CN104920215A - Simplified sugarcane test-tube plantlet rooting and seedling method - Google Patents
Simplified sugarcane test-tube plantlet rooting and seedling method Download PDFInfo
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Abstract
The invention relates to a simplified sugarcane test-tube plantlet rooting and seedling method. The method comprises steps as follows: a sugarcane test-tube plantlet undergone a multiplication culture cycle is transferred to a rooting culture medium and cultured for 24-96 hours in a culture room; the rootless test-tube plantlet is divided, sterilized for 3-6 minutes and then planted in a planting pot containing a culture medium, and after a young leaf of the test-tube plantlet grows, a nursery bed management method is the same with a traditional nursery bed management method. According to the seedling method, the rootless test-tube plantlet replaces a traditional test-tube plantlet with a root for transplantation, autotrophic and heterotrophic rooting replaces traditional heterotrophic rooting, the rooting culture time of the sugarcane test-tube plantlet is shortened from 15 days to 1-4 days, the production efficiency is greatly improved, the production cost is reduced, and large-scale production and popularization of health sugarcane test-tube plantlets are facilitated.
Description
Technical field
The present invention relates to the heterotrophism+photoautotrophy rooting method of a Plants test-tube plantlet, be specifically related to a kind of sugarcane rooting of vitro seedling and seedling-cultivating method of simplification.
Background technology
Plant Tissue Breeding generally will through four-stage, i.e. Initial culture (or setting up aseptic culture system), shoot proliferation cultivation, culture of rootage and acclimatization and transplants.At present, on producing, the sugarcane test-tube plantlet fast breeding technology of large-scale application also includes above four-stage.But this traditional sugarcane test-tube plantlet production process is complicated, and high to culture environment condition, cause the cost of sugarcane tissue culture factorial praluction test-tube plantlet to remain high, can not be accepted by market, limit plantation and the popularization of sugarcane health seedling.Thus, simplifying sugarcane test-tube plantlet production routine, reduce production cost, is solve the most effective way that this difficult problem is promoted in the large-scale production of restriction sugarcane health seedling.
Research shows, in plant tissue culture course, the cost of taking root of test-tube plantlet accounts for 35% ~ 75% of whole test-tube plantlet production cost, therefore simplifies rooting of vitro seedling technology and plant test-tube plantlet production cost can be made significantly to decline.Advance in non-tube rootage technology is the successful advanced group training rooting technique of research in recent years, is the important component part that plant test-tube plantlet simplifies production technology.Advance in non-tube rootage technology is directly cultivated by unrooted test-tube plantlet to take root in nursery, greenhouse, by traditional take root and tame two stages combine, eliminate the traditional program of the in vitro rooting of unrooted test-tube plantlet, taking root by autotrophy replaces traditional heterotrophism to take root, shorten growing-seedling period, save production cost.Meanwhile, the development of plant tissue culture technique is very fast, is studied both at home and abroad, and obtains successfully large quantities of plant Advance in non-tube rootage technologies such as babysbreath, Momordica grosvenori, tree peony, sugarcane, strawberries.
General sugarcane rooting of vitro seedling needs more than 15 days, and test-tube plantlet quality is poor, and production cost is high, seriously constrains production efficiency and reduces productivity effect.Study a kind of scientific and reasonable sugarcane rooting of vitro seedling and seedling-cultivating method, shorten rooting of vitro seedling time and growing-seedling period, to the production capacity and the range of application that improve sugarcane test-tube plantlet, tool is of great significance.
Summary of the invention
The object of the invention is the deficiency overcoming conventional sugarcane rooting of vitro seedling medium and seedling-cultivating method, a kind of sugarcane rooting of vitro seedling and seedling-cultivating method of simplification are provided.
The present invention realizes in the following way:
The sugarcane rooting of vitro seedling of simplification and a seedling-cultivating method, is characterized in that, comprises the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20mg/L+ sucrose 50g/L.The each composition of MS medium and content as shown in table 1.
Table 1 MS medium component and content
(2) root media sterilizing: by root media autoclave sterilization 5 ~ 10min at 121 DEG C.The culture of rootage time of the present invention shortens to 1-4 days by original 14-15 days, is therefore suitably shortened by conventional sterilization time 15min and medium can not be caused to pollute, also can not affect the growth of test-tube plantlet.
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling.The culture of rootage time of the present invention shortens to 1-4 days by original 14-15 days, sugarcane test-tube plantlet not vulnerable to pollution, and sterile working standard can suitably reduce, and cleans superclean bench number of times as reduced.
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 25 ~ 30 DEG C, humidity 65 ~ 80%, intensity of illumination 1000 ~ 2000lux, cultivates 24-96 hour.Sugarcane test-tube plantlet adventive root occurs in theory will through 4 stages, i.e. dedifferentiation stage (after rooting treatment 0-24 hour), induction period (24-96 hour) and differential period (after 96 hours), in differential period root restriction elongation, and break through epidermis, form visible root.Traditional sugarcane rooting of vitro seedling cultivation cycle takes 14-15 days, and namely adventive root completes above-mentioned three growth and development stages, and when visible root all appears in the test-tube plantlet of 80%, can transplanted seedling garden after namely completing culture of rootage.Present invention utilizes sugarcane test-tube plantlet adventive root genesis and development irreversible principle of its process of taking root after the dedifferentiation stage, the unrooted test tube seedling direct transplantation of culture of rootage 1-4 days is carried out autotrophy in nursery and takes root.
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1cm, the test-tube plantlet being highly less than 1cm involves body segmentation in a criminal case by 2 ~ 3, cleans medium, remove Lao Ye with clear water.
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.2 ~ 0.3% liquor potassic permanganate soaking disinfection 3 ~ 6 minutes.
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water.
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, cover sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 2000 ~ 10000lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 2000 ~ 10000lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
Preferably, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1mg/L+ methyl α-naphthyl acetate 0.01mg/L+ sucrose 20g/L.
Preferably, described cultivation matrix is nursery vermiculite and the organic nutrition earth mixtures of volume ratio 1:1, or the rural area soil of weathering, or the yellow mud of volume ratio 1:1 and river sand mixture.
Preferably, the composition of described organic nutrient soil comprises: organic>=30%, humic acid>=5%, N+P
2o
5+ K
2o>=3%.
The invention has the beneficial effects as follows:
1. the sugarcane rooting of vitro seedling time of the present invention foreshortened to 1-4 days from traditional 15 days, test-tube plantlet survival rate is up to more than 90%, test-tube plantlet quality is good, dark green, without yellow leaf, survival rate transplants level of viability to there being root test tube nursery, significantly improves production efficiency and the economic benefit of sugarcane test-tube plantlet.
2. the root media autoclave sterilization time of the present invention shortened to 5 to 10 minutes from 15 minutes, saved high-voltage time, manpower, electric power.
3. aseptic tube of the present invention is easy and simple to handle, is beneficial to saving human and material resources, reduces production cost.
4. the present invention's unrooted test-tube plantlet replaces traditional root test-tube plantlet that has to transplant, traditional heterotrophism is replaced to take root with heterotrophism+autotrophy rooting method, saving culturing room takes up room, improve culturing room utilizes turnover rate, improves output, saves labour and energy input, considerably improves production efficiency.
5. rooting of vitro seedling cultivation matrix of the present invention is price comparatively cheap and the more rich nursery vermiculite of nutrition and organic nutrition earth mixtures, or the rural area soil of weathering, or yellow mud and river sand mixture, replaces expensive MS root media, is beneficial to and reduces costs.
6. rooting of vitro seedling and nursery taming organically combine by the present invention, and the production cycle shortens 12-15 days than conventional method.
7. the present invention is simple to operate, with low cost, good in economic efficiency, has good market prospects.
Embodiment
Below in conjunction with specific embodiment, the invention will be further described, but do not limit the scope of the invention and range of application.
Embodiment 1
The sugarcane rooting of vitro seedling of simplification and a seedling-cultivating method, is characterized in that, comprises the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20mg/L+ sucrose 50g/L;
(2) root media sterilizing: by root media autoclave sterilization 5min at 121 DEG C;
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling;
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 25 DEG C, humidity 80%, and intensity of illumination 1200lux, cultivates 24 hours;
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1cm, the test-tube plantlet being highly less than 1cm involves body segmentation in a criminal case by 2, cleans medium, remove Lao Ye with clear water;
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.3% liquor potassic permanganate soaking disinfection 3 minutes;
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water;
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, covers sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 4000lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 6000lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
Wherein, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1mg/L+ methyl α-naphthyl acetate 0.01mg/L+ sucrose 20g/L; Described cultivation matrix is nursery vermiculite and the organic nutrition earth mixtures of volume ratio 1:1, and the composition of described organic nutrient soil comprises: organic>=30%, humic acid>=5%, N+P
2o
5+ K
2o>=3%.
Plant and add up test-tube plantlet for the 20th day, plant 50 strains/clump, 46 strains of surviving/clump, survival rate is 92%.
Embodiment 2
The sugarcane rooting of vitro seedling of simplification and a seedling-cultivating method, is characterized in that, comprises the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20mg/L+ sucrose 50g/L;
(2) root media sterilizing: by root media autoclave sterilization 10min at 121 DEG C;
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling;
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 28 DEG C, humidity 70%, and intensity of illumination 1000lux, cultivates 48 hours;
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1cm, the test-tube plantlet being highly less than 1cm involves body segmentation in a criminal case by 3, cleans medium, remove Lao Ye with clear water;
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.2% liquor potassic permanganate soaking disinfection 6 minutes;
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water;
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, covers sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 2000lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 2000lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
Wherein, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1mg/L+ methyl α-naphthyl acetate 0.01mg/L+ sucrose 20g/L; Described cultivation matrix is the rural area soil of weathering.
Plant and add up test-tube plantlet for the 20th day, plant 50 strains/clump, 42 strains of surviving/clump, survival rate is 84%.
Embodiment 3
The sugarcane rooting of vitro seedling of simplification and a seedling-cultivating method, is characterized in that, comprises the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20mg/L+ sucrose 50g/L; (2) root media sterilizing: by root media autoclave sterilization 8min at 121 DEG C;
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling;
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 30 DEG C, humidity 75%, and intensity of illumination 2000lux, cultivates 72 hours;
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1cm, the test-tube plantlet being highly less than 1cm involves body segmentation in a criminal case by 3, cleans medium, remove Lao Ye with clear water;
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.3% liquor potassic permanganate soaking disinfection 4 minutes;
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water;
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, covers sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 6000lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 8000lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
Wherein, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1mg/L+ methyl α-naphthyl acetate 0.01mg/L+ sucrose 20g/L; Described cultivation matrix is yellow mud and the river sand mixture of volume ratio 1:1.
Plant and add up test-tube plantlet for the 20th day, plant 50 strains/clump, 43 strains of surviving/clump, survival rate is 86%.
Embodiment 4
The sugarcane rooting of vitro seedling of simplification and a seedling-cultivating method, is characterized in that, comprises the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20mg/L+ sucrose 50g/L; (2) root media sterilizing: by root media autoclave sterilization 6min at 121 DEG C;
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling;
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 26 DEG C, humidity 65%, and intensity of illumination 1500lux, cultivates 96 hours;
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1cm, the test-tube plantlet being highly less than 1cm involves body segmentation in a criminal case by 2, cleans medium, remove Lao Ye with clear water;
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.2% liquor potassic permanganate soaking disinfection 5 minutes;
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water;
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, covers sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 10000lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 10000lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
Wherein, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1mg/L+ methyl α-naphthyl acetate 0.01mg/L+ sucrose 20g/L; Described cultivation matrix is nursery vermiculite and the organic nutrition earth mixtures of volume ratio 1:1, and the composition of described organic nutrient soil comprises: organic>=30%, humic acid>=5%, N+P
2o
5+ K
2o>=3%.
Plant and add up test-tube plantlet for the 20th day, plant 50 strains/clump, 47 strains of surviving/clump, survival rate is 94%.
Comparative example 1
As a comparison, adopt the sugarcane rooting of vitro seedling of conventional sugarcane rooting of vitro seedling culture technique and simplification of the present invention and seedling-cultivating method to cultivate sugarcane test-tube plantlet respectively respectively, and carry out stroke analysis and compare, result is as shown in table 2:
Table 2 distinct methods cultivates the Technical comparing of sugarcane test-tube plantlet
The sugarcane rooting of vitro seedling time was foreshortened to 1-4 days from traditional 15 days by the sugarcane rooting of vitro seedling of a kind of simplification of the present invention and seedling-cultivating method, the root media autoclave sterilization time was foreshortened to 5 ~ 10 minutes from 15 minutes, and simplify the aseptic tube operation of sugarcane test-tube plantlet, significantly shorten the production cycle, improve production efficiency and the economic benefit of sugarcane test-tube plantlet; Traditional root test-tube plantlet that has is replaced to transplant with unrooted test-tube plantlet, taking root with heterotrophism+autotrophy replaces traditional heterotrophism to take root, sugarcane test-tube plantlet quality better, and save culturing room space, labour, energy input, raising culturing room's availability and production capacity, significantly reduce production cost.Therefore, the root media of simplification sugarcane rooting of vitro seedling of the present invention and seedling-cultivating method are beneficial to large-scale production and the popularization of sugarcane health test-tube plantlet.
Claims (4)
1. the sugarcane rooting of vitro seedling simplified and a seedling-cultivating method, is characterized in that, comprise the following steps:
(1) root media is prepared: according to following formulated root media: 1/2MS macroelement+MS trace element+MS molysite+MS organic matter+methyl α-naphthyl acetate 5 ~ 20 mg/L+ sucrose 50 g/L;
(2) root media sterilizing: by root media autoclave sterilization 5 ~ 10 min at 121 DEG C;
(3) aseptic tube: in desinfection chamber or superclean bench, the sugarcane test-tube plantlet completing the Multiplying culture cycle is transferred in the root media of cooling;
(4) culture of rootage: the sugarcane test-tube plantlet in root media is moved into culturing room, controlling culturing room's environment is temperature 25 ~ 30 DEG C, humidity 65 ~ 80%, intensity of illumination 1000 ~ 2000 lux, cultivates 24-96 hour;
(5) split test-tube plantlet: after culture of rootage completes, from bottle, take out test-tube plantlet, be highly greater than the test-tube plantlet individual plant segmentation of 1 cm, the test-tube plantlet being highly less than 1 cm involves body segmentation in a criminal case by 2 ~ 3, cleans medium, remove Lao Ye with clear water;
(6) sterilize: the test-tube plantlet after segmentation is clean with 0.2 ~ 0.3% liquor potassic permanganate soaking disinfection 3 ~ 6 minutes;
(7) plant: the test-tube plantlet after sterilization is divided into individual plant or little Cong plants in the planting pot being placed with cultivation matrix, waters sufficient normal root water;
(8) cultivating and managing: the planting pot with test-tube plantlet is moved into the seedling cultivation greenhouse automatically regulating temperature humidity, covers sunshade net, regulate seedling cultivation greenhouse environment: in 10 days after plantation, chamber humidity controls more than 80%, intensity of illumination 2000 ~ 10000 lux; Plant after 10 days, chamber humidity controls more than 70%, intensity of illumination 2000 ~ 10000 lux; After sugarcane test-tube plantlet when 90% grows young leaves, namely open sunshade net, it is 70% that chamber humidity controls, every 30 minutes trickles 30 seconds, until test-tube plantlet goes out garden; After test-tube plantlet grows young leaves, start applying liquid fertilizer and weeding, nursery bed management is identical with conventional sugarcane test-tube plantlet seedling-cultivating method.
2. the seedling-cultivating method of simplification sugarcane rooting of vitro seedling according to claim 1, it is characterized in that, its medium of described Multiplying culture is MS+6-benzyl aminoadenine 1 mg/L+methyl α-naphthyl acetate 0.01 mg/L+sucrose 20 g/L.
3. the seedling-cultivating method of simplification sugarcane rooting of vitro seedling according to claim 1 and 2, it is characterized in that, described cultivation matrix is nursery vermiculite and the organic nutrition earth mixtures of volume ratio 1:1, or the rural area soil of weathering, or the yellow mud of volume ratio 1:1 and river sand mixture.
4. the seedling-cultivating method of simplification sugarcane rooting of vitro seedling according to claim 3, it is characterized in that, the composition of described organic nutrient soil comprises: organic>=30%, humic acid>=5%, N+P
2o
5+ K
2o>=3%.
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| CN109197504A (en) * | 2018-08-29 | 2019-01-15 | 广西壮族自治区农业科学院 | A kind of sugarcane test tube seedling seedling nutritious soil and its preparation method and application |
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