig. 1 is the Chinese yam tender stem explant according to acquisition.
Fig. 2 be according to the method for the present invention in the indefinite bud point that induces of bud inducement cultivation base primary training culture.
Fig. 3 is the aseptic seedling that Initial culture induces according to the method for the present invention.
Fig. 4 is the aseptic seedling that middle Multiplying culture obtains according to the method for the present invention.
Fig. 5 is the tissue-cultured seedling of middle culture of rootage according to the method for the present invention.
Fig. 6 is the seedling that middle tissue-cultured seedling is transplanted to crop field according to the method for the present invention.
Specific embodiment
With reference to the accompanying drawing, specific embodiments of the present invention will be described in detail, it is to be understood that guarantor of the invention
Shield range is not limited by the specific implementation.
Unless otherwise explicitly stated, otherwise in entire disclosure and claims, term " includes " or its change
Changing such as "comprising" or " including " etc. will be understood to comprise stated element or component, and not exclude other members
Part or other component parts.
Embodiment 1
A kind of method of south yampi type Chinese yam tissue-culturing quick-propagation, comprising the following steps:
1. explant processing and disinfection
June, 8~10cm shoot of clip Chinese yam robust plant rinses surface dirt under water tap, cuts away
One cutout end, it is tender slightly to leave about 6~8cm, is placed in a beaker, and 30min or more is rinsed under flowing water；Superclean bench is brought into,
With 75% alcohol impregnate 30~45s after, then with added with 2~3 drop Tween-20s 0.1%HgCl26~8min of soaking disinfection, every
1min rolling makes a movement, and is allowed to come into full contact with thimerosal.After removing mercuric chloride solution, with aseptic water washing 4~5 times, in sterile suction
Suck dry moisture on water paper.Shoot is as shown in Figure 1.
2. bud inducement cultivation primary
Take in step 1 it is tender slightly cut stem segments and stem apex as explant, stem segments cut blade, are then cut into length
About 1cm retains the dissection of a leaf segment, and stem apex can directly be inoculated with, be inoculated in bud inducement cultivation base, 5 explants of every bottle of inoculation
Body.Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, the photoperiod is illumination 10hd-1/ dark 14hd-1,
Culture 50d days, induces Chinese yam aseptic seedling.The indefinite bud point that bud inducement cultivation base training culture primary induces is as described in Figure 2, nothing
Vaccine is as shown in Figure 3.
Wherein, the bud inducement cultivation base be MS culture medium+kinetin (KT)+methyl α-naphthyl acetate (NAA)+active carbon (AC)+
Sucrose+agar, the concentration of the KT are 3.0mg/L, and the concentration of the NAA is 0.2mg/L, and the concentration of the AC is 2.0g/L,
The concentration of the sucrose is 30g/L, and the concentration of the agar is 8g/L, pH5.8.
3. Multiplying culture
The Chinese yam aseptic seedling for taking step 2 culture to obtain cuts the Chinese yam aseptic seedling list of robust growth in superclean bench
Section band base of leaf dissection, morphology lower end are inoculated in proliferated culture medium, and every bottle of 5~6 stem sections of inoculation or stem apex mark after inoculation
Date；Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, photoperiod illumination 10hd-1/ dark 14hd-1,
After cultivating 50d, a large amount of aseptic seedlings are obtained.The aseptic seedling that Multiplying culture obtains is as shown in Figure 4.
Wherein, the proliferated culture medium is MS culture medium+KT+NAA+AC+ sucrose+agar, and the concentration of the KT is
The concentration of 1.0mg/L, the NAA are 0.1mg/L, and the concentration of the AC is 0.5g/L, and the concentration of the sucrose is 30g/L, institute
The concentration for stating agar is 8g/L, pH5.8.
4. Rooting and hardening-off culture
The aseptic seedling for taking step 3 Multiplying culture to go out, is inoculated in Rooting and hardening-off culture base, and every bottle is inoculated with 5 plants, marks after inoculation
Infuse the date；Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, photoperiod illumination 10hd-1/ dark 14hd-1, there is root long to go out within culture 1 week, cultivate 28d, carry out acclimatization and transplants.The tissue-cultured seedling of Rooting and hardening-off culture is as shown in Figure 5.
Wherein, the Rooting and hardening-off culture base is 1/2MS culture medium+NAA+AC+ sucrose+agar, and the concentration of the NAA is
The concentration of 0.1mg/L, the AC are 0.1g/L, and the concentration of the sucrose is 30g/L, and the concentration of the agar is 8g/L,
pH5.9。
5. hardening and transplanting
The tissue-cultured seedling of taking root of step 4 is taken out from culturing room, gradually adjusting temperature and humidity makes tissue-cultured seedling reform of nature environment,
First set outdoor sunshade hardening 2d, after open bottle cap sunshade and practice seedling 5d, finally take out seedling, carefully clean root culture medium, is placed on
0.15%KMnO41~2min is impregnated in solution, is then transplanted to sterilized detritus soil: perlite=3: in 1 matrix,
Covered plastic film Small plastic shed, gradually taking off film and leaking informaton reduces humidity, takes off canopy, regular fertilizer and water management, after hardening 55 days after 18d completely
Crop field can be transplanted to.The tissue-cultured seedling for being transplanted to crop field is as shown in Figure 6.
Embodiment 2
A kind of method of south yampi type Chinese yam tissue-culturing quick-propagation, comprising the following steps:
1. explant processing and disinfection
July, 8~10cm shoot of clip Chinese yam robust plant rinses surface dirt under water tap, cuts away
One cutout end, it is tender slightly to leave about 6~8cm, is placed in a beaker, and 30min or more is rinsed under flowing water；Superclean bench is brought into,
With 75% alcohol impregnate 30~45s after, then with added with 2~3 drop Tween-20s 0.1%HgCl26~8min of soaking disinfection, every
1min rolling makes a movement, and is allowed to come into full contact with thimerosal.After removing mercuric chloride solution, with aseptic water washing 4~5 times, in sterile suction
Suck dry moisture on water paper.
2. bud inducement cultivation primary
Take in step 1 it is tender slightly cut stem segments and stem apex as explant, stem segments cut blade, are then cut into length
About 1cm retains the dissection of a leaf segment, and stem apex can directly be inoculated with, be inoculated in bud inducement cultivation base, 5 explants of every bottle of inoculation
Body.Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, the photoperiod is illumination 10hd-1/ dark 14hd-1,
Culture 50d days, induces Chinese yam aseptic seedling.
Wherein, the bud inducement cultivation base be MS culture medium+kinetin (KT)+methyl α-naphthyl acetate (NAA)+active carbon (AC)+
Sucrose+agar, the concentration of the KT are 2.0mg/L, and the concentration of the NAA is 0.3mg/L, and the concentration of the AC is 1.5g/L,
The concentration of the sucrose is 30g/L, and the concentration of the agar is 8g/L, pH5.9.
3. Multiplying culture
The Chinese yam aseptic seedling for taking step 2 culture to obtain cuts the Chinese yam aseptic seedling list of robust growth in superclean bench
Section band base of leaf dissection, morphology lower end are inoculated in proliferated culture medium, and every bottle of 5~6 stem sections of inoculation or stem apex mark after inoculation
Date；Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, photoperiod illumination 10hd-1/ dark 14hd-1,
After cultivating 50d, a large amount of aseptic seedlings are obtained.
Wherein, the proliferated culture medium is MS culture medium+KT+NAA+AC+ sucrose+agar, and the concentration of the KT is
The concentration of 2.0mg/L, the NAA are 0.2mg/L, and the concentration of the AC is 0.5g/L, and the concentration of the sucrose is 30g/L, institute
The concentration for stating agar is 8g/L, pH5.9.
4. Rooting and hardening-off culture
The aseptic seedling for taking step 3 Multiplying culture to go out, is inoculated in Rooting and hardening-off culture base, and every bottle is inoculated with 5 plants, marks after inoculation
Infuse the date；Condition of culture are as follows: 25 ± 1 DEG C of temperature, 1500~2000lux of light intensity, photoperiod illumination 10hd-1/ dark 14hd-1, there is root long to go out within culture 1 week, cultivate 30d, carry out acclimatization and transplants.
Wherein, the Rooting and hardening-off culture base is 1/2MS culture medium+NAA+AC+ sucrose+agar, and the concentration of the NAA is
The concentration of 0.1mg/L, the AC are 0.1g/L, and the concentration of the sucrose is 30g/L, and the concentration of the agar is 8g/L,
pH5.8。
5. hardening and transplanting
The tissue-cultured seedling of taking root of step 4 is taken out from culturing room, gradually adjusting temperature and humidity makes tissue-cultured seedling reform of nature environment,
First set outdoor sunshade hardening 3d, after open bottle cap sunshade and practice seedling 4d, finally take out seedling, carefully clean root culture medium, is placed on
0.15%KMnO41~2min is impregnated in solution, is then transplanted to sterilized detritus soil: perlite=4: in 1 matrix,
Covered plastic film Small plastic shed, gradually taking off film and leaking informaton reduces humidity, takes off canopy, regular fertilizer and water management, after hardening 45 days after 20d completely
Crop field can be transplanted to.
Southern yampi type Chinese yam tissue cultures are quickly bred using the above method, and detect all data index,
As shown in table 1, table 2, table 3 and table 4.Table 1 is the statistics of explant pollution rate, and table 2 is explant induction differentiation budding situation system
Meter, table 3 are Multiplying culture situation statistics, and table 4 is transplanting survival rate statistics.
1 explant pollution rate of table statistics
As shown in table 1, using the explant processing of the method for the present invention and sterilization method, pollution rate is down to 7.14%, significantly
Reduce pollution rate.
2 explant of table induction differentiation budding situation statistics
As shown in table 2, high using the method for the present invention bud induction differentiation rate primary, explant is easy to get, and Initial culture
Directly induction budding, without the callus tissue culture stage, conducive to the stabilization for keeping kind inhereditary feature.
3 Multiplying culture situation of table statistics
As shown in table 3, the culture medium prescription and cultural method provided using the method for the present invention can make tissue-cultured seedling fast breeding,
After cultivating 50d, induces the proper healthy and strong aseptic seedling quantity of form and reach as high as 6.5 times, 4.5 times of average out to, i.e. proliferation rate is high.
4 transplanting survival rate of table statistics
| It transplants tissue-cultured seedling number (strain) | Survival seedling number (strain) | Survival rate (%) |
| 150 | 143 | 95.33 |
As shown in table 4, high using the method for the present invention Chinese yam tissue-cultured seedling transplanting survival rate, it is 95% or more.
The aforementioned description to specific exemplary embodiment of the invention is in order to illustrate and illustration purpose.These descriptions
It is not wishing to limit the invention to disclosed precise forms, and it will be apparent that according to the above instruction, can much be changed
And variation.The purpose of selecting and describing the exemplary embodiment is that explaining specific principle of the invention and its actually answering
With so that those skilled in the art can be realized and utilize a variety of different exemplary implementation schemes of the invention and
Various chooses and changes.The scope of the present invention is intended to be limited by claims and its equivalents.
Claims (9)
1. a kind of method of south yampi type Chinese yam tissue-culturing quick-propagation, which comprises the following steps:
(1) explant is handled: clip Chinese yam plant shoot carries out disinfection shoot；
(2) bud inducement cultivation primary: cutting stem section and stem apex as explant for the shoot after disinfection, and stem section cuts blade,
Retain a leaf segment, then stem section and stem apex are inoculated in bud inducement cultivation base and cultivated 40-60 days, condition of culture are as follows: temperature
20-30 DEG C, light intensity 1500-2000lux, periodicity of illumination are illumination 8-10hd-1, dark 14-16hd-1, induce and differentiate Huaihe River
Mountain aseptic seedling；
(3) Multiplying culture: taking Chinese yam aseptic seedling, cuts the band base of leaf dissection of Chinese yam aseptic seedling single-unit, morphology lower end is inoculated in increasing
It grows in culture medium, cultivates 40-60 days, condition of culture are as follows: 20-30 DEG C of temperature, light intensity 1500-2000lux, periodicity of illumination are illumination
8-10h·d-1, dark 14-16hd-1, Multiplying culture goes out a large amount of Chinese yam aseptic seedlings；
(4) Rooting and hardening-off culture: taking the aseptic seedling of Multiplying culture to be inoculated in Rooting and hardening-off culture base, cultivates 25-30 days, culture
Condition are as follows: 20-30 DEG C of temperature, light intensity 1500-2000lux, periodicity of illumination are illumination 8-10hd-1, dark 14-16hd-1,
Obtain tissue-cultured seedling of taking root；
(5) practice seedling and transplanting: the tissue-cultured seedling that will take root carries out being transplanted to crop field after practicing seedling.
2. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(1) disinfection includes: the fracture end for cutting away shoot, is rinsed with water, is placed in after impregnating 30-45s in alcoholic solution, is taken out tender
Branch is placed in the 0.1% mercuric chloride soaking disinfection 6-8min added with 2-3 drop Tween-20, after removing mercuric chloride solution, uses aseptic water washing
4-6 times, the suck dry moisture on sterile blotting paper.
3. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 2, which is characterized in that step
(1) clip Chinese yam plant 8-10cm shoot in, then the fracture end of shoot is cut away, stay the tender tip of 6-8cm.
4. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(2) raw material of bud inducement cultivation base includes: MS culture medium, kinetin, methyl α-naphthyl acetate, active carbon, sucrose and agar in；
Wherein, the concentration of kinetin is 2.0-3.0mg/L, and the concentration of methyl α-naphthyl acetate is 0.2-0.3mg/L, and the concentration of active carbon is
1.5-2.0g/L, the concentration of sucrose are 30-35g/L, and the concentration of agar is 8-9g/L, pH5.8-5.9.
5. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(2) each culture bottle is inoculated with 5-6 explant in.
6. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(3) raw material of proliferated culture medium includes: MS culture medium, kinetin, methyl α-naphthyl acetate, active carbon, sucrose and agar in；
Wherein, the concentration of kinetin is 1.0-2.0mg/L, and the concentration of methyl α-naphthyl acetate is 0.1-0.2mg/L, and the concentration of active carbon is
0.3-0.5g/L, the concentration of sucrose are 30-35g/L, and the concentration of agar is 8-9g/L, pH5.8-5.9.
7. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(4) raw material of Rooting and hardening-off culture base includes: 1/2MS culture medium, methyl α-naphthyl acetate, active carbon, sucrose and agar in；
Wherein, the concentration of methyl α-naphthyl acetate is 0.1-0.2mg/L, and the concentration of active carbon is 0.1-0.3g/L, and the concentration of sucrose is 30-
35g/L, the concentration of agar are 8-9g/L, pH5.8-5.9.
8. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 1, which is characterized in that step
(5) practicing seedling is, the tissue-cultured seedling that will first take root, which is placed in outdoor sunshade, to be practiced seedling 2-3 days, after open culture bottle cap, sunshade experienced seedling 4-5 days,
Seedling is taken out, root culture medium is cleaned, being placed on mass concentration is 0.15%KMnO41~2min is impregnated in solution, is then transplanted to
In sterilized matrix, covered plastic film Small plastic shed, gradually taking off film and leaking informaton reduces humidity, takes off canopy completely after 15-20d, fixed
Phase fertilizer and water management, hardening were transplanted to crop field after 1.5-2 months.
9. the method for south yampi type Chinese yam tissue-culturing quick-propagation according to claim 8, which is characterized in that described
Matrix include detritus soil and perlite, the volume ratio of detritus soil and perlite is 3-4: 1.
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