CN101743909A - Tissue culture and propagation method of illicium plant illicium benryi dies - Google Patents
Tissue culture and propagation method of illicium plant illicium benryi dies Download PDFInfo
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- CN101743909A CN101743909A CN200910232666A CN200910232666A CN101743909A CN 101743909 A CN101743909 A CN 101743909A CN 200910232666 A CN200910232666 A CN 200910232666A CN 200910232666 A CN200910232666 A CN 200910232666A CN 101743909 A CN101743909 A CN 101743909A
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- illicium
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Abstract
The invention discloses a tissue culture and propagation method of illicium plant illicium benryi dies. In the method, an explant which is in tissue culture and propagation with a sprout on the top of a current-year branch of the illicium benryi dies is firstly subjected to surface treatment in an ethanol solution and then is subjected to sterilization treatment in an HgC12 solution; then a bud slice is cut down from the branch to be inoculated to an initial medium and cultured under the conditions of illumination for 12-15h everyday with the illumination intensity of 1500-2000LX and the temperature of 23 DEG C plus/minus 1 DEG C; and new leaves are transferred onto a subculture medium to be continuously cultured after germinating, and a tissue culture seedling is obtained. The invention solves the problems of less resources, acarpia, incapability of harvest seeds, difficult vegetative propagation and critical shortage of seedling of the traditional illicium benryi dies, can accelerate propagation, provide a new way to satisfy the market demands of new variety seedling, increase a new type as a rare ornamental plant and have a favorable application potential in landscaping.
Description
Technical field
The present invention relates to the tissue culture propagation of a kind of Winteraceae plant " my child lotus ", be specifically related to the selection of explant kind, initial culture method, successive transfer culture method and medium component thereof in " my child lotus " tissue culture procedures.
Background technology
Winteraceae (Illiciaceac) is a single genus section plant, about 50 kinds of the whole world, and there are 28 kinds and 2 mutation in China, mainly is distributed in China west and south to the eastern region; They are evergreen dungarunga or shrub, and fragrance is arranged, and like dark and damp environment, grow in woodland stream limit more; Flower both sexes, armpit are given birth to or give birth on nearly top, redness, and tepal is 7～21 pieces, the florescence is generally in 4～June, the ornamental value that tool is certain.The Winteraceae plant is mainly by seminal propagation, and its fruit is poisonous mostly, all has certain toxicity as the fruits of Winteraceae plants such as thick grass, red fennel and anisetree bark, causes the application of this type of plant in afforestation to be subjected to the restriction of Environmental security factor.
Rare ornamental plants " my child lotus " is Winteraceae Illicium (Illicium Linn.) plant, evergreen dungarunga, the about 3～8m of the height of tree, tree crown obovate; The bark taupe, single leaf alternate or closely verticillate, lanceolar, blade keratin degree height, doubling is easily broken, branches and leaves tool peat-reek; Flower shape lotus shape, pattern is ruddy, and the florescence, when blooming, pattern was ruddy in 4～May, was the reversal of the natural order of things lotus shape, and as small lantern, several are hung on the end of the branch, and it very is beautiful being embedded between the greenery, has very high ornamental plantation and is worth.Especially " my child lotus " do not observe its normal phenomenon as a result so far in Suzhou District, thereby can avoid people and animals to eat fruit by mistake and cause the defective of poisoning, and the greening and the garden that can be widely used in street, park beautify.
Carry according to " Taihu Lake appendix " secretary: " my child lotus, wood are like osmanthus, and flower is big as chess piece, look shape and lotus flower together, Hua Buxiang, its leaf of Norway is smelt it, Xin Fen is fennel seemingly.Wuzhong is not to there being this kind, and Dongshan Weng Hanjin is the west of a river, Yunnan county magistrate between Shunzi, takes and return, and the back owns for Xi Shi, and treasure is curiosa.The figured cloth knot, root does not have rudiment, its kind of tendency to develop non-availability ... ".Though " my child lotus " imports the history in existing more than 300 year in Jiangsu and Zhejiang Provinces area into, its resource is most deficient, and artificial plantation few at present, at Suzhou District poke strain bearing tree only, can be rated as rare tree species.This is because " my child lotus " is barren, can't gather in the crops seed; Grafting lacks stock, and the cottage propagation survival rate is very low, and the vegetative propagation difficulty causes a seedling difficulty to be asked, and these have all limited " my child lotus " application in afforestation.
In recent years, the application of plant tissue culture technique on ornamental plants more and more widely, especially the application on the low plant of natural breeding rate enjoys people to pay attention to, a series of successfully report is also being arranged aspect the tissue-culturing rapid propagation of woody plant, but in the Winteraceae plant, do not succeed so far, problems such as forefathers did not carry out system research to the key technique of " my child lotus " tissue culture yet, and were seriously polluted such as explant in its group training process, that the blastogenesis length that exsomatizes stops, easy brownization is withered are not effectively solved.
Summary of the invention
The purpose of this invention is to provide explant kind, Biocidal treatment method, initial culture method, successive transfer culture method and medium component thereof that a cover is applicable to Winteraceae plant " my child lotus " tissue culture, solve that " my child lotus " explant pollutes easily, difficult problems such as " mashed core " and brownization, for the quick breeding of rare plant " my child lotus " provides effective technical support.
For achieving the above object, technical scheme of the present invention is: a kind of Winteraceae plant---the tissue culture propagation of " my child lotus ", cultivate in vitro tissue, and comprise the steps:
1) bud of sprouting with " my child lotus " current-year branch top is a starting material, after cleaning solution immersion, flowing water flushing operation are cleaned, dries indoor, as the explant of tissue culture propagating;
2) above-mentioned explant is carried out the surface treatment in 5～6 seconds in concentration is the ethanolic solution of volume ratio 70～75% after, be the HgCl of w/v 0.1% in concentration
2Carry out 5～6 minutes sterilization processing in the solution, fully clean 5～6 times with sterile water again, finish the sterilization of explant is handled;
3) branch behind the above-mentioned sterilization cuts the bud sheet, be inoculated on the initial culture base, illumination every day 12～15 hours, intensity of illumination is that 1500～2000LX, temperature are to cultivate under 23 ℃ ± 1 ℃ the condition, treat to transfer to after young leaves is sprouted and continue on the subculture medium to cultivate, obtain normotrophic tissue cultivating seedling.
Described initial culture base is White medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1, add sucrose and agar powder, volume ratio by weight, sucrose concentration is 3% of an initial culture base, agar powder concentration is 0.6% of initial culture base; Regulating pH is 5.6～5.8.
Described subculture medium is White medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.1mg.L
-1, add sucrose and agar powder, volume ratio by weight, sucrose concentration is 3% of an initial culture base, agar powder concentration is 0.6% of initial culture base; Regulating pH is 5.6～5.8.
The present invention compared with prior art has following advantage:
1, the vegetable material of the present invention's use is " a my child lotus ", it shows as the not result of only blooming in certain areas, these are different with the most poisonous situation of other Winteraceae fruits, " my child lotus " tissue cultivating seedling that adopts technical solution of the present invention to provide, can be used as a kind of ornamental plants new varieties, the application in afforestation has better safety.
2, the technical method that proposes of the present invention has the characteristics of simple and easy to do, operability and favorable reproducibility.
Embodiment
Below in conjunction with embodiment the present invention is further described:
Embodiment: adopting the bud of not sprouting on " my child lotus " current-year branch top respectively is starting material with the bud of sprouting, and compares test.With above-mentioned two kinds of materials through cleaning solution soak, after the flowing water flushing, respectively with concentration be carry out the surface treatment in 5 seconds in the ethanolic solution of volume ratio 75% after, be the HgCl of w/v 0.1% in concentration
2Carry out 6 minutes sterilization processing in the solution, use sterile water wash at last 5～6 times, put and blot excessive moisture on the filter paper, cut explant at the sprout downside and be inoculated in medium.
Cultivation results is, the culture effect of " my child lotus " bud of not sprouting is relatively poor, easily produces endogenous and pollutes, and brownization rate is higher to be 22.2～46.9%, and survival rate is lower, only is 2.8～6.2%.Secondly, be the occasion of explant at the employing bud of sprouting, use 0.1%HgCl
2When solution sterilization was handled 3 minutes, pollution rate was up to 100%; When sterilization time was 9 minutes, though do not see pollution, sprout stem apex lethality reached 33.3%, and brownization rate reaches 50.0%; And sterilize 6 minutes the time, pollution rate only is 8.3%, survival rate can be up to 91.7%.Show that thus " my child lotus " bud of sprouting more is applicable to and the explant material of tissue-culturing rapid propagation helps setting up the aseptic culture system.
With MS (Murashige ﹠amp; Skoog, 1962) be minimal medium, anti-brownization agent PVP, 6-BA and the NAA of additional variable concentrations compare test as the initial culture base.Wherein medium I is MS+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1, medium ii is 1/2MS+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1, medium ii I is B5 medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1, medium IV is White medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1Each medium all adds sucrose and agar powder, volume ratio by weight, and sucrose concentration is 3% of an initial culture base, agar powder concentration is 0.6% of initial culture base; Regulate pH 5.6～5.8.
Cultivation results is, the upgrowth situation of explant in medium I, II is relatively poor, and brownization rate is more than 90%; In medium ii I, IV, the explant that upgrowth situation is good accounts for more than 80%, and brownization rate is lower.In medium I, II, III, the explant lower cut-out all produces callus, have only the explant lower cut-out among the medium IV not produce callus, and the explant ratio that upgrowth situation is good has reached 90.9%, show that medium IV ratio is more suitable for the initial culture of " my child lotus " stripped bud, has also obtained similar cultivation results in successive transfer culture.
In subculture medium, add 500mg.L respectively
-1And 1000mg.L
-1Anti-brownization agent PVP is not to add PVP in contrast, brownization of the explant situation after the comparison different disposal.The result shows that when not adding PVP, whole brownization of explant lower cut-out spread to whole explant then; When adding 1000mg.L
-1During PVP, though explant lower cut-out and itself brownization rate have reduction, its retarded growth; When adding 500mg.L
-1When resisting brownization agent PVP, explant lower cut-out and itself brownization rate are all very low, and it is good to grow.Therefore, add 500mg.L
-1Anti-brownization agent PVP explant is had preferably anti-brownization effect, and several nothings influences of growing that the bud that exsomatizes is cultivated.
Claims (3)
1. the tissue culture propagation of a Winteraceae plant " my child lotus " is characterized in that, carries out in vitro tissue and cultivates, and comprises the steps:
1) bud of sprouting with " my child lotus " current-year branch top is a starting material, after cleaning solution immersion, flowing water flushing operation are cleaned, dries indoor, as the explant of tissue culture propagating;
2) above-mentioned explant is carried out the surface treatment in 5～6 seconds in concentration is the ethanolic solution of volume ratio 70～75% after, be the HgCl of w/v 0.1% in concentration
2Carry out 5～6 minutes sterilization processing in the solution, fully clean 5～6 times with sterile water again, finish the sterilization of explant is handled;
3) branch behind the above-mentioned sterilization cuts the bud sheet, be inoculated on the initial culture base, illumination every day 12～15 hours, intensity of illumination is that 1500～2000LX, temperature are to cultivate under 23 ℃ ± 1 ℃ the condition, treat to transfer to after young leaves is sprouted and continue on the subculture medium to cultivate, obtain normotrophic tissue cultivating seedling.
2. the tissue culture propagation of Winteraceae plant according to claim 1 " my child lotus " is characterized in that: described initial culture base is White medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.5mg.L
-1, add sucrose and agar powder, volume ratio by weight, sucrose concentration is 3% of an initial culture base, agar powder concentration is 0.6% of initial culture base; Regulating pH is 5.6～5.8.
3. the tissue culture propagation of Winteraceae plant according to claim 1 " my child lotus " is characterized in that: described subculture medium is White medium+500mg.L
-1Anti-brownization agent PVP+6-BA 2.0mg.L
-1+ NAA 0.1mg.L
-1, add sucrose and agar powder, volume ratio by weight, sucrose concentration is 3% of an initial culture base, agar powder concentration is 0.6% of initial culture base; Regulating pH is 5.6～5.8.
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| CN2009102326669A CN101743909B (en) | 2009-12-04 | 2009-12-04 | Tissue culture and propagation method of ''Haierlian'' of Illiciaceae plant |
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| CN2009102326669A CN101743909B (en) | 2009-12-04 | 2009-12-04 | Tissue culture and propagation method of ''Haierlian'' of Illiciaceae plant |
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| CN101743909A true CN101743909A (en) | 2010-06-23 |
| CN101743909B CN101743909B (en) | 2012-06-20 |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103734014A (en) * | 2014-01-06 | 2014-04-23 | 广西壮族自治区药用植物园 | Tissue culture rapid propagation method for anisetree barks |
| CN107509635A (en) * | 2017-10-11 | 2017-12-26 | 陈金水 | A kind of in vitro tissue culture and rapid propagation method of Chinese anise |
| CN115250923A (en) * | 2022-09-26 | 2022-11-01 | 云南省农业科学院园艺作物研究所 | Tissue culture and rapid propagation method of star anise |
-
2009
- 2009-12-04 CN CN2009102326669A patent/CN101743909B/en not_active Expired - Fee Related
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103734014A (en) * | 2014-01-06 | 2014-04-23 | 广西壮族自治区药用植物园 | Tissue culture rapid propagation method for anisetree barks |
| CN103734014B (en) * | 2014-01-06 | 2016-03-30 | 广西壮族自治区药用植物园 | A kind of quick breeding method for tissue culture of anisetree bark |
| CN107509635A (en) * | 2017-10-11 | 2017-12-26 | 陈金水 | A kind of in vitro tissue culture and rapid propagation method of Chinese anise |
| CN115250923A (en) * | 2022-09-26 | 2022-11-01 | 云南省农业科学院园艺作物研究所 | Tissue culture and rapid propagation method of star anise |
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| CN101743909B (en) | 2012-06-20 |
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