CN102144569B - Method for rapidly propagating root and rhizome of Divaricate Gynura - Google Patents
Method for rapidly propagating root and rhizome of Divaricate Gynura Download PDFInfo
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- CN102144569B CN102144569B CN2011101155222A CN201110115522A CN102144569B CN 102144569 B CN102144569 B CN 102144569B CN 2011101155222 A CN2011101155222 A CN 2011101155222A CN 201110115522 A CN201110115522 A CN 201110115522A CN 102144569 B CN102144569 B CN 102144569B
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Abstract
The invention discloses a method for rapidly propagating root and rhizome of Divaricate Gynura. The method comprises the following steps of: inducing culture, namely inoculating in a murashige and skoog (MS) start culture medium which does not contain any hormone so as to induce the generation of axillary buds; and proliferation and rooting culture, namely cutting a start seedling into single stem segments with buds, and inoculating in a culture medium, namely 1/2MS culture medium plus 0.2mg/L naphthylacetic acid (NAA) plus 20g/L sucrose plus 6g/L agar for synchronous proliferation and rooting, wherein culture conditions are both that the temperature is 25+/-1 DEG C, the illumination intensity is 2000lx, and the illumination time is 16h/d. By the method, high-quality seedlings of the root and rhizome of Divaricate Gynura, which have pure seed character and are healthy and neat, can be rapidly, efficiently and massively produced.
Description
Technical field
The invention belongs to the Plant Tissue Breeding field, be specifically related to a kind of quick-breeding method of Radix et Rhizoma Gynurae divaricatae.
Background technology
Radix et Rhizoma Gynurae divaricatae (Gynura divaricata (L.) DC.) belongs to composite family pseudo-ginseng platymiscium; Have another name called Bai Dongfeng, white chessman grass, beautiful loquat, asiatic toddalia root-bark, thick musculus cutaneus, chicken dish, big fertile ox, white kind of amaranth, Bai Hongcai, rich and honour dish, Gynura divaricata etc., be distributed in Taiwan to south China, a southwestern band.Its root, cauline leaf all can be used as medicine; Its root nature and flavor are sweet, cool, ability clearing heat and cooling blood, eliminating stasis to subdue swelling; Its stem, the leaf salty little suffering, cold of distinguishing the flavor of, poisonous, have heat-clearing, Shujin, hemostasis, the effect of eliminating the phlegm.On Fujian and other places, among the people with its cauline leaf make tea drinking-water take, be used to treat diabetes etc., curative effect is fine.Radix et Rhizoma Gynurae divaricatae is rich in flavonoids and glycosides compound (Huang Qi etc., the mensuration of Gynura divaricata general flavone content, Fujian Normal University's journal, 2006,22 (4): 118-120; Lee is pretty etc., the comparison of rich and honour dish stem and different development stage leaf flavonoids content, Hunan forestry science and technology, 2007,34 (3): 13-14; Hu Yong etc., the chemical composition of Radix et Rhizoma Gynurae divaricatae acrial part, Chinese natural drug, 2006,4 (2): 156-158; Li Limei etc., the Radix et Rhizoma Gynurae divaricatae chemical constitution study, the time precious traditional Chinese medical science traditional Chinese medicines, 2008; 19 (1): 118-119), polysaccharose (Jiang Manhua etc., the extraction purifying and the Determination on content of Radix et Rhizoma Gynurae divaricatae polysaccharide, the time precious traditional Chinese medical science traditional Chinese medicines; 2008,19 (9): 2147-2149), alcohol compound and derivative (Hu Yong etc., the chemical composition of Radix et Rhizoma Gynurae divaricatae acrial part; China's natural drug, 2006,4 (2): 156-158; Li Limei etc., the Radix et Rhizoma Gynurae divaricatae chemical constitution study, the time precious traditional Chinese medical science traditional Chinese medicines; 2008,19 (1): 118-119) etc., be important Traditional health care type vegetables of China and Chinese herbal medicine resource (Li Yanlin etc.; The present Research and the prospect of the rich and honour dish of medicinal plant, Chinese wild plant resource, 2011; 30 (1): 10-13,19).
Rich and honour dish is planted at me and mainly is distributed in south, and it receives the requirement of growing environment, and its righttest growing environment temperature is 20-30 ℃, is lower than 15 ℃ of cauline leaf poor growths; Become strain can restrain oneself 40 ℃ high temperature, 3 ℃ low temperature, freeze to death (Zheng Hua etc., rich and honour dish artificial cultivation technique of acrial part in the time of-2 ℃; China's Vegetable, 2004, (4): 54-55); The plant of often gathering simultaneously seldom blossoms and has seeds, (He Dongfang, the rich and honour dish high-yield high-grade culture technique of not setting seeds of only blooming in Delta of the Pearl River area, south, China Guangdong in addition; The Changjiang river vegetables, 2003, (11): 29-33; Zhao Shile etc., rich and honour dish and cultivation thereof, extraordinary economic animals and plants, 2002, (6): 36-39).This reproductive characteristic has brought certain difficulty for the germ plasm resource storage and transport of Radix et Rhizoma Gynurae divaricatae, but the conventional artificial cottage propagation is restricting the implant mass of Radix et Rhizoma Gynurae divaricatae.Therefore (Lee is pretty, the research of rich and honour dish flavones content and cultured in vitro, Agricultural University Of Hunan's Master's thesis, 2007,6 to have set up the tissue culturing system of Radix et Rhizoma Gynurae divaricatae through Radix et Rhizoma Gynurae divaricatae regenerating system structure, adventitious organogenesis, lateral bud clump bud approach such as induce; Lee is pretty etc., the foundation of rich and honour dish cultured in vitro system, Changjiang University's journal (natural science edition), 2009,6 (2): 56-60; Lee is pretty etc., the organogenetic research of rich and honour dish, Hunan agricultural science, 2009, (6): 4-6; Huang Lili etc., the tissue culture of Gynura divaricata and breeding fast, Plant Physiology Communications, 46 (4): 377-378).
Above-mentioned 4 kinds of methods produce callus with Radix et Rhizoma Gynurae divaricatae stem section or blade, neomorph takes place then form plant, the formation whole plant of taking root at last; Perhaps form plant, but reproduction coefficient is lower with the lateral bud inducing culture.
Summary of the invention
The object of the present invention is to provide a cover to utilize the biotechnology batch production to produce the method for the seedling of Radix et Rhizoma Gynurae divaricatae nursery stock; Overcome the defective that above-mentioned existing propagation technique or method exist, kind of property is pure, healthy, the high-quality Radix et Rhizoma Gynurae divaricatae seedling of neat and consistent so that produce fast, efficiently, on a large scale.
The objective of the invention is to realize in the following manner.
A kind of quick-breeding method of Radix et Rhizoma Gynurae divaricatae may further comprise the steps:
(1) the drawing materials and sterilizing of explant: choose young sprout Radix et Rhizoma Gynurae divaricatae stem section annual or that give birth to then, cut off blade and stay petiole, clean, sterilize, be cut into the 0.5-1.0cm stem with bud with 70% alcohol and 2% clorox;
(2) inducing culture: the stem section band bud explant that disinfects is seeded in the MS that does not contain any hormone starts medium; Induce axillalry bud to produce, condition of culture is 25 ℃ ± 1 ℃, and intensity of illumination is 2000lx; Light application time is 16h/d, obtains the high startup seedling of 5-8cm after 20 days;
(3) propagation and culture of rootage: will start seedling and be cut into single stem with bud; Being seeded in medium MS+IBA 0.3mg/L+IAA 0.3mg/L+ sucrose 20g/L+ agar 6g/L breeds simultaneously and takes root; Condition of culture is 25 ℃ ± 1 ℃, and intensity of illumination is 2000lx, and light application time is 16h/d;
(4) refining seedling and transplanting: the seedling after will taking root carries out acclimatization and transplants.
The annual or annual shoot tip in the said step (1) derives from the young sprout that spring, Radix et Rhizoma Gynurae divaricatae was sprouted, any cuttage in season branch, old young sprout, seed germination bud or the group training transplanted seedling of pinching and being sprouted.
Said step (1) cleaning and sterilizing process is specific as follows: earlier explant is added detergent with running water and clean; Again with running water flowing water flushing 1 hour; Through 70% alcohol disinfecting 15s and 2% clorox sterilization 12min, subsequent use behind the aseptic water washing 5 times in superclean bench.
The every 4-6 of propagation system week subculture once in the said step (3).On medium MS+IBA 0.3mg/L+IAA 0.3mg/L+ sucrose 20g/L+ agar 6g/L, after 2 weeks, the explant base portion produces the callus of a small amount of white at enrichment culture, and axillalry bud begins to sprout simultaneously, takes root; Cultivate the complete seedling about 3 weeks back formation 3cm;
Said step (4) specifically comprises with lower refining seedling and transplanting process: the container of the seedling of will taking root is directly transferred to normal room and was placed 2 days, shifts out with the tweezers seedling of will taking root then, under running water, washes and removes medium; Be transplanted on the seedbed of refining seedling matrix, water permeablely, and build the little shed of plastics; Guarantee the interior humidity of canopy more than 80%, in plastic tunnel 15-30 ℃ cultivation, two all left and right sides seedlings adapt to culture environment gradually; Can slowly remove little shed, carry out normal rich water quality management.
Described refining seedling matrix is cotton seed hulls (offcuts after the Edible Fungi): river sand: perlitic weight ratio is 3: 1: 1 a mixture.
This invention has the following advantages: (1) explant collection aspect, and pollution rate is low, used disinfectant non-environmental-pollution; (2) enrichment culture obtains aseptic seedling through aseptic seedling axillalry bud approach, and reproduction coefficient is high; (3) enrichment culture is cultivated through one step of axillalry bud approach and is obtained whole plant, and cultivation cycle is short; (4) rooting rate and planting survival rates are high, and genetic stability is good.
The method can realize breeding fast Radix et Rhizoma Gynurae divaricatae, for the large-scale planting Radix et Rhizoma Gynurae divaricatae provides high quality seedling; Especially can cultivate in a step and obtain whole plant fast, plant is exponential propagation mode to be increased, and every 4-6 week repeats subculture once, and transplanting survival rate reaches more than 95%.Can realize the purpose of breeding fast and implant mass at short notice through the Plant Tissue Breeding plant; Breed seedling and do not receive the influence of extraneous poor environment, this method is being widely used aspect the rescue of the expansion plantation of rare flowers, rare traditional Chinese medicine and endangered species.
Description of drawings
Fig. 1 is each the Radix et Rhizoma Gynurae divaricatae plant that is obtained by tender tip stem section in the inventive method in stage, and wherein: the startup of A. stem section is cultivated; B. breed and culture of rootage; C. take root transplantation of seedlings to the seedbed (after 70 days).
Embodiment
Following examples are intended to further specify the present invention, and can not limit the present invention.
Embodiment 1
1) the drawing materials and sterilizing of explant: the tender tip of choosing about 20 days is made explant; Cut off blade (reservation petiole); Clean with an amount of detergent earlier, again with running water flowing water flushing 1 hour, warp 70% alcohol disinfecting 15s and 2% clorox sterilization 12min in superclean bench; Be placed in the superclean bench behind the aseptic water washing 5 times, it is subsequent use to be cut into the 0.5-1.0cm stem with bud.
2) inducing culture: in superclean bench, the stem section band bud explant that disinfects is seeded in the MS that does not contain any hormone and starts medium; Induce axillalry bud to produce; Condition of culture is 25 ℃ ± 1 ℃; Intensity of illumination is about 2000lx, and the photoperiod is 16h/8h, obtains the high startup seedling of 5-8cm after about 20 days; (Fig. 1, A)
3) propagation and culture of rootage: terminal bud and stem with bud to start seedling are explant; Be seeded in root media MS+IBA0.3mg/L+IAA0.3mg/L+ sucrose 20g/L+ agar 6g/L and carry out root induction; (the Fig. 1 of can taking root in 9 days; B), the many and moderate length of root quantity is transplanted after can directly refining seedling.Plant is exponential propagation mode to be increased, and the coefficient of taking root reaches 100%.
4) refining seedling and transplanting: the container of the seedling of will taking root is directly transferred to normal room and was placed 2 days, shifts out with the tweezers seedling of will taking root then, under running water, washes and removes medium; Be transplanted on the seedbed of refining seedling matrix, water permeablely, and build the little shed of plastics; Guarantee the interior humidity of canopy more than 80%, with 25 ℃ of cultivations of plastic tunnel, two all left and right sides seedlings adapt to culture environment gradually; Can progressively remove little shed, carry out normal rich water quality management.Plant and the seedling indifference of become living, keep fully female parent hereditary feature (Fig. 1, C).Survival rate reaches more than 95%.
Claims (4)
1. the quick-breeding method of a Radix et Rhizoma Gynurae divaricatae is characterized in that, may further comprise the steps:
(1) the drawing materials and sterilizing of explant: choose young sprout Radix et Rhizoma Gynurae divaricatae stem section annual or that give birth to then, cut off blade and stay petiole, clean, sterilize, be cut into the 0.5-1.0cm stem with bud with 70% alcohol and 2% clorox;
(2) inducing culture: the stem section band bud explant that disinfects is seeded in the MS that does not contain any hormone starts medium; Induce axillalry bud to produce, condition of culture is 25 ℃ ± 1 ℃, and intensity of illumination is 2000lx; Light application time is 16h/d, obtains the high startup seedling of 5-8cm after 20 days;
(3) propagation and culture of rootage: will start seedling and be cut into single stem with bud; Being seeded in medium MS+IBA 0.3mg/L+IAA 0.3mg/L+ sucrose 20g/L+ agar 6g/L breeds simultaneously and takes root; Condition of culture is 25 ℃ ± 1 ℃, and intensity of illumination is 2000lx, and light application time is 16h/d;
(4) refining seedling and transplanting: the seedling after will taking root carries out acclimatization and transplants.
2. method according to claim 1 is characterized in that,
The annual or annual shoot tip in the said step (1) derives from the young sprout that spring, Radix et Rhizoma Gynurae divaricatae was sprouted, any cuttage in season branch, old young sprout, seed germination bud or the group training transplanted seedling of pinching and being sprouted.
3. method according to claim 1 is characterized in that,
Said step (1) cleaning and sterilizing process is specific as follows: earlier explant is added detergent with running water and clean; Again with running water flowing water flushing 1 hour; Through 70% alcohol disinfecting 15s and 2% clorox sterilization 12min, subsequent use behind the aseptic water washing 5 times in superclean bench.
4. method according to claim 1 is characterized in that,
Said step (4) specifically comprises with lower refining seedling and transplanting process: the container of the seedling of will taking root is directly transferred to normal room and was placed 2 days, shifts out with the tweezers seedling of will taking root then, under running water, washes and removes medium; Be transplanted on the seedbed of refining seedling matrix, water permeablely, and build the little shed of plastics; Guarantee that the humidity in the canopy is not less than 80%, in plastic tunnel 15-30 ℃ cultivation, after two all seedlings adapt to culture environment; Remove little shed, carry out normal rich water quality management.
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| CN102972264A (en) * | 2012-11-16 | 2013-03-20 | 云南农业大学 | Radix notoginseng culture medium prepared by using zeolite, perlite and organic fertilizer |
| CN104381105B (en) * | 2014-10-29 | 2017-09-12 | 德州学院 | A kind of method of Radix et Rhizoma Gynurae divaricatae Water culture Rapid Rooting |
| CN104686340A (en) * | 2015-02-22 | 2015-06-10 | 杨业容 | Method for establishing cell suspension culture system of gynura divaricate (L.) DC. |
| CN107853174A (en) * | 2017-10-31 | 2018-03-30 | 安徽新华学院 | A kind of method for improving angelica keiskei koidz lateral bud survival rate |
| CN107873412A (en) * | 2017-12-06 | 2018-04-06 | 广西沙田仙人滩农业投资有限公司 | A kind of implantation methods of Bulbilus boussingaultiae pseudobaselloidis good quality and high output |
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