CN101336613A - Black bamboo tissue culture medium and tissue culture and rapid propagation method - Google Patents
Black bamboo tissue culture medium and tissue culture and rapid propagation method Download PDFInfo
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- CN101336613A CN101336613A CNA2008100633846A CN200810063384A CN101336613A CN 101336613 A CN101336613 A CN 101336613A CN A2008100633846 A CNA2008100633846 A CN A2008100633846A CN 200810063384 A CN200810063384 A CN 200810063384A CN 101336613 A CN101336613 A CN 101336613A
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/10—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in agriculture
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/40—Afforestation or reforestation
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Abstract
The invention relates to a tissue culture medium for the purple bamboo, which is composed of a clustered bud induction culture medium and a root induction culture medium. The clustered bud induction culture medium comprises the following components: MS, 3.0 mg/L of BA, 0.01 mg/L of TDZ, 0.25% of the total weight of crystal agar and 3% of the total weight of sucrose. The root induction culture medium comprises the following components: MS, 3mg/L of BA, 1mg/L of NAA, 0.25% of the total weight of crystal agar and 3% of the total weight of sucrose. Both the pH values of the two culture media are 5.0. The method of tissue culture and rapid propagation using the culture media comprises the following four steps: collection and sterilization of explants, induction of clustered buds, multiplication and rapid propagation of adventitious buds, and root induction and transplanting. With the tissue culture and rapid propagation method using the culture media, the clustered bud induction rate reaches 42%, and the adventitious bud multiplication coefficient is as high as 13 times, the root induction rate reaches 90%, and the survival rate of transplanting test-tube plantlets is about 90%. The invention overcomes the difficulty in tissue culture and plant regeneration of the running bamboo and provides a new approach to the rapid propagation and cultivation of the purple bamboo for yard planting, landscaping and large-area afforestation.
Description
[technical field]
The present invention relates to the tissue culture medium (TCM) and the tissue culture and rapid propagation method of tissue culture medium (TCM) and tissue culture and rapid propagation method, the especially black bamboo of a kind of middle scattered bamboo of path.
[background technology]
Black bamboo (Phyllostachys nigra), have another name called black bamboo, Wu Zhu, grass family Bambusoideae Phyllostachys, scattered bamboo kind, new bar green, biennial bar fades to atropurpureus, and greenery are verdant, and attitude is elegant, has very high ornamental value, be the excellent material of making furniture, musical instrument and handicraft, breed by transplanting female bamboo or bamboo whip all the time that speed is slow, land used is many, transportation is inconvenient, cost is high and restricted by season again.In recent years, the strong-willed person who explores bamboo class tissue culture and fast breeding technique is increasing, and achievement is respectively arranged.Comprehensive bamboo class Study on tissue culture present situation, the tender material of children induces the callus ratio, and to induce regeneration plant easy, the explantation tissue that takes from ripe bamboo induces into embryonal connective tissue and regeneration plant, difficulty is bigger, pick up from scattered ripe bamboo explantation tissue and induce regeneration plant, for the bamboo that grows thickly, extremely difficult especially.By retrieval and market survey, do not see that so far the explantation tissue with ripe black bamboo induces the document announcement and relevant in kind appearance of regeneration plant.
[summary of the invention]
Induce successfully the realistic situation of regeneration plant at the scattered bamboo that does not see maturation so far especially black bamboo explantation tissue, task of the present invention has and provides the medium that a kind of induced bundle is sprouted at three: one, two provide a kind of medium of root induction, and three provide a whole set of black bamboo quick breeding method for tissue culture.
Above-mentioned task can realize by following measure:
This black bamboo tissue culture medium, be divided into induced bundle sprout medium and root induction medium, it all is minimal medium with MS, be by MS add 0.3 1 or 3 or the BA of 10mg/L be the benzyl aminoadenine add again 0.001 0.01 or 0.1 or the TDZ of 1mg/L be the thiadiazoles phenylurea, the additional quartzy agar that accounts for gross weight 0.25%, 3% sucrose are mixed with the induced bundle medium of sprouting; By MS add 0.3 1 or 3 or the BA of the 10mg/L NAA that adds 1.0mg/L again be α-naa, additionally account for the quartzy agar of gross weight 0.25%, 3% sucrose is mixed with the root induction medium.The pH value of two medium is 3 or 4 or 5 or 5.7 or 6 or 7.
The preferred version of this black bamboo tissue culture medium is that the sprout addition of BA in the medium of induced bundle is that the addition of 3mg/L, TDZ is 0.01mg/L; The addition of BA in the medium of root induction is 3mg/L, and the pH value of two medium is 5.
Carry out tissue-culturing rapid propagation process the following step with this black bamboo tissue culture medium:
(1) explant collection and sterilization: sprout tender stem eye the spring of collection greenhouse pot culture black bamboo, take back the laboratory, peel off the bar sheaths of bamboo shoots, running water flushing 2h, in the 5%NaCl0 solution of 10 times of dilutions, vacuum condition soaks 10min down, with twice of aseptic water washing, soak 4min again in the NaCl0 of same concentrations solution, behind the aseptic water washing three times, microscopically cuts the stem-tip tissue that is about 0.5mm;
(2) grow thickly the inducing of bud: will cut stem-tip tissue be inoculated in the medium that induced bundle as claimed in claim 1 sprouts, under intensity of illumination 2500Lux, 25 ± 2 ℃ of temperature, the illumination 16h/d condition, the bud beginning sprouts behind the 7d, base portion bears the new bud of growing thickly behind the 45d;
(3) adventitious bud proliferation is with numerous soon: the bud cutting-out of will be above-mentioned growing thickly is inoculated in induced bundle as claimed in claim 1 and sprouts in the medium intensity of illumination 2500Lux, illumination 16h/d under 25 ± 2 ℃ of conditions, extends behind the 15d, successive transfer culture, growth coefficient reaches 13 times behind the 100d;
(4) root induction and transplanting: the bud 3-4 of growing thickly after will extending is individual to be one clump, place 300 respectively, 1000, the IBA of 3000ppm is 5-15min in the indolebutyric acid solution, be inoculated in the root media as claimed in claim 1 more respectively, intensity of illumination 2500Lux, illumination 16h/d, under 25 ± 2 ℃ of conditions, begin behind the 20d to take root, after cultivating 125d, rooting rate reaches 70-90%, get well-grown regeneration plant and place the domestication chamber, 2 weeks of domestication transplant under the high light 20000Lux, use the medium of clear water flush away root earlier, planting in volume ratio is vermiculite: perlite: in the matrix of peat=1: 1: 1, single flower pot coat transparent plastic bag, this bag cut an osculum in per two days, abandoned bag immigration greenhouse plantation after the week.
The invention has the beneficial effects as follows and broken through the ripe scattered bamboo of middle path, especially the stem apex explantation tissue of black bamboo induces the difficult problem of regeneration plant, change traditional method that female bamboo is transplanted, the bamboo whip is transplanted, female bamboo, land area, cultivation time and cost of transportation have been saved, and be not subjected to the restriction in season, for the sight spot provides the effective way of breeding the black bamboo seedling fast sufficiently with bamboo, large tracts of land greening with bamboo.
[embodiment]
The present invention is described in further detail below in conjunction with embodiment: the bamboo class tissue culture of China goes out regeneration plant, still is in the starting stage, and scattered bamboo is again because of a little less than its neomorph ability, than the more difficult success of the bamboo that grows thickly.What move ahead is with the numerous bud of bud, the mode through dedifferentiation does not realize, and what select for use is tissue such as plant, embryo childhood, does not see that useful adult plant explant group trains into the report of regeneration plant, and numerous bamboo class groups training practitioners generally acknowledge the scattered bamboo kind tissue cultivating seedling difficulty of taking root.Therefore, main direction of the present invention is to create good artificial environment, sets about from histocyte propagation, the condition of taking root of improving black bamboo, and captures scattered bamboo group training difficult point.For this reason, to the selecting for use, match of culture medium raw material, concentration, pH value, illumination, temperature have been done series of contrast, in the medium of sprouting at the preparation induced bundle, adopted seven kinds of medium that the black bamboo stem-tip tissue is inoculated, filtered out inductivity up to 42% medium; Filter out the medium of growth coefficient, thereby provide material base for histiocytic propagation up to 13 times.At scattered bamboo kind tissue cultivating seedling take root the difficulty problem, successively soak 5-15min in the IBA solution with 300ppm, 1000ppm, three kinds of variable concentrations of 3000ppm, inoculate in the root media of MS+BA 3mg/L+NAA 1mg/L, rooting rate reaches 70-90%, the scheme that the contrast back is selected 1000ppm concentration, soaked 10min.The purpose that clump bud after the elongation is soaked in IBA earlier is because black bamboo is the bamboo kind of easy brownization, the IBA of high concentration is inoculated in the medium that contains the BA basic element of cell division after stimulating, can alleviate brownization degree, to strengthen rootability, therefore also need inoculate in the medium of this root induction, just realize 90% rooting rate.In a word, the black bamboo group is trained out regeneration plant, is that the treatment conditions, medium, matrix etc. to explant are carried out strict results of screening.
This black bamboo medium is minimal medium with MS, appositional growth plain and the basic element of cell division, quartzy agar and sucrose, and addition is chosen to be the quartzy agar that accounts for medium gross weight 0.25%, 3% sucrose, the pH value of medium is decided to be 5.0.Again under other composition and addition same case, press the different of growth hormone and basic element of cell division composition and addition, be mixed with the medium of two kinds of different induction periods: the one, the medium that induced bundle is sprouted is the MS+BA 3+TDZ 0.01 (mg/L of unit, down together), the 2nd, the medium of root induction is MS+BA 3+NAA 1.Above-mentioned medium is prepared according to a conventional method.The bud cultivation effect of the pH value 4-5.7 of medium than 3,6-7 good, the best is 5.
Stem-tip tissue with black bamboo is made explant, the step of carrying out tissue culture and rapid propagation method following (each numerical value of employing is the numerical value of preferred plan, therefore, can be considered most preferred embodiment):
(1) explant collection and sterilization: sprout tender stem eye the spring of collection greenhouse pot culture black bamboo, take back the laboratory, peel off the bar sheaths of bamboo shoots, running water flushing 2h, in the 5%NaCl0 solution of 10 times of dilutions, vacuum condition soaks 10min down, with twice of aseptic water washing, soak 4min again in the NaCl0 of same concentrations solution, behind the aseptic water washing three times, microscopically cuts the stem-tip tissue that is about 0.5mm;
(2) grow thickly the inducing of bud: will cut stem-tip tissue be inoculated in the medium of sprouting as claim 1,2 described induced bundles, under intensity of illumination 2500Lux, 25 ± 2 ℃ of temperature, the illumination 16h/d condition, the bud beginning sprouts behind the 7d, and base portion bears the new bud of growing thickly behind the 45d;
(3) adventitious bud proliferation is with numerous soon: the above-mentioned bud of growing thickly is downcut, be inoculated in the medium of sprouting as claim 1,2 described induced bundles intensity of illumination 2500Lux, illumination 16h/d under 25 ± 2 ℃ of conditions, extends behind the 15d, successive transfer culture, growth coefficient reaches 13 times behind the 100d;
(4) root induction and transplanting: the bud 3-4 of growing thickly after will extending is individual to be one clump, the IBA that places 1000ppm is that indolebutyric acid solution soaks 10min, inoculate as claim 1, in the 2 described root medias, intensity of illumination 2500Lux, illumination 16h/d, under 25 ± 2 ℃ of conditions, begin behind the 20d to take root, after cultivating 125d, rooting rate reaches 90%, get well-grown regeneration plant and place the domestication chamber, 2 weeks of domestication transplant under the high light 20000Lux, use the medium of clear water flush away root earlier, planting in volume ratio is vermiculite: perlite: in the matrix of peat=1: 1: 1, every single flower pot coat transparent plastic bag, this bag cut an osculum in per two days, abandoned bag immigration greenhouse plantation after the week.
In comparative trial, the addition of basic element of cell division BA is selected 0.3mg/L, 1mg/L, 3mg/L, 10mg/L respectively, and add and do not add growth hormone NAA, carried out comparative trial.Do not adding under the NAA situation, the inducing clumping bud rate reaches 42%; Adding BA is 10 buds of growing thickly of appearance of 3mg/L, and performance is good; All do not sprout after adding NAA, so induced bundle is sprouted and can not be added NAA in the medium.
In the addition comparative trial of pair cell mitogen TDZ, addition is respectively 0.001mg/L, 0.01mg/L, 0.1mg/L, 1mg/L, can both induce the bud of growing thickly, the bud that with the addition is 0.01mg/L is maximum, therefore, induce the situation of effect after comprehensive two kinds of growth hormone add, the decision induced bundle is sprouted, and to select best proportioning be the TDZ of the BA+0.01mg/L of 3mg/L to medium.
In the root induction medium, select addition or the 3mg/L of basic element of cell division BA, the addition of growth hormone NAA is 1mg/L, soaks the growth hormone IBA concentration of usefulness and selects 1000ppm, soak time 10min; The propagation situation of medium optimal pH 5.0 buds is best.
Other MS, quartzy agar, sucrose are definite value, carefully do not state one by one.
Under the cultivation with above-mentioned all conditions, test after 125 days, the bud of growing thickly of black bamboo group training is 2-4, the long 13-16mm of bud, the long 10-12cm of root, 4 of radicals.
Claims (3)
1, a kind of black bamboo tissue culture medium, be divided into induced bundle sprout medium and root induction medium, it all is minimal medium with MS, it is characterized in that by MS+0.3 1 or 3 or the BA of 10mg/L be the benzyl aminoadenine and 0.001 or 0.01 0.1 or the TDZ of 1mg/L be the thiadiazoles phenylurea, the additional quartzy agar that accounts for gross weight 0.25%, 3% sucrose are mixed with the induced bundle medium of sprouting; By MS+0.3 1 or 3 or the BA of 10mg/L and the NAA of 1.0mg/L be α-naa, additionally account for the quartzy agar of gross weight 0.25%, 3% sucrose is mixed with the root induction medium.The pH value of two medium is 3 or 4 or 5 or 5.7 or 6 or 7.
2, black bamboo tissue culture medium as claimed in claim 1 is characterized in that the sprout addition of BA in the medium of said induced bundle is that the addition of 3mg/L, TDZ is 0.01mg/L; The addition of BA in the medium of root induction is 3mg/L, and the pH value of two medium is 5.
3, black bamboo tissue culture medium as claimed in claim 1 or 2 carries out the method for tissue-culturing rapid propagation, it is characterized in that through the following step:
(1) explant collection and sterilization: sprout tender stem eye the spring of collection greenhouse pot culture black bamboo, take back the laboratory, peel off the bar sheaths of bamboo shoots, running water flushing 2h, in the 5%NaC10 solution of 10 times of dilutions, vacuum condition soaks 10min down, with twice of aseptic water washing, soak 4min again in the NaC10 of same concentrations solution, behind the aseptic water washing three times, microscopically cuts the stem-tip tissue that is about 0.5mm;
(2) grow thickly the inducing of bud: will cut stem-tip tissue be inoculated in the medium that induced bundle as claimed in claim 1 sprouts, under intensity of illumination 2500Lux, 25 ± 2 ℃ of temperature, the illumination 16h/d condition, the bud beginning sprouts behind the 7d, base portion bears the new bud of growing thickly behind the 45d;
(3) adventitious bud proliferation is with numerous soon: the bud cutting-out of will be above-mentioned growing thickly is inoculated in induced bundle as claimed in claim 1 and sprouts in the medium intensity of illumination 2500Lux, illumination 16h/d under 25 ± 2 ℃ of conditions, extends behind the 15d, successive transfer culture, growth coefficient reaches 13 times behind the 100d;
(4) root induction and transplanting: the bud 3-4 of growing thickly after will extending is individual to be one clump, place 300 respectively, 1000, the IBA of 3000ppm soaks 5-15min in the indolebutyric acid solution, inoculate in the root media as claimed in claim 1, intensity of illumination 2500Lux, illumination 16h/d, under 25 ± 2 ℃ of conditions, begin behind the 20d to take root, after cultivating 125d, rooting rate reaches 90%, get well-grown regeneration plant and place the domestication chamber, 2 weeks of domestication transplant under the high light 20000l ux, use the medium of clear water flush away root earlier, planting in volume ratio is vermiculite: perlite: in the matrix of peat=1: 1: 1, single flower pot coat transparent plastic bag, this bag cut an osculum in per two days, abandoned bag immigration greenhouse plantation after the week.
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| CN2008100633846A CN101336613B (en) | 2008-08-12 | 2008-08-12 | Black bamboo tissue culture medium and tissue culture and rapid propagation method |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2479983C1 (en) * | 2011-10-27 | 2013-04-27 | Федеральное государственное бюджетное образовательное учреждение высшего профессионального образования "Горский государственный аграрный университет" | Method of increasing net reproduction of meristematic potato tubers |
| CN108174786A (en) * | 2018-01-18 | 2018-06-19 | 广西正匠农业科技有限公司 | A kind of yushania method for tissue culture |
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- 2008-08-12 CN CN2008100633846A patent/CN101336613B/en not_active Expired - Fee Related
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2479983C1 (en) * | 2011-10-27 | 2013-04-27 | Федеральное государственное бюджетное образовательное учреждение высшего профессионального образования "Горский государственный аграрный университет" | Method of increasing net reproduction of meristematic potato tubers |
| CN108174786A (en) * | 2018-01-18 | 2018-06-19 | 广西正匠农业科技有限公司 | A kind of yushania method for tissue culture |
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| CN101336613B (en) | 2010-12-29 |
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