CN101507415A - In-vitro culture method of antlerpilose grass - Google Patents
In-vitro culture method of antlerpilose grass Download PDFInfo
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- CN101507415A CN101507415A CNA2009100969550A CN200910096955A CN101507415A CN 101507415 A CN101507415 A CN 101507415A CN A2009100969550 A CNA2009100969550 A CN A2009100969550A CN 200910096955 A CN200910096955 A CN 200910096955A CN 101507415 A CN101507415 A CN 101507415A
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Abstract
The invention discloses an isolated culture method for Monochasma Savatieri Franch.ex Maxim, which comprises the following steps in turn: 1) washing a burgeon of the Monochasma Savatieri Franch.ex Maxim; 2) after conventional disinfection, cutting the burgeon into a small segment of burgeon I and inoculating the small segment of burgeon I to an adventitious bud culture medium for culture; 3) cutting an adventitious bud growing out on the small segment of burgeon I, and inoculating the obtained small segment of burgeon II to a multiplication culture medium for culture; 4) cutting an adventitious bud growing out on the small segment of burgeon II, and inoculating an obtained small segment of burgeon III to a multiplication culture medium for culture; 5) cutting an adventitious bud growing out on the small segment of burgeon III, and inoculating an obtained seedling III into a rooting culture medium; and 6) obtaining the seedling capable of being planted out of a bottle until the seedling III grows out at least three roots greater than 2 cm. With the method, people can obtain a great number of seedlings of the Monochasma Savatieri Franch.ex Maxim.
Description
Technical field
The present invention relates to a kind of cultured in vitro method of wool savatier monochasma herb.
Background technology
The wool savatier monochasma herb belongs to the Scrophulariaceae savatier monochasma herb and belongs to, and effect such as have clearing heat and detoxicating, the promoting the circulation of qi of dispeling the wind, cooling blood and hemostasis, eliminate the phlegm is used for the treatment of flu, chronic bronchitis, pneumonia etc., and medical value is very high; It is distributed in Jiangsu, Zhejiang one band, the present domestic artificial cultivation of also not realizing, and therefore the quantity of wild wool savatier monochasma herb is fewer and feweri.
Summary of the invention
The technical problem to be solved in the present invention provides a kind of cultured in vitro method of cotton wool savatier monochasma herb, adopts this method can obtain the seedling of a large amount of wool savatier monochasma herbs.
In order to solve the problems of the technologies described above, the invention provides a kind of in vitro culture method of cotton wool savatier monochasma herb, may further comprise the steps successively:
1), the wool savatier monochasma herb tender shoots of getting no scab, robust growth carries out the flowing water flushing;
2), the tender shoots after the flushing of above-mentioned flowing water is sterilized through routine, be cut into the long segment tender shoots I of 0.8～1.2cm then, above-mentioned segment tender shoots I be inoculated on the evoking adventive bud medium cultivate; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
3), when treating that the indefinite bud that grows on the segment tender shoots I grows to the high seedling I of 2.0～5.0cm, extract seedling I; This seedling I is cut into the long segment tender shoots II of 0.8～1.2cm, above-mentioned segment tender shoots II is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
4), II when treating that the indefinite bud that grows on the segment tender shoots II grows to the high seedling of 2.0～5.0cm, extract seedling II; This seedling II is cut into the long segment tender shoots III of 0.8～1.2cm, above-mentioned segment tender shoots III is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
5), when treating that the indefinite bud that grows on the segment tender shoots III grows to the high seedling III of 3.0～5.0cm, extract seedling III; III is inoculated in the root media with this seedling;
6), when treating that above-mentioned seedling III grows at least 3 roots greater than 2cm, but the seedling of bottle outlet plantation.
Improvement as the in vitro culture method of cotton wool savatier monochasma herb of the present invention: the evoking adventive bud medium step 2) is: MS minimal medium+BA0.5～1.0mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
The preparation method of evoking adventive bud medium is specific as follows: as the basis, add 6-benzyladenine (BA), sugar and agar with the MS minimal medium respectively, evenly mix, utilizing the KOH of 1mol/L or the HCl adjusting pH of 1mol/L is 5.5～6.0; Add 0.5～1.0mg BA, 20～30g sugar and 5～9g agar in the MS minimal medium of every 1L.
Further improvement as the in vitro culture method of cotton wool savatier monochasma herb of the present invention: the proliferated culture medium in step 3) and the step 4) is: MS minimal medium+BA 0.3～0.5mg/l+NAA 0.2～0.5mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
The preparation method of proliferated culture medium is specific as follows: as the basis, add 6-benzyladenine (BA) and methyl (NAA), sugar and agar with the MS minimal medium respectively, evenly mix, utilizing the KOH of 1mol/L or the HCl adjusting pH of 1mol/L is 5.5～6.0; The MS minimal medium of every 1L adds 0.3～0.5mg BA, 0.2～0.5mg NAA, 20～30g sugar and 5～9g agar.
Further improvement as the in vitro culture method of cotton wool savatier monochasma herb of the present invention: the root media in the step 5) is: 1/2MS minimal medium+NAA0.2～0.5mg/l+ sugar 30g/l+ agar 5～9g/l+ active carbon 2g/l, pH is 5.5～6.0.
The preparation method of root media is specific as follows: with 1/2MS minimal medium (content of all substances is half of MS minimal medium in the ie in solution) as the basis, add methyl (NAA), sugar, agar and active carbon respectively, evenly mix, utilizing the KOH of 1mol/L or the HCl adjusting pH of 1mol/L is 5.5～6.0; The 1/2MS minimal medium of every 1L adds 0.2～0.5mg NAA, 30g sugar, 5～9g agar and 2g active carbon.
Further improvement as the in vitro culture method of cotton wool savatier monochasma herb of the present invention: step 1) is: wool savatier monochasma herb tender shoots is put into beaker, running water flushing 1～2 hour.
The in vitro culture method of wool savatier monochasma herb of the present invention belongs to a kind of wool savatier monochasma herb method for tissue culture of breeding fast of inducing.According to the totipotent principle of cell in the Plant Tissue Breeding, can produce the high quality seedling that a large amount of genetic backgrounds are identical, growing way is consistent (seed) at short notice, and rely on that the laboratory can realize the anniversary high quality seedling is provided for a long time.In the method for the invention, the wool savatier monochasma herb tender shoots of stalwartness is sterilized earlier, induce by suitable inducing culture again to obtain a certain amount of aseptic seedling.Adopt method of the present invention, but the general seedling that only needs can obtain in 60～80 days the bottle outlet plantation; Therefore the reproduction coefficient in the wool savatier monochasma herb test-tube plantlet anniversary is 3 in theory
10Doubly.Wool savatier monochasma herb in vitro culture method of the present invention is a kind of factor affecting such as season that is not subjected to, and the method for high-quality wool savatier monochasma herb seedling can be provided efficiently, fast, can quicken improved variety popularization speed, improves field kind plantation output.
The seedling of gained of the present invention can adopt following method to plant: shift out blake bottle having at least 3 seedlings of being longer than the 2cm root, clean its root medium, be transplanted in the matrix (is that peat, perlite and the vermiculite of 3:1:1 formed by volume ratio), in the climatic cabinate indoor cultivation, condition of culture: illumination in 12 hours, intensity of illumination 15～25 μ mol m
-2.s
-1, temperature is 23～25 ℃; 12 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed; Humidity 85%～90%.After 3～5 weeks, make into: intensity of illumination is 30～40 μ mol m
-2.s
-1, all the other conditions are the same.After 2 months, survival rate reaches more than 60%.
And the identical radical of natural income is cultivated under above-mentioned identical environment with the long wild[l of root, and survival rate only is about 10% after 2 months.
Embodiment
The method of embodiment 1, a kind of wool savatier monochasma herb cultured in vitro, carry out following steps successively:
1), the wool savatier monochasma herb tender shoots of getting no scab, robust growth is put into beaker, running water flushing 1～2 hour;
2) (be 0.1%w/v HgCl through the routine sterilization, with above-mentioned tender shoots
2Handle after 6～10 minutes, aseptic water washing 5～8 times blots with aseptic filter paper then) after, be cut into the segment tender shoots I about 1cm, be inoculated on the evoking adventive bud medium and cultivate; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
The evoking adventive bud medium is: MS minimal medium+BA0.5～1.0mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
3), when treating that the indefinite bud that grows on the segment tender shoots I grows to the high seedling I of 2.0～5.0cm, extract seedling I; This seedling I is cut into segment tender shoots II about 1cm, above-mentioned segment tender shoots II is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
Proliferated culture medium is: MS minimal medium+BA 0.3～0.5mg/l+NAA 0.2～0.5mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
4), II when treating that the indefinite bud that grows on the segment tender shoots II grows to the high seedling of 2.0～5.0cm, extract seedling II; This seedling II is cut into the long segment tender shoots III of 0.8～1.2cm, above-mentioned segment tender shoots III is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
Proliferated culture medium is with the proliferated culture medium of step 3).
5), when treating that the indefinite bud that grows on the segment tender shoots III grows to the high seedling III of 3.0～5.0cm, extract seedling III; III is inoculated in the root media with this seedling; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
Root media is: 1/2MS minimal medium+NAA 0.2～0.5mg/l+ sugar 30g/l+ agar 5～9g/l+ active carbon 2g/l, pH is 5.5～6.0.
6), when treating that above-mentioned seedling III grows at least 3 roots greater than 2cm, but the seedling of bottle outlet plantation.
According to said method, but only needed 60～80 days can obtain the seedling (test-tube plantlet) that bottle outlet is planted; Therefore adopt method of the present invention can obtain a large amount of test-tube plantlets for a long time.
At last, it is also to be noted that what more than enumerate only is a specific embodiment of the present invention.Obviously, the invention is not restricted to above embodiment, many distortion can also be arranged.All distortion that those of ordinary skill in the art can directly derive or associate from content disclosed by the invention all should be thought protection scope of the present invention.
Claims (5)
1, a kind of in vitro culture method of cotton wool savatier monochasma herb is characterized in that may further comprise the steps successively:
1), the wool savatier monochasma herb tender shoots of getting no scab, robust growth carries out the flowing water flushing;
2), the tender shoots after the flushing of above-mentioned flowing water is sterilized through routine, be cut into the long segment tender shoots I of 0.8～1.2cm then, above-mentioned segment tender shoots I be inoculated on the evoking adventive bud medium cultivate; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
3), when treating that the indefinite bud that grows on the segment tender shoots I grows to the high seedling I of 2.0～5.0cm, extract seedling I; This seedling I is cut into the long segment tender shoots II of 0.8～1.2cm, above-mentioned segment tender shoots II is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
4), II when treating that the indefinite bud that grows on the segment tender shoots II grows to the high seedling of 2.0～5.0cm, extract seedling II; This seedling II is cut into the long segment tender shoots III of 0.8～1.2cm, above-mentioned segment tender shoots III is inoculated on the proliferated culture medium cultivates; Condition of culture is: illumination in 16 hours, intensity of illumination 10～30 μ mol m
-2.s
-1, temperature is 26～28 ℃; 8 hours dark cultivations, temperature is 20～22 ℃; Above-mentioned illumination and dark the cultivation are hocketed;
5), when treating that the indefinite bud that grows on the segment tender shoots III grows to the high seedling III of 3.0～5.0cm, extract seedling III; III is inoculated in the root media with this seedling;
6), when treating that above-mentioned seedling III grows at least 3 roots greater than 2cm, but the seedling of bottle outlet plantation.
2, the in vitro culture method of cotton wool savatier monochasma herb according to claim 1 is characterized in that: step 2) in the evoking adventive bud medium be: MS minimal medium+BA0.5～1.0mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
3, the in vitro culture method of cotton wool savatier monochasma herb according to claim 2, it is characterized in that: the proliferated culture medium in step 3) and the step 4) is: MS minimal medium+BA 0.3～0.5mg/l+NAA 0.2～0.5mg/l+ sugar 20～30g/l+ agar 5～9g/l, pH is 5.5～6.0.
4, the in vitro culture method of cotton wool savatier monochasma herb according to claim 3 is characterized in that: the root media in the step 5) is: 1/2MS minimal medium+NAA 0.2～0.5mg/l+ sugar 30g/l+ agar 5～9g/l+ active carbon 2g/l, pH is 5.5～6.0.
5, the in vitro culture method of cotton wool savatier monochasma herb according to claim 4 is characterized in that: described step 1) is: wool savatier monochasma herb tender shoots is put into beaker, running water flushing 1～2 hour.
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| CN2009100969550A CN101507415B (en) | 2009-03-26 | 2009-03-26 | In-vitro culture method of antlerpilose grass |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102498781A (en) * | 2011-10-09 | 2012-06-20 | 中国科学院华南植物园 | Method for Monochasma sheareri Maxim.ex Franch.et Savat seed germination and seedling breeding |
| CN104770300A (en) * | 2015-04-17 | 2015-07-15 | 浙江省中药研究所有限公司 | Method for quickly breeding monochasma Savatieri Franch.ex Maxim seedlings |
| CN104782367A (en) * | 2015-04-15 | 2015-07-22 | 万邦德（湖南）天然药物有限公司 | Artificial cultivation method for savatier monochasma herb |
| CN109757313A (en) * | 2019-03-12 | 2019-05-17 | 浙江省中药研究所有限公司 | A kind of method for transplanting of wool savatier monochasma herb |
| CN110859130A (en) * | 2019-12-04 | 2020-03-06 | 中国林业科学研究院亚热带林业实验中心 | Method for rooting tissue culture seedlings of medicinal plant of cornua cervi pantotrichum outside bottle |
| CN111386981A (en) * | 2020-04-10 | 2020-07-10 | 惠州市九惠制药股份有限公司 | Method for planting savatier saururus in pot |
| CN113692928A (en) * | 2021-04-02 | 2021-11-26 | 宜春市炳晨农业科技发展有限公司 | Breeding method of savatier monochasma herb |
| CN113692927A (en) * | 2021-04-06 | 2021-11-26 | 宜春市炳晨农业科技发展有限公司 | Artificial cultivation method of savatier monochasma |
| CN113812337A (en) * | 2021-04-02 | 2021-12-21 | 宜春市炳晨农业科技发展有限公司 | Hydroponic rooting method for savatier monochasma |
-
2009
- 2009-03-26 CN CN2009100969550A patent/CN101507415B/en not_active Expired - Fee Related
Cited By (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102498781A (en) * | 2011-10-09 | 2012-06-20 | 中国科学院华南植物园 | Method for Monochasma sheareri Maxim.ex Franch.et Savat seed germination and seedling breeding |
| CN104782367A (en) * | 2015-04-15 | 2015-07-22 | 万邦德（湖南）天然药物有限公司 | Artificial cultivation method for savatier monochasma herb |
| CN104770300A (en) * | 2015-04-17 | 2015-07-15 | 浙江省中药研究所有限公司 | Method for quickly breeding monochasma Savatieri Franch.ex Maxim seedlings |
| CN109757313A (en) * | 2019-03-12 | 2019-05-17 | 浙江省中药研究所有限公司 | A kind of method for transplanting of wool savatier monochasma herb |
| CN109757313B (en) * | 2019-03-12 | 2023-09-26 | 浙江省中药研究所有限公司 | Transplanting method of hairy antler grass |
| CN110859130B (en) * | 2019-12-04 | 2022-02-01 | 中国林业科学研究院亚热带林业实验中心 | Method for rooting tissue culture seedlings of medicinal plant of cornua cervi pantotrichum outside bottle |
| CN110859130A (en) * | 2019-12-04 | 2020-03-06 | 中国林业科学研究院亚热带林业实验中心 | Method for rooting tissue culture seedlings of medicinal plant of cornua cervi pantotrichum outside bottle |
| CN111386981A (en) * | 2020-04-10 | 2020-07-10 | 惠州市九惠制药股份有限公司 | Method for planting savatier saururus in pot |
| CN113692928A (en) * | 2021-04-02 | 2021-11-26 | 宜春市炳晨农业科技发展有限公司 | Breeding method of savatier monochasma herb |
| CN113812337A (en) * | 2021-04-02 | 2021-12-21 | 宜春市炳晨农业科技发展有限公司 | Hydroponic rooting method for savatier monochasma |
| CN113692928B (en) * | 2021-04-02 | 2023-02-14 | 宜春市炳晨农业科技发展有限公司 | Breeding method of savatier monochasma herb |
| CN113692927A (en) * | 2021-04-06 | 2021-11-26 | 宜春市炳晨农业科技发展有限公司 | Artificial cultivation method of savatier monochasma |
| CN113692927B (en) * | 2021-04-06 | 2023-02-14 | 宜春市炳晨农业科技发展有限公司 | Artificial cultivation method of savatier monochasma |
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| CN101507415B (en) | 2011-01-05 |
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