CN101015279A - Tissue culture method for fast propagation of primula poissonii - Google Patents
Tissue culture method for fast propagation of primula poissonii Download PDFInfo
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- CN101015279A CN101015279A CNA2007100640559A CN200710064055A CN101015279A CN 101015279 A CN101015279 A CN 101015279A CN A2007100640559 A CNA2007100640559 A CN A2007100640559A CN 200710064055 A CN200710064055 A CN 200710064055A CN 101015279 A CN101015279 A CN 101015279A
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- 235000001587 Primula poissonii Nutrition 0.000 title 1
- 241000400342 Primula poissonii Species 0.000 title 1
- 238000012136 culture method Methods 0.000 title 1
- 238000000034 method Methods 0.000 claims abstract description 28
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 14
- 230000001939 inductive effect Effects 0.000 claims description 13
- 238000005286 illumination Methods 0.000 claims description 11
- 230000001954 sterilising effect Effects 0.000 claims description 9
- 239000002689 soil Substances 0.000 claims description 6
- 229960002523 mercuric chloride Drugs 0.000 claims description 5
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims description 5
- 238000012546 transfer Methods 0.000 claims description 4
- 230000003203 everyday effect Effects 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 2
- 241000245063 Primula Species 0.000 abstract description 35
- 235000000497 Primula Nutrition 0.000 abstract description 33
- 241000196324 Embryophyta Species 0.000 abstract description 8
- 230000004083 survival effect Effects 0.000 abstract description 5
- 238000004519 manufacturing process Methods 0.000 abstract description 3
- 238000005516 engineering process Methods 0.000 abstract description 2
- 238000004321 preservation Methods 0.000 abstract description 2
- 235000016311 Primula vulgaris Nutrition 0.000 abstract 1
- 230000006698 induction Effects 0.000 abstract 1
- 238000004904 shortening Methods 0.000 abstract 1
- 238000002054 transplantation Methods 0.000 abstract 1
- 239000002609 medium Substances 0.000 description 12
- 238000009395 breeding Methods 0.000 description 8
- 230000001488 breeding effect Effects 0.000 description 7
- 230000004069 differentiation Effects 0.000 description 4
- 238000011160 research Methods 0.000 description 4
- 241000238631 Hexapoda Species 0.000 description 3
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- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 238000012258 culturing Methods 0.000 description 2
- 239000006160 differential media Substances 0.000 description 2
- 238000011010 flushing procedure Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000008929 regeneration Effects 0.000 description 2
- 238000011069 regeneration method Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- TWFZGCMQGLPBSX-UHFFFAOYSA-N Carbendazim Natural products C1=CC=C2NC(NC(=O)OC)=NC2=C1 TWFZGCMQGLPBSX-UHFFFAOYSA-N 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- JNPZQRQPIHJYNM-UHFFFAOYSA-N carbendazim Chemical compound C1=C[CH]C2=NC(NC(=O)OC)=NC2=C1 JNPZQRQPIHJYNM-UHFFFAOYSA-N 0.000 description 1
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- 239000000463 material Substances 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 230000001902 propagating effect Effects 0.000 description 1
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Classifications
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- Y02P60/216—
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
本发明提供了一种海仙报春的组培快繁方法，包括外植体选择、表面消毒、不定芽诱导、继代培养、生根培养与试管苗移栽。本发明的海仙报春(Primula poisonii)组培快繁方法，一个腋芽可诱导产生40～50个左右的新芽，经过继代30～40天左右增殖系数为3～4，生根率可达到95％以上，移栽成活率几乎为100％，实现了通过快繁技术对优良海仙报春(Primula poisonii)单株的保存，同时大大提高了海仙报春的育苗生产能力，缩短成苗时间又可降低成本，为报春花的大面积推广应用提供了技术支持。The invention provides a method for tissue culture and rapid propagation of Primula chinensis, comprising explant selection, surface disinfection, adventitious bud induction, subculture, rooting culture and test-tube seedling transplantation. According to the tissue culture and rapid propagation method of Primula poisonii of the present invention, one axillary bud can be induced to produce about 40 to 50 new buds, and after about 30 to 40 days of subculture, the multiplication coefficient is 3 to 4, and the rooting rate can reach 95 % or more, the transplanting survival rate is almost 100%, which realizes the preservation of the excellent Primula poisonii (Primula poisonii) single plant through rapid propagation technology, and greatly improves the seedling production capacity of Primula poisonii at the same time, shortening the seedling time It can also reduce the cost, and provides technical support for the large-scale popularization and application of the primrose.
Description
Technical field
The present invention relates to Plant Tissue Breeding, specifically, relate to the herald spring quick breeding method for tissue culture of (Primulapoisonii) of Hai Xian.
Background technology
Hai Xian is heralded spring (Primula poisonii) for the Primulaceae primula, is a kind of very beautiful ornamental plantation flowers.Hai Xian herald spring (Primula poisonii) be the perennial herb flowers, whole plant is rosette-stape, the root-like stock prosperity, upwards give birth to 5 to a plurality of leafages, based on seed propagation, generally in spring after planting, through nourishing and growing in beginning May next year flower, it is slower to grow, and reproduction coefficient is low.Except that seed propagation, the primula vegetative propagation can with method have: plant division (leafage) breeding, tissue culture propagating, the perennial kind of minority can adopt cottage propagation, however the method for division propagation far can not satisfy the production demand.
Tissue culture is an important channel of breeding the good plant kind fast, the existing many relevant reports of having carried out tissue culture of planting of primula, the tissue culture of (Primulapoisonii) does not but have correlative study and report but Hai Xian is heralded spring, therefore need the research Hai Xian tissue culture of (Primula poisonii) of heralding spring, and form the special fast breeding technique system of a cover.
Summary of the invention
The purpose of this invention is to provide the herald spring tissue culture and rapid propagation method of (Primula poisonii) of a kind of Hai Xian, improve Hai Xian herald spring reproduction coefficient, rooting rate and the transplanting survival rate of (Primula poisonii), and, make the herald spring seedling production, quality saving etc. of (Primulapoisonii) of Hai Xian can access better guarantees in conjunction with other relevant technical measures.
In order to realize the object of the invention, the herald spring tissue culture and rapid propagation method of (Primula poisonii) of a kind of Hai Xian of the present invention, explant selection, surface sterilization, adventitious bud inducing, successive transfer culture, culture of rootage and test-tube seedling transplanting.
Described explant is the herald spring axillalry bud of (Primula poisonii) of Hai Xian.
The medium of described adventitious bud inducing and subculture is MS+2.0mg/l 6-BA+0.1mg/lNAA, and pH is 5.8～5.9.Its illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 1500～2000Lux.Lux (lux) is the unit of illumination.
Described root media is MS or MS+0.2mg/l NAA, is preferably MS+0.2mg/lNAA, and pH is 5.8～5.9.
Its illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 2500～3000Lux.
Through cultivated for 2～3 weeks in root media after, hardening was transplanted and is cultivated to the thin turfy soil in greenhouse in 3～4 days.
Explant adopts 0.1% mercuric chloride solution sterilization, 3～4min.
Tissue cultivating seedling suitable culture temperature is between 20～23 ℃, the tissue cultivating seedling flavescence above 25 ℃ even dead gradually.
The transplanting preference temperature of tissue cultivating seedling is between 18～25 ℃, too high or too lowly all is unfavorable for surviving of test-tube plantlet.
Tissue culture and rapid propagation method is particularly: selecting the herald spring axillalry bud of (Primula poisonii) of Hai Xian is explant, rinse well through water, adopt 0.1% mercuric chloride solution sterilization, 3～4min, under aseptic condition, clean repeatedly through distilled water, be seeded among the inducing culture MS+2.0mg/l6-BA+0.1mg/l NAA, pH is 5.8, and to be natural scattering light 2500～3000Lux add artificial fill-in light 1500～2000Lux with interior to illumination condition; Move to same medium propagation afterwards, subculture 2～3 times, root media are MS+0.2mg/l NAA, and pH is 5.8, and illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 2500～3000Lux; Temperature remains between 20～23 ℃; In 2～3 weeks of culture of rootage, hardening moved in the thin turfy soil and cultivates after 3～4 days, kept humidity to be higher than 80%, sprayed water every day 2～3 times.Progressively throwing off coverlay after 2 weeks normally cultivates.
In general with the axillalry bud be explant, bigger to the destructiveness of plant, but relatively precious for some, and be badly in need of expanding numerous material, it is numerous with the axillalry bud to be that explant expands, and can obtain a large amount of regeneration plants at short notice; By the Hai Xian of axillalry bud regeneration (Primulapoisonii) test-tube plantlet growth stalwartness of heralding spring, growth of later stage observed find, can keep its good strains of seeds constant substantially.The test-tube plantlet of tissue-culturing rapid propagation in process of growth, is not vulnerable to infecting of damage by disease and insect, but in order further to prevent the generation of damage by disease and insect, it is for 3～4 times good should spraying carbendazim, flolimat etc. in growth season; Temperature during test-tube seedling transplanting should be controlled between 18～25 ℃, too high or too lowly all is unfavorable for surviving of test-tube plantlet, and should not carry out the transplanting of test-tube plantlet summer.
Tissue culture and rapid propagation method of the present invention is bred Hai Xian herald spring (Primula poisonii), the internal breeding coefficient reached 3～4 in 2 months, rooting rate reaches more than 95%, transplanting survival rate is almost 100%, greatly improved the herald spring reproduction coefficient of (Primula poisonii) of Hai Xian, shorten into the seedling cycle, can reduce cost again, for large tracts of land Landscape Application and factorial seedling growth provide very effective method from now on; Simultaneously, be that explant is bred and numerous soon with the axillalry bud, can keep the herald spring kind of (Primula poisonii) of Hai Xian constant, for the preservation of the comparatively precious rare mutation that occurs in the rearing new variety provides guarantee.
Compared with prior art, the herald spring advantage of tissue culture and rapid propagation method of (Primula poisonii) of Hai Xian of the present invention is:
1. set up a kind of Hai Xian and heralded spring the tissue culture and rapid propagation method of (Primula poisonii), compare traditional primula propagation method of growing seedlings, as seed propagation and division propagation, greatly improved the herald spring reproduction coefficient of (Primula poisonii) of Hai Xian, for scientific research research with produce cultivation technical support is provided;
2. the seedling that obtains by tissue-culturing rapid propagation, it is strong to grow, and the leaf look dark green, and uniformity is strong, becomes seedling easy, and adaptability is stronger, and damage by disease and insect is few, is easy to management;
3. utilize tissue culture and rapid propagation method to breed Hai Xian and herald spring (Primula poisonii), removed the restriction of season and weather, thereby shortened growing-seedling period, increased breeding amount;
4. utilize tissue culture and rapid propagation method to breed Hai Xian and herald spring (Primula poisonii), can keep the characteristic of good species or kind constant, thereby provide effective method for making a variation to preserve in the breeding of new variety research.
Embodiment
Following examples are used to illustrate the present invention, but are not used for limiting the scope of the invention.
Embodiment 1
With Hai Xian (Primula poisonii) blade of heralding spring is that explant is probed into the adventitious bud inducing optimum medium: select for use the seedling leaves that grows fine to probe into adventitious bud inducing optimal medium prescription as explant.Get the herald spring young leaflet tablet running water flushing 2～3h of (Primula poisonii) of Hai Xian, drip cleaning agent concussion 15min removing surface impurity in water, running water is rinsed well repeatedly; Surface sterilization: 70% alcohol disinfecting, 10～15s, aseptic water washing 4～5 times, mercuric chloride sterilization 3～4min of 0.1% uses aseptic water washing 4～5 times again.The blade that to sterilize is cut into 1cm then
2Fritter (band portion vein) be seeded on the inducing culture of the bud of growing thickly, 5～6 of every bottle graft kinds, every kind of medium connects 10 bottles, the dark cultivation.Wherein, the concentration gradient of 6-BA: 0.5mg/l, 1.0mg/l and 2.0mg/l; The concentration gradient of NAA: 0.05mg/l, 0.1mg/l and 0.5mg/l, medium pH is 5.8, adopts the design of two factor completely randomized experiments.
The herald spring differentiation of (Primula poisonii) blade of different 6-BA and NAA concentration affects Hai Xian.Inoculate after 4～5 days, blade begins the swelling shrinkage, and 15 days left and right sides incision thickening of growing fat after 30 days, has medium kind blade to begin the generation that produces callus and a small amount of indefinite bud is arranged.The optium concentration of differentiation adventitious buds is 6-BA 2.0mg/l and NAA 0.1mg/l, and this medium Shanghai celestial being (Primula poisonii) inductivity of heralding spring reaches 22.5%, and the part differentiation and bud formation.
Embodiment 2
With Hai Xian (Primula poisonii) axillalry bud of heralding spring is that explant carries out just cultivating and successive transfer culture: choose the axillalry bud on the plant of robust growth, running water flushing 2～3h, the same blade of sterilization process, be seeded on the MS+6-BA 2.0mg/+NAA 0.1mg/l inducing clumping bud medium 2～3 of every bottle graft kinds; Illumination condition is that natural scattering light 3000Lux adds artificial secondary light source 1800 ± 200Lux, light application time 14h.
When axillalry bud is seeded on the differential medium, constantly extract young leaves out, beginning in 10 days, the petiole of extracting young leaves out is sturdy, and base portion reddens, and forms the light green color callus gradually, begins to have the differentiation of indefinite bud after 2 weeks; Inoculate after 40 days, on the new petiole of extracting out and blade of axillalry bud, differentiate a large amount of indefinite buds.
After 2 months, on average to differentiate the number of indefinite bud be 40～50 to each axillalry bud to statistical result showed in inoculation on the medium of MS+6-BA 2mg/l+NAA 0.1mg/l, significantly the quantity of the indefinite bud of inducing more than blade.Simultaneously, temperature plays very crucial effect for inducing with the growth of tissue cultivating seedling of indefinite bud, and between 20～23 ℃, tissue cultivating seedling leaf look dark green, robust growth; Surpass 25 ℃, the tissue cultivating seedling flavescence is also dead gradually.Under such condition of culture, 2 months value-added coefficient of test-tube plantlet reaches 3～4, has greatly improved the herald spring reproduction coefficient of (Primula poisonii) of Hai Xian.
Embodiment 3
The herald spring culture of rootage of (Primula poisonii) tissue cultivating seedling of Hai Xian: when indefinite bud grows 2～3 leaves on differential medium, it is excised from callus, be transferred to root media, prescription: among MS, the MS+NAA (0.1mg/l, 0.2mg/l, 0.5mg/l), pH is 5.8; Illumination condition is that natural scattering light 3000 Lux add artificial secondary light source 3000Lux, light application time 14h.
From taking root the needed time, the required time of taking root in medium MS and MS+0.2mg/l NAA is the shortest, it is 8～10 days, but in MS+0.2mg/l NAA, tissue cultivating seedling amount of taking root and growing way are significantly better than the MS minimal medium, the tissue cultivating seedling average height can reach 4.8cm, and rooting rate is more than 95%.
Embodiment 4
The herald spring transplanting of (Primula poisonii) tissue cultivating seedling of Hai Xian: culture of rootage is after two weeks, treats that the average every strain of tissue cultivating seedling takes root 4～5, when root reaches 1cm, can begin to prepare for transplanting.Hardening is the transition before the test-tube seedling transplanting, can help test-tube plantlet progressively to adapt to external environment, makes test-tube plantlet more healthy and stronger, and the leaf look more dark green, thereby improves survival rate.
Hai Xian was heralded spring (Primula poisonii) test-tube plantlet hardening after 3～4 days, transplanted to the thin turfy soil in greenhouse, and turfy soil is paved with the cave dish in advance, and clear water soaks into; Keep humidity more than 80% behind the test-tube seedling transplanting, cover and spray water film and every day 2～3 times, 2 weeks can progressively be carried out normal management later, and transplanting survival rate is almost 100%.
Though above the present invention is described in detail with a general description of the specific embodiments, on basis of the present invention, can make some modifications or improvements it, this will be apparent to those skilled in the art.Therefore, these modifications or improvements all belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Claims (9)
1. tissue culture and rapid propagation method that extra large celestial being is heralded spring, comprise explant selection, surface sterilization, adventitious bud inducing, successive transfer culture, culture of rootage and test-tube seedling transplanting, it is characterized in that, described explant is the axillalry bud that Hai Xian is heralded spring, the medium of described adventitious bud inducing and subculture is MS+2.0mg/l 6-BA+0.1mg/l NAA, pH is 5.8～5.9, and described root media is MS or MS+0.2mg/l NAA, and pH is 5.8～5.9.
2. tissue culture and rapid propagation method according to claim 1 is characterized in that, when described adventitious bud inducing, successive transfer culture, illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 1500～2000Lux.
3. tissue culture and rapid propagation method according to claim 1 and 2 is characterized in that, described root media is MS+0.2mg/l NAA.
4. according to any described tissue culture and rapid propagation method of claim 1～3, it is characterized in that during described culture of rootage, illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 2500～3000Lux.
5. according to any described tissue culture and rapid propagation method of claim 1～4, it is characterized in that the cultivation temperature of described tissue cultivating seedling is 20～23 ℃.
6. according to any described tissue culture and rapid propagation method of claim 1～5, it is characterized in that the transplanting temperature of described tissue cultivating seedling is 18～25 ℃.
7. according to any described tissue culture and rapid propagation method of claim 1～6, it is characterized in that described explant adopts 0.1% mercuric chloride solution sterilization, 3～4min.
8. according to any described tissue culture and rapid propagation method of claim 1～7, it is characterized in that described tissue cultivating seedling is through cultivating 2-3 after week in root media, hardening was transplanted and is cultivated to the thin turfy soil in greenhouse in 3～4 days.
9. according to any described tissue culture and rapid propagation method of claim 1～8, it is characterized in that, the axillalry bud that selects Hai Xian to herald spring is an explant, rinse well through water, adopt 0.1% mercuric chloride solution sterilization, 3～4min, under aseptic condition, clean repeatedly, be seeded among the inducing culture MS+2.0mg/l 6-BA+0.1mg/l NAA through distilled water, pH is 5.8, and illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 1500～2000Lux; Move to same medium propagation afterwards, subculture 2～3 times, root media are MS+0.2mg/l NAA, and pH is 5.8, and illumination condition is that natural scattering light 2500～3000Lux adds artificial fill-in light 2500～3000Lux; Temperature remains between 20～23 ℃; In 2～3 weeks of culture of rootage, hardening moved in the thin turfy soil and cultivates after 3～4 days, kept humidity to be higher than 80%, sprayed water every day 2～3 times.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101965796A (en) * | 2010-10-14 | 2011-02-09 | 北京林业大学 | Method for performing tissue culture and rapid propagation on primula saxatilis |
| CN103070064A (en) * | 2012-12-17 | 2013-05-01 | 中国科学院昆明植物研究所 | Artificial hybrid breeding method of Primula L. |
| CN107041308A (en) * | 2017-04-26 | 2017-08-15 | 江苏农林职业技术学院 | A kind of primrose root tissue culture propagation method |
| CN112293251A (en) * | 2020-10-19 | 2021-02-02 | 云南中医药大学 | Artificial efficient primula forbesii breeding method |
-
2007
- 2007-02-16 CN CN2007100640559A patent/CN101015279B/en not_active Expired - Fee Related
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101965796A (en) * | 2010-10-14 | 2011-02-09 | 北京林业大学 | Method for performing tissue culture and rapid propagation on primula saxatilis |
| CN101965796B (en) * | 2010-10-14 | 2012-07-25 | 北京林业大学 | Method for performing tissue culture and rapid propagation on primula saxatilis |
| CN103070064A (en) * | 2012-12-17 | 2013-05-01 | 中国科学院昆明植物研究所 | Artificial hybrid breeding method of Primula L. |
| CN103070064B (en) * | 2012-12-17 | 2013-12-18 | 中国科学院昆明植物研究所 | Artificial hybrid breeding method of Primula L. |
| CN107041308A (en) * | 2017-04-26 | 2017-08-15 | 江苏农林职业技术学院 | A kind of primrose root tissue culture propagation method |
| CN112293251A (en) * | 2020-10-19 | 2021-02-02 | 云南中医药大学 | Artificial efficient primula forbesii breeding method |
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| CN101015279B (en) | 2011-05-11 |
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