CN101268758A - Quick replication method for Xuan pawpaw tissue cultivation - Google Patents
Quick replication method for Xuan pawpaw tissue cultivation Download PDFInfo
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- CN101268758A CN101268758A CNA2008100988843A CN200810098884A CN101268758A CN 101268758 A CN101268758 A CN 101268758A CN A2008100988843 A CNA2008100988843 A CN A2008100988843A CN 200810098884 A CN200810098884 A CN 200810098884A CN 101268758 A CN101268758 A CN 101268758A
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Abstract
The invention relates to a rapid breeding method which uses MS as the basic culture medium and respectively performs the tissue culture of a xuancheng pawpaw through inducing a sprout and a root. The rapid breeding method is characterized in that the cotyledon or the embryo of the seed of the xuancheng pawpaw is adopted to perform the tissue culture. When the cotyledon of the seed is adopted to perform the tissue culture, the best culture medium formulation for inducing an adventitious bud is MS plus NAA0.5mg/L plus TDZ1.0mg/L, the best culture medium formulation for inducing rooting is 1/2MS plus IBA0.2mg/L; when the embryo of the seed is adopted to perform the tissue culture, the best culture medium formulation for inducing an adventitious bud is MS plus BA0.5mg/L plus NAA0.2mg/L, and the best culture medium formulation for inducing rooting is 1/2MS plus NAA0.2mg/L.
Description
One, technical field
The invention belongs to the plant regeneration technical field of passing through tissue culture in the forestry, be specifically related to a kind of method of declaring the pawpaw tissue-culturing quick-propagation.
Two, background technology
A surname pawpaw (Chaenomeles speciosa Nakai.) belongs to the rose family, is fallen leaves or half evergreen shrubs or dungarunga, and the close thorniness of branch can be made green Li.Single leaf alternate has short handle and stipule.Hua Dansheng or cluster, sepal 5, petal 5, fruit are the operatic circle, plant leather matter, no endosperm.A surname's pawpaw also common cultivation in various places, early spring is the flower posterior lobe earlier, and pattern is bright red, pink, milky white and polyphyll and half double variety arranged.
A surname pawpaw is medicinal certified products, is one of China's four big traditional Chinese medicines, apart from modern existing more than 2,000 year cultivation history.A surname pawpaw is produced with the Xuancheng, Anhui declares pawpaw the most famous, is written into " Mingyi Bielu ", classifies middle product as.A surname pawpaw is recorded in fruit portion by Compendium of Material Medica, draws Su Songyu: " pawpaw has it everywhere, and the Xuancheng person is good ".A surname pawpaw contains 19 seed amino acids, 18 kinds of mine trace elements, and a large amount of vitamin Cs, also contain saponin, flavones, malic acid, oleanolic acid, citric acid, citric acid, tartaric acid, ascorbic acid, fumaric acid, tannin etc. simultaneously, contain catalase, phenol oxidase, oxidase, be rich in hepatocuprein (SOD) etc. especially.Be used as medicine after the dried fruit system, wind dispelling, Shujin, active, analgesia, detumescence, pleasant effect are arranged.Can eat after the processing of a surname's pawpaw fruit, can be made into Nakai beverage, papaya ester, papaya wine etc.Owing to be grown in the different environment, all there is certain difference in a surname pawpaw of Different Provenances aspect proterties and the medicament contg.There are some researches show that the content of a surname's pawpaw organic acid and alcohols is respectively 2 times and 3 times of common wrinkled papaya.Therefore strengthen protection, can be the medicinal pawpaw new varieties of following cultivation abundant genetic resources is provided the Different Provenances wrinkled papaya.
The modes of reproduction of tradition a surname pawpaw seed and the cottage propagations that adopt more, reproductive efficiency is lower, and plant tissue can rapidly and efficiently be finished the reproductive process of plant.Plant Tissue Breeding was 20 beginnings of the century, was the emerging technology that base growth is got up with plant physiology.On the cell theory basis of Schleiden and Schwann foundation, 1902, German botanist Haberlandt prophesy " plant cell has totipotency, and each cell can become a complete plant through culture in vitro all as embryonic cell ".Nineteen thirty-seven, people such as French botanist Gautheret use the little block organization of stem section cambium and the carrot root of tobacco for the first time, make cell proliferation and evoked callus, but do not induce organ.1958, people such as Steward successfully induced in the carrot cell suspension culture and have formed embryoid, and by embryoid growth formation whole plant.
Enter after the sixties in 20th century, Plant Tissue Breeding work most countries all over the world, and obtained the development of advancing by leaps and bounds, tissue and organ culture technology reach its maturity and are perfect, at present, plant tissue culture technique has become one of important research technology in the biological subject and means.The foundation of Plant Tissue Breeding system, for breeding of the rare famous and precious kind of plant and detoxification, germ plasm resource preserve, somatic mutants bring out and the aspects such as production, cell engineering and gene engineering of screening, artificial seed manufacturing, cell secondary metabolites all have great importance.Plant tissue culture technique is research and application further, will be to traditional plant production mode producing far-reaching influence.
At present, plant tissue culture technique mainly contains following four kinds of methods:
1. the breeding fast of exsomatizing of plant
Plant is exsomatized and breeds fast is that present Plant Tissue Breeding is used at most, the most effective a kind of method.The genetic stockss such as useful mutant of the breeding of newly breeding or newly introducing, fine individual plant, nature and the induced mutations of selection.By the rapid propagation in vitro of tissue, can breed in a large number at short notice, can extensive use on producing in one or two years.As the bud mutation material under the natural conditions, the good genetic stocks that the induced mutations breeding obtains etc. can be bred in a large number by tissue culture, thereby are applied to produce.
In the plant rapid propagation in vitro, the stem apex detoxify technology plays a very important role to the quality that improves some economic crops.In case the plant infective virus gently then causes tree type decline, output falls sharply, and product qualitative change is bad, and is serious then will produce destructive injury to the plant of infection.Based on the floristics of nourishing and generating such as potato, sweet potato, garlic, lily etc., long-term vegetative propagation accumulates virus in vivo, causes virus disease, has a strong impact on the ornamental value of yield and quality and the flower and the leaf etc. of fruit.But disease plant is not that each position all has virus.Utilize plant stem apex detoxification technology, can obtain detoxic seedling, thereby improve the quality and yield of plant greatly.As far back as nineteen forty-three, White just finds that near the virus concentration the growing tips of the plant is very low even virus-free.It is very fast at stem apex position plant growth rate that stem apex does not have the infective virus main cause, and the transfer velocity of virus is relatively slow.Therefore, the stem apex of intercepting plant health just can obtain detoxic seedling as explant through cultivating, and promptly can obtain a large amount of detoxic seedlings through quick breeding again.Nineteen sixty, first obtains the detoxification orchid with the Shoot Tip Culture method Morel, after this succeeds in various plants.
2. the breeding of monoploid, polyploid
Haplobiont is to induce the plant of generation by having single doubly (single cover) chromosomal pollen or egg cell.Haploid breeding has high speed, high efficiency, genotype advantage such as isozygoty.Therefore,, come out, and cultivated the crop new varieties that form establishing in large scale as a kind of brand-new breeding technique by the haploid breeding that flower pesticide or pollen are cultivated.Guha (1964) and Maheshiwari (1966) success go out haplobiont by pollen induction in the anther culture of datura.Up to now, existing plant more than 75 kinds obtains haplophyte in the world wide.
Polyploid is commonplace in distributed in nature, and it is bigger to show colored shape usually, bigger polyploid plant distinctive " huge " feature that waits of fruit.Therefore have higher ornamental value and economic worth.Usually obtain polyploid by following two kinds of approach: the one, double naturally; The 2nd, induced mutations, as add an amount of mutagen (colchicine etc.).If induced mutations and tissue culture technique are combined, the efficient of polyploid breeding will be improved greatly.Tissue culture technique and induced mutations combine, make day lily (Hemerocallis citrine Baroni.) double, select and fast breeding carry out synchronously, thereby shortened time of breeding.Utilize this technology, in the various plants material, successfully induced polyploid at present.
3. embryo culture and endosperm are cultivated
The stripped embryo culture of plant can solve the abortion problem of distant hybridization, and those can not form the hybrid combination with vitality seed can obtain hybrid plant by the cultivation of the embryo that exsomatizes.In breeding, Lai Bahe solves the problem that the hybrid seed of Iinum peerenne L (Linum peerenne L.) and Austrian flax can not be sprouted with embryo culture technique, makes their ripe and obtain hybrid plant.Pioneer's work of Lay Bach is laid a good foundation for the obstacle that the method with the culture of ex vivo embryo overcomes hybridity, cultivates the epoch that are applied to production practices thereby started plant embryos.Nineteen ninety, female Sillim etc. utilize embryo rescue technology successfully to obtain the species hybrid plant of delicious kiwi fruit * actinidia eriantha.After this, stripped embryo culture launches between various plants.
Endosperm is cultivated can obtain triploid, has opened up a new way for inducing the formation triploid plant.People such as Nakano differentiation from immature paddy endosperm is cultivated obtains 2 plant, proves that cereal class endosperm has the potential of differentiation plant equally.Utilize citrus endosperm cultured in vitro can obtain seedless citrus, and ratio 2 * faster with 4 * hybridization acquisition triploid.At present carried out cultured in vitro with the endosperm of 30 various plants.
4. artificial seed and stripped quality saving
Artificial seed refers to have the somatic cell parcel artificial endosperm of good growth and people's work post skin and a kind of reproductive structure of forming.Because artificial seed is not subjected to time restriction, can breed virus-free material in a large number in the short time, therefore for perennial plant with fertility is bad or the plant that is difficult to carry out sexual propagation, have remarkable advantages by the artificial seed breeding.Productive manpower seed at first needs a large amount of embryoids, and at present, the plant that can produce embryoid in a large number has 43 sections approximately, kind surplus in the of 100.The application prospect of artificial seed depends on production cost, and the high production cost of artificial seed is the biggest obstacle of its development.
Utilize traditional method to preserve germ plasm resource and need expend great amount of manpower and financial resources, and be difficult to long preservation.The process that ultralow temperature is preserved is with sterilizable material such as aseptic bud, somatic embryo etc., through after the freezing protective treatment, deposits in-196 ° of liquid nitrogen and preserves.When the needs germplasm materials, can induce organ differentiation and plant regeneration to obtain through thawing, cultivating again.Because ultralow temperature is preserved to have and is economized on resources and advantage such as germplasm materials, therefore opens up a new way for quality saving.In recent years, abroad the horticultural crop kind of successfully preserving with the vitrification ultra-low temperature preservation method had kind surplus cherry, sweet cherry, pears, the apple etc. tens, had domesticly also obtained certain progress in this field.
Up to now, producing regeneration plant by tissue culture organ occurring mode is the most successful forest cultured in vitro modes of reproduction, has obtained success in many plants.The tissue culture of plant has three kinds of regeneration approach, i.e. embryoid regeneration, callus regeneration and direct induction buds sprouting.Organ generation regeneration plant approach in the Plant Tissue Breeding needs separated bud induction period of experience and root induction phase.Organ has indirect and direct dual mode.Under indirect mode, indefinite bud induces from callus.Under direct mode, the bud of regeneration is to induce from the bud that has existed.When bud is when those positions that should not produce bud induce, this bud is called indefinite bud, and when bud be when regenerate in the position that can produce bud usually, this bud is called lateral bud and axillalry bud.
The formation of root is the organ occurring mode that the frequency of occurrences is the highest in the Plant Tissue Breeding.Two kinds of different root ex vivo differentiation forms are arranged, and a kind of is that the bud that has formed is taken root in cultivation, thereby forms regrowth, is called and takes root.Another kind of then be to be divided into the original hase of root and further to grow by the class meristematic tissue in the callus to form root, be called the root generation.By tissue culture elder generation evoking adventive bud, and then dissolving root in its base section, is a kind of common isolated organ occurring mode.In this case, the differentiation of bud and growth obviously can change the composition and the ratio of endogenous hormones in the tissue, thereby the formation to root exerts an influence, and this process that obtains regeneration plant through taking root by indefinite bud also is to utilize tissue culture to carry out one of important channel of clonal reproduction.Root generation phenomenon in the callus is often not as the general and easy acquisition like that of taking root, and the differentiation capability of callus root can weaken gradually along with the increase of subculture number so that forfeiture.
At present, the research data of a surname pawpaw tissue culture aspect is less.1993, Wu Guoliang etc. carried out cultured in vitro and fast numerous research to common flowering quince stem section, found that BA has good facilitation to the pawpaw indefinite bud, and add caseinhydrolysate (CH) breeding and the growth of indefinite bud were had negative interaction.2004, Fan Yilian etc. carried out the correlative study of common flowering quince stem section callus induction and regeneration.According to another report, utilize the tender stem section of common flowering quince to carry out the research of quick proliferation, show that the rate of sprouting of tender stem eye is 0%～22%, the rate of sprouting is relatively low; In addition, utilize the common flowering quince kind in addition---the research of aspect of regenerating of the spire of Nanjing brocade and stem section, all reach 100% though show callus induction rate, regeneration rate is lower, is generally 2%～10%.
Up to now, in all kinds of documents and materials of relevant plant tissue culture technique, except that Japan has the relevant patented technology report that belongs to close kind of Chaenomeles pawpaw (Chaenomeles lagenari) extract together, Shang Weijian organizes the complete set technology of training and delivers on a surname pawpaw.To breed based theoretical in a short time in a large number for a surname pawpaw kind to the research that a surname pawpaw breeds fast, thereby help to change achievement in research into productivity, and the production practices of a surname pawpaw be had important use be worth with merit.
Three, summary of the invention
The objective of the invention is to declare the research of pawpaw method for quickly breeding, seek and to be applicable to a surname pawpaw tissue culture from regenerating, thereby quicken the seed selection and the popularization of a surname pawpaw improved seeds to the packaged technology of fast breeding.
Technical solution of the present invention is: a kind ofly make minimal medium with MS, induce the method for declaring the pawpaw tissue-culturing quick-propagation with root induction by bud respectively, it is characterized in that adopting a surname's pawpaw seed cotyledon or a surname pawpaw seed embryo to carry out tissue culture.
When adopting a surname pawpaw seed cotyledon to carry out tissue culture, the optimal medium prescription of evoking adventive bud is MS+NAA0.5mg/L+TDZ1.0mg/L; The optimal medium prescription of root induction is 1/2MS+IBA0.2mg/L.
When adopting a surname pawpaw seed embryo to carry out tissue culture, the optimal medium prescription of evoking adventive bud is MS+BA0.5mg/L+NAA0.2mg/L; The optimal medium prescription of root induction is 1/2MS+NAA0.2mg/L.
The standard recipe of MS medium (Murashing and Skoog medium) is as follows:
NH
4NO
3 1650mg/L
KNO
3 1900mg/L
CaCl
2·2H
2O 440mg/L
MgSO
4·7H
2O 370mg/L
KH
2PO
4 170mg/L
KI 0.83mg/L
H
3BO
3 6.2mg/L
MnSO
4·H
2O 16.9mg/L
ZnSO
4·7H
2O 8.6mg/L
Na
2MoO
4·2H
2O 0.25mg/L
CuSO
4·5H
2O 0.025mg/L
CoCl
2·6H
2O 0.025mg/L
FeSO
4·7H
2O 27.8mg/L
Na
2-EDTA 37.3mg/L
Inositol 100mg/L
Nicotinic acid 0.5mg/L
Thiamine hydrochloride 0.1mg/L
Puridoxine hydrochloride 0.5mg/L
Glycine 2mg/L
MS/2, i.e. the MS medium that reduces by half of macroelement content; MS/4, promptly macroelement content is 1/4th MS medium.
Four, description of drawings
Accompanying drawing 1 cotyledon is induced the indefinite bud of generation
Accompanying drawing 2 betides the shape indefinite bud of growing thickly of cotyledon base portion
Accompanying drawing 3 the inducing of embryo children stem indefinite bud of sprouting
The propagation of accompanying drawing 4 indefinite buds
Accompanying drawing 5 adventitious bud inducings are taken root
Five, embodiment
Embodiment 1: adopt a surname pawpaw seed cotyledon to carry out tissue culture
A. the combination of medium, sterilization and condition of culture
MS is a minimal medium, adds TDZ and NAA, and the growth regulating plant of dissimilar combinations is added in the minimal medium with different concentration, and interpolation sucrose 30g/L, agar 6.5g/L, regulate medium pH 5.8～6.0, be sub-packed in the glass triangle flask of 100ml every bottle of about 40ml.The damp and hot sterilization of high pressure is adopted in the sterilization of medium and culture vessel.The medium that branch is installed places in the high-pressure sterilizing pot with experiment apparatus such as the culture vessel that bandages, tweezers, scalpel, and 121 ℃, 1.1kg/cm
3Sterilized 20 minutes.
B. the sterilization of explant
Divest exosper in a surname's pawpaw seed, with seed on superclean bench with 70% alcohol disinfecting 5～10 minutes, sterile distilled water washes 1 time.Use 0.1% or 0.15%HgCl then
2(adding a Tween-80) sterilization 5～15 minutes, cotyledon is taken out in sterile distilled water flushing 4～5 times at last from disinfection seed.
C. inoculation
To be inoculated into through the cotyledon of disinfecting in the following MS medium that contains different growth regulatory substances.Cultivate through 40 days that the back is observed and the record result, as following table:
The different growth regulatory substances of table 1 are to the influence of cotyledon evoking adventive bud
| PGR(mg/l) | Become the seedling inductivity |
| NAA0.1+TDZ1 NAA0.5+TDZ1 NAA1+TDZ0.1 NAA1+TDZ0.5 NAA1+TDZ1 NAA1+TDZ2 | 10％ 25％ 0 10％ 0 20％ |
Annotate: PGR---plant growth regulating substance, as follows.
Condition of culture: light application time is 8～16h/ days, intensity of illumination 1000～3000lx, 24 ± 3 ℃ of temperature;
Wherein, when adopting the medium of MS+NAA 0.5mg/L+TDZ 1.0mg/L, can obtain optimal result.
D. the propagation of indefinite bud
When treating that the indefinite bud height grows to the 3cm left and right sides, with its cutting-out, be inoculated among the proliferated culture medium MS+BA0.5mg/L+NAA0.2mg/L, indefinite bud can fast breeding.
E. take root and transplanting
1/2MS is a minimal medium, add different growth regulatory substance BA, IBA, NAA, be combined to form four types of IBA, NAA, BA+IBA, BA+NAA, add sucrose 30g/l, agar 6.5g/l again, regulate medium pH to 5.8～6.0, be sub-packed in the glass triangle flask of 100ml every bottle of about 40ml.
When the indefinite bud height in the proliferated culture medium reaches 2cm, cut indefinite bud and be inoculated in the above root media, observed result and record data behind the 40d, the result is as follows:
The influence of the different growth regulator confrontation of table 2 adventitious bud rooting
Condition of culture: light application time is 8～16h/ days, intensity of illumination 1000～3000lx, 24 ± 3 ℃ of temperature;
Wherein, when adopting the medium of 1/2MS+IBA 0.2mg/L, can obtain optimal result.
Treat that adventive root quantity reaches 3～4, when length reaches 4cm, transplant test-tube plantlet, promptly progressively open and seal indoor one week of hardening of film according to conventional method, subsequently at ambient temperature, with seedling replanting to containing peat soil: in the plastic basin of vermiculite=1: 1.The hardening initial stage seals with plastic film, removes film after the week, makes tissue cultivating seedling progressively adapt to extraneous variations in temperature, improves the survival rate of transplanting seedling.Transplanting survival rate can reach 80%.
A surname pawpaw cotyledon is inoculated in the medium of MS+NAA0.5mg/L+TDZ1.0mg/L, can obtain the inductivity up to 25%.The cotyledon evoking adventive bud, the occurrence positions of its indefinite bud is positioned at base portion, and only has indefinite bud to form at base portion, does not all see the generation that indefinite bud is arranged at otch and other position of cotyledon.Experimental result as can be known, TDZ is to the good effect of having induced of a surname's pawpaw cotyledon indefinite bud, and certain density TDZ is necessary to successful evoking adventive bud.Have than big-difference between the indefinite bud that TDZ induces, the indefinite bud that has is grown too vigorous and is presented vitrification phenomenon, and the indefinite bud poor growth, the stem section that have are shorter, and the indefinite bud that also has is the shape of growing thickly.Correlation between the TDZ concentration and the indefinite bud type of being induced is not obvious.When utilizing a surname pawpaw cotyledon as explant, BA does not show very high activity.2,4-D has inhibition in the process of inducing of indefinite bud, and a surname pawpaw cotyledon all can induce indefinite bud in a plurality of combinations of TDZ and NAA, and as TDZ and 2,4-D then can induce indefinite bud without any a kind of combination during combination.From the combination of the growth regulatory substance of adventitious bud inducing as can be seen, because in TDZ and 2, do not have indefinite bud to take place among the 4-D, think that therefore the existence of NAA also is essential to the generation of indefinite bud, but the concentration of NAA is then less relatively to the influence of adventitious bud inducing.
Embodiment 2: adopt a surname pawpaw seed embryo to carry out tissue culture
A. the combination of medium, sterilization and condition of culture
Divest exosper in a surname's pawpaw seed, with seed on superclean bench with 70% alcohol disinfecting 10 minutes, sterile distilled water washes 1 time, uses 0.15%HgCl
2(adding a Tween-80) sterilization 15 minutes, sterile distilled water flushing 5 times is inoculated into sterilization back seed in the germination medium of MS+BA 0.2mg/L.
B. adventitious bud proliferation
MS, 1/2MS or 1/4MS are minimal medium, different growth regulatory substances are combined to form seven types of TDZ+IBA, TDZ+NAA, BA+IBA, BA+NAA, BA+IBA+NAA, KT+IBA, KT+NAA, growth regulatory substance in the combinations thereof is added in the minimal medium with different concentration, and add sucrose 30g/L, agar 6.5g/L, regulate medium pH to 5.8-6.0.
When the seedling height of seedling in the germination medium reaches 2～3cm, cut aseptic seedling stem sections and be inoculated in the above-mentioned proliferated culture medium, observed result is as follows behind the cultivation 40d.
The influence of the different growth regulator confrontation of table 3 adventitious bud proliferation
Condition of culture: light application time is 8～16h/ days, intensity of illumination 1000～3000lx, 24 ± 3 ℃ of temperature.
Wherein, when adopting the medium of MS+BA 0.5mg/L+NAA 0.2mg/L, can obtain optimal result.
C. adventitious bud inducing is taken root
1/2MS is a minimal medium, add different growth regulatory substance BA, IBA, NAA, be combined to form four types of IBA, NAA, BA+IBA, BA+NAA, add sucrose 30g/l, agar 6.5g/l again, regulate medium pH to 5.8～6.0, be sub-packed in the glass triangle flask of 100ml every bottle of about 40ml.
When the indefinite bud height in the proliferated culture medium reaches 3cm, cut indefinite bud and be inoculated in the above root media, observed result and record data behind the 40d, the result is as follows:
The influence of the different growth regulator confrontation of table 4 adventitious bud rooting
Condition of culture: light application time is 8～16h/ days, intensity of illumination 1000～3000lx, 24 ± 3 ℃ of temperature;
Wherein, when adopting the medium of 1/2MS+NAA0.2mg/L, can obtain optimal result.
D. hardening and transplanting
When adventive root length surpasses 3～4 of the quantity of 4cm, adventive root, one week of uncork hardening, transplant test-tube plantlet according to conventional method then, transplanting survival rate can reach more than 85%.
Owing to adopted seed as initial experiment material, so whole experiment is not subjected to the influence of the plant growing rhythm and pace of moving things, can improve the efficient of declaring the pawpaw crossbreeding, acceleration good variety selection and popularization.Adopted the stem section behind the seed germination as experiment material, the explant survival rate of inoculation is 100%, and high proliferation coefficient is 4.48, is a breeding cycle according to experimentation 40d, and the highest can the propagation of indefinite bud forms more than 70 ten thousand indefinite buds in 1 year.Therefore, a surname pawpaw fast breeding has higher efficient with respect to traditional plant propagation mode.
In the adventitious bud proliferation process, five kinds of growth regulatory substances are chosen in this experiment, observe drawing from experiment, and no matter the indefinite bud that BA and KT induce is the propagation position of indefinite bud or the growth conditions in young shoot period does not all have very big difference.And when adding the TDZ evoking adventive bud, then have bigger difference with the above two.Illustrate that the activity of TDZ or the mode of action may exist differently with BA and KT, but with regard to a surname pawpaw indefinite bud fast with regard to the breeding, the effect of BA is better than TDZ.Wherein the culture effect of MS+BA0.5mg/L+NAA0.2mg/L combination is best, and its growth coefficient can reach 4.48.
Also obtain following conclusion the experiment of inventor beyond above-mentioned two embodiment:
One, the MS macroelement is to the influence of adventitious bud proliferation
The nutritive element that plant growing need be enriched, because nutritive element is the plant corpus constituent on the one hand, on the other hand, the concentrating of nutrients also has certain regulating action to the growth of plant, and growth is absolutely necessary therefore suitable nutritive element concentration to plant health.From experimental result as can be known, the macroelement of variable concentrations is bred fast to a surname pawpaw has certain influence, and when using the MS medium, the indefinite bud growth is normal and growth coefficient is high; And when answering the 1/2MS medium, the growth coefficient of a surname pawpaw reduces, but difference does not clearly appear in the external form of young shoot; When macroelement is reduced to 1/4, the flavescence of young shoot blade, poor growth, plant is short and small.
Two, the optimization of root media
Stripped the taking root of plant is a relatively easier process, and the indefinite bud of some plants can be taken root in the medium of no growth regulatory substance.Experimental result shows that the root media of a surname pawpaw indefinite bud need add auxins and regulate material, otherwise indefinite bud can not be taken root.NAA, IBA induce the effect of a surname pawpaw adventitious bud rooting close, and IBA is taken root more neat, and it is more relatively to take root, and therefore has effect preferably; The root that NAA induces is thicker.A surname pawpaw indefinite bud can't be taken root in the medium of no growth regulatory substance, and it is inhibited to taking root to add BA.The auxins of high concentration is regulated material and is promoted the indefinite bud base portion to form callus, but generation that can not inducing adventitious root, therefore can not add the growth regulatory substance of higher concentration in root media.
Beneficial effect: the method that adopts a surname pawpaw tissue-culturing quick-propagation provided by the present invention, not only can on the basis that superior families is selected, declare fast and efficiently the multiple target breeding research of pawpaw, also can in production of forestry, provide the method for culturing seedlings that a kind of cycle is short, breeding potential is high, with low cost by the implant mass for a surname pawpaw.
Claims (3)
1. make minimal medium with MS for one kind, induce the method for declaring the pawpaw tissue-culturing quick-propagation with root induction by bud respectively, it is characterized in that adopting a surname's pawpaw seed cotyledon or a surname pawpaw seed embryo to carry out tissue culture.
2. the method for a surname pawpaw tissue-culturing quick-propagation as claimed in claim 1 is characterized in that the optimal medium prescription of evoking adventive bud is MS+NAA0.5mg/L+TDZ 1.0mg/L when adopting a surname pawpaw seed cotyledon to carry out tissue culture; The optimal medium prescription of root induction is 1/2MS+IBA0.2mg/L.
3. the method for a surname pawpaw tissue-culturing quick-propagation as claimed in claim 1 is characterized in that the optimal medium prescription of evoking adventive bud is MS+BA0.5mg/L+NAA 0.2mg/L when adopting a surname pawpaw seed embryo to carry out tissue culture; The optimal medium prescription of root induction is 1/2MS+NAA0.2mg/L.
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| CN102002086A (en) * | 2010-09-15 | 2011-04-06 | 安徽农业大学 | Method for preparing compound oleanolic acid from Xuanzhou pawpaw by cell culture method |
| CN102835315A (en) * | 2012-09-26 | 2012-12-26 | 钦州市林业科学研究所 | Culture medium for cultivating papaya tissue |
| CN103004589A (en) * | 2012-12-03 | 2013-04-03 | 中国计量学院 | Tissue culture and rapid propagation method of agrimony |
| CN104206279A (en) * | 2014-09-21 | 2014-12-17 | 云南集创园艺科技有限公司 | Induction medium for green spherical bodies of hemionitis arifolia |
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2008
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102002086A (en) * | 2010-09-15 | 2011-04-06 | 安徽农业大学 | Method for preparing compound oleanolic acid from Xuanzhou pawpaw by cell culture method |
| CN102002086B (en) * | 2010-09-15 | 2013-05-08 | 安徽农业大学 | Method for preparing compound oleanolic acid from Xuanzhou pawpaw by cell culture method |
| CN102835315A (en) * | 2012-09-26 | 2012-12-26 | 钦州市林业科学研究所 | Culture medium for cultivating papaya tissue |
| CN102835315B (en) * | 2012-09-26 | 2013-08-28 | 钦州市林业科学研究所 | Culture medium for cultivating papaya tissue |
| CN103004589A (en) * | 2012-12-03 | 2013-04-03 | 中国计量学院 | Tissue culture and rapid propagation method of agrimony |
| CN103004589B (en) * | 2012-12-03 | 2014-08-06 | 中国计量学院 | Tissue culture and rapid propagation method of agrimony |
| CN104206279A (en) * | 2014-09-21 | 2014-12-17 | 云南集创园艺科技有限公司 | Induction medium for green spherical bodies of hemionitis arifolia |
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