CN105210831A - The breeding method of blbizzia falcata tissue cultures explant - Google Patents
The breeding method of blbizzia falcata tissue cultures explant Download PDFInfo
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- CN105210831A CN105210831A CN201510683550.2A CN201510683550A CN105210831A CN 105210831 A CN105210831 A CN 105210831A CN 201510683550 A CN201510683550 A CN 201510683550A CN 105210831 A CN105210831 A CN 105210831A
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- seed
- sterile water
- tissue cultures
- breeding method
- blbizzia falcata
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G31/00—Soilless cultivation, e.g. hydroponics
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01C—PLANTING; SOWING; FERTILISING
- A01C1/00—Apparatus, or methods of use thereof, for testing or treating seed, roots, or the like, prior to sowing or planting
Abstract
The present invention relates to a kind of breeding method of blbizzia falcata tissue cultures explant, comprise the steps such as seed disinfection, seed soaking, vernalization, inoculation, nursery.By aseptic nursery flow process of the present invention, pollution rate can be controlled within 5%; Pregermination device provided by the invention provides the aseptic seedling raising environment that temperature and humidity is suitable for, is conducive to young root growth; Raise seedling culture medium in the method can promote that seedling is healthy and strong, grow fast; Operation is simple for the method, the cycle is short, it is low to pollute, output is high, can obtain more than 95% aseptic healthy and strong explant, can meet the needs of blbizzia falcata tissue-culturing quick-propagation Establishing in 10 days.
Description
Technical field
The present invention relates to biological technical field, particularly relate to a kind of breeding method of blbizzia falcata tissue cultures explant.
Background technology
Blbizzia falcata (Paraserianthesfalcataria (L.) Nielsen) is Mimosaceae (Mimosaceae), blbizzia falcata belongs to aiphyllium seeds, originate in Malaysia and Indonesia, the feature such as strong with its fast-growing, fixed nitrogen is fostered and apply fertilizer and by extensive introducing and planting in China torrid zone and South Subtropical Area of China.At present, blbizzia falcata production kind is substantially all introduce a fine variety in early days on a small amount of scattered elite stand that remains to gather, and has that growth rate differs, branch quantity and a Differentiation Problems such as not of uniform size.Breed blbizzia falcata by tissue culture technique both can solve standing forest and break up large problem, in a short time a small amount of fine individual plant can be bred a large amount of nursery stock again, contribute to clone and the improved variety of blbizzia falcata afforestation.
Explant source now for blbizzia falcata tissue cultures mainly contains 2 approach, and one is adopt large tree girdling method, gathers rudiment bar as explant in blbizzia falcata standing forest; Another kind is in standing forest, gather branch grafting, then carries out topping process to grafting, and the terminal bud, the axillalry bud that gather the edible tender branch germinated then are explant.The explant that these 2 kinds of methods obtain is due to long term growth lowered in field environment, and be difficult to kill the mushroom of organization internal by surface sterilization and cause polluting, sterile system is difficult to set up.
Summary of the invention
Based on this, the object of the present invention is to provide a kind of breeding method of blbizzia falcata tissue cultures explant, overcome the deficiency of existing blbizzia falcata tissue cultures explant acquiring technology, quickly breeding quality is good, quantity is many, easy sterilizing and the convenient explant collected, and meets the needs of blbizzia falcata tissue-culturing quick-propagation Establishing.
The concrete technical scheme realizing foregoing invention object is as follows:
A breeding method for blbizzia falcata tissue cultures explant, comprises the following steps:
(1) seed disinfection: aseptically, by the seed of blbizzia falcata successively with sterile water washing, liquor potassic permanganate immersion, sterile water washing, HgCl
2the washing of aqueous solution soaking, sterile water carries out disinfection;
(2) soak seed: the sterile water of the seed after sterilization with 85 ~ 90 DEG C is soaked 22 ~ 26 hours;
(3) vernalization: aseptically, the seed after seed soaking is successively with sterile water washing, HgCl
2vernalization container is put into, vernalization 2 ~ 4 days under 25 ~ 35 DEG C of aseptic dark conditions after the washing of aqueous solution soaking, sterile water;
Described vernalization container is the sealable culture vessel of high temperature high voltage resistant, and the thickness that bottom tiling sterile water soaks into is the good material of the water imbibition of 2 ~ 5mm; Described vernalization container was through autoclave sterilization 15 ~ 25 minutes, for subsequent use after cooling;
(4) inoculate: aseptically, the seed exposing white young root is taken out, successively with sterile water washing, HgCl
2dry after the washing of aqueous solution soaking, sterile water, young root is inserted raise seedling culture medium;
Described raise seedling culture medium is composed of the following components: 1/2MS medium, 0.15 ~ 0.25mg/L indolebutyric acid, 0.6 ~ 1.0mg/L methyl α-naphthyl acetate, 20 ~ 30g/L sucrose, 6 ~ 8g/L carragheen; The pH of described raise seedling culture medium is 5.6 ~ 6;
(5) nursery: cultivate after inoculation and can obtain described explant in 6 ~ 8 days, condition of culture is: temperature 20 ~ 30 DEG C, illumination every day 14 ~ 18 hours, and the intensity of illumination is 750 ~ 1500lx.
Wherein in some embodiments, described sterile water washing is for washing 2 ~ 5 times with sterile water, and described liquor potassic permanganate soaks for soaking 12 ~ 18 minutes with the liquor potassic permanganate of 4 ~ 6g/L, described HgCl
2aqueous solution soaking is the HgCl with 1.3 ~ 1.7g/L
2aqueous solution soaking 10 ~ 20 minutes.
Wherein in some embodiments, the concentration of described liquor potassic permanganate is 5g/L, described HgCl
2the concentration of the aqueous solution is 1.5g/L.
Wherein in some embodiments, described culture vessel is tissue culture bottle, and the good material of described water imbibition is filter paper.
Wherein in some embodiments, further comprising the steps of before step (1):
A () is collected seed: annual late July to mid-August, gather from cyan transfer yellowish-brown or black to and just cracking time pod;
(b) production of hybrid seeds and seed selection: shine under pod being placed in sunlight and take out seed to natural cracking, choose the seed without damage by disease and insect, full grains, for subsequent use.
Wherein in some embodiments, step (4) described raise seedling culture medium is composed of the following components: 1/2MS medium, 0.2mg/L indolebutyric acid, 0.8mg/L methyl α-naphthyl acetate, 25g/L sucrose, 7g/L carragheen; The pH of described raise seedling culture medium is 5.8.
Wherein in some embodiments, the time of step (5) described cultivation is 7 days, and condition of culture is: temperature 25 ~ 28 DEG C, illumination every day 16 hours, and the intensity of illumination is 1000lx.
Wherein in some embodiments, step (1)-(3) described aseptic condition is the aseptic condition of superclean bench.
The breeding method of blbizzia falcata tissue cultures explant provided by the invention has following beneficial effect:
Overcome deficiency (field acquisition or the cultivation difficulty of existing blbizzia falcata tissue cultures explant acquiring technology, it is many that tissue includes mushroom, easy pollution), operation is simple for the method for the invention, the cycle is short, it is low to pollute, output is high, can quickly breeding quality good, quantity is many, easy sterilizing and the convenient explant collected.By aseptic nursery flow process of the present invention, pollution rate can be controlled within 5%; Pregermination device provided by the invention provides the aseptic seedling raising environment that temperature and humidity is suitable for, is conducive to young root growth, the filter paper that tiles bottom pregermination device replaces the materials such as gauze of the prior art, the young root of damage or the seed that fractures when can prevent from taking out the seed germinateed in inoculation step; Raise seedling culture medium of the present invention can promote that seedling is healthy and strong, grow fast.Result of study finds that the breeding method of blbizzia falcata tissue cultures explant provided by the invention can obtain more than 95% aseptic healthy and strong explant in 10 days, can meet the needs of blbizzia falcata tissue-culturing quick-propagation Establishing.
Embodiment
Below in conjunction with specific embodiment, the present invention is further elaborated.
In the introduction blbizzia falcata provenance of building at Boluo County of Guangdong Province forestry scientific research and family trial woods, selecting from the fine individual plant BLS16 in Malaysian superior families M4 is seed collecting elite stand, 17 years elite stand ages, ripe pod (pod when transferring yellowish-brown to from cyan and just ftracture) 250g is gathered on August 15th, 2013, cultivate the explant of blbizzia falcata tissue cultures with this pod, breeding method comprises the following steps:
(1) production of hybrid seeds and seed selection: shine under gathered pod is placed in sunlight and take out seed to natural cracking, 100, the seed chosen without damage by disease and insect, full grains puts into tissue culture bottle;
(2) seed disinfection and seed soaking: under superclean bench aseptic condition, the seed sterile water chosen is washed 3 times, and the liquor potassic permanganate being placed in 5g/L soaks 15min, and sterile water cleans the HgCl being placed on 1.5g/L
2soak 15min in the aqueous solution, then wash 4 times with sterile water, the sterile water that the seed after sterilization puts into 85 ~ 90 DEG C is soaked 24h;
(3) vernalization: under superclean bench aseptic condition, washs 3 times by the seed sterile water after seed soaking, is placed in the HgCl of 1.5g/L
2soak 15min in the aqueous solution, then wash 4 times with sterile water, put into 5 vernalization containers, in each vernalization container, place 20, seed, vernalization 3 days under 28 ~ 30 DEG C of aseptic dark conditions; Vernalization container is tissue culture bottle, and bottom tiling absorbent filter, after soaking into sterile water, through autoclave sterilization 20min;
(4) inoculate: under superclean bench aseptic condition, the seed exposing white young root is taken out, washs 3 times with sterile water, be placed in the HgCl of 1.5g/L
2soak 10min in the aqueous solution, then wash 4 times with sterile water, take out and dry surface moisture, young root is inserted raise seedling culture medium; Consisting of of described raise seedling culture medium: 1/2MS+ indolebutyric acid (IBA, 0.2mg/L)+methyl α-naphthyl acetate (NAA, 0.8mg/L)+sucrose (25g/L)+carragheen (7g/L), pH value is 5.8; 1/2MS refers to that the composition in MS medium is reduced to original half;
(5) nursery: cultivate after inoculation and obtain aseptic seedling 95 strain in 7 days, condition of culture is: temperature 25 ~ 28 DEG C, illumination every day 16 hours, and the intensity of illumination is 1000lx; The height of seedling of gained 95 strain aseptic seedling is 5 ~ 8cm, and nursery stock is healthy and strong, can directly as group training explant.
Each technical characteristic of the above embodiment can combine arbitrarily, for making description succinct, the all possible combination of each technical characteristic in above-described embodiment is not all described, but, as long as the combination of these technical characteristics does not exist contradiction, be all considered to be the scope that this specification is recorded.
The above embodiment only have expressed several embodiment of the present invention, and it describes comparatively concrete and detailed, but can not therefore be construed as limiting the scope of the patent.It should be pointed out that for the person of ordinary skill of the art, without departing from the inventive concept of the premise, can also make some distortion and improvement, these all belong to protection scope of the present invention.Therefore, the protection domain of patent of the present invention should be as the criterion with claims.
Claims (7)
1. a breeding method for blbizzia falcata tissue cultures explant, is characterized in that, comprises the following steps:
(1) seed disinfection: aseptically, by the seed of blbizzia falcata successively with sterile water washing, liquor potassic permanganate immersion, sterile water washing, HgCl
2the washing of aqueous solution soaking, sterile water carries out disinfection;
(2) soak seed: the sterile water of the seed after sterilization with 85 ~ 90 DEG C is soaked 22 ~ 26 hours;
(3) vernalization: aseptically, the seed after seed soaking is successively with sterile water washing, HgCl
2vernalization container is put into, vernalization 2 ~ 4 days under 25 ~ 35 DEG C of aseptic dark conditions after the washing of aqueous solution soaking, sterile water;
Described vernalization container is the sealable culture vessel of high temperature high voltage resistant, and the thickness that bottom tiling sterile water soaks into is the good material of the water imbibition of 2 ~ 5mm; Described vernalization container was through autoclave sterilization 15 ~ 25 minutes, for subsequent use after cooling;
(4) inoculate: aseptically, the seed exposing white young root is taken out, successively with sterile water washing, HgCl
2dry after the washing of aqueous solution soaking, sterile water, young root is inserted raise seedling culture medium;
Described raise seedling culture medium is composed of the following components: 1/2MS medium, 0.15 ~ 0.25mg/L indolebutyric acid, 0.6 ~ 1.0mg/L methyl α-naphthyl acetate, 20 ~ 30g/L sucrose, 6 ~ 8g/L carragheen; The pH of described raise seedling culture medium is 5.6 ~ 6;
(5) nursery: cultivate after inoculation and can obtain described explant in 6 ~ 8 days, condition of culture is: temperature 20 ~ 30 DEG C, illumination every day 14 ~ 18 hours, and the intensity of illumination is 750 ~ 1500lx.
2. the breeding method of blbizzia falcata tissue cultures explant according to claim 1, it is characterized in that, described sterile water washing is for washing 2 ~ 5 times with sterile water, and described liquor potassic permanganate soaks for soaking 12 ~ 18 minutes with the liquor potassic permanganate of 4 ~ 6g/L, described HgCl
2aqueous solution soaking is the HgCl with 1.3 ~ 1.7g/L
2aqueous solution soaking 10 ~ 20 minutes.
3. the breeding method of blbizzia falcata tissue cultures explant according to claim 3, is characterized in that, the concentration of described liquor potassic permanganate is 5g/L, described HgCl
2the concentration of the aqueous solution is 1.5g/L.
4. the breeding method of blbizzia falcata tissue cultures explant according to claim 1, is characterized in that, step (3) described culture vessel is tissue culture bottle, and the good material of described water imbibition is filter paper.
5. the breeding method of the blbizzia falcata tissue cultures explant according to claim 1-4, is characterized in that, further comprising the steps of before step (1):
A () is collected seed: annual late July to mid-August, gather from cyan transfer yellowish-brown or black to and just cracking time pod;
(b) production of hybrid seeds and seed selection: shine under pod being placed in sunlight and take out seed to natural cracking, choose the seed without damage by disease and insect, full grains, for subsequent use.
6. the breeding method of the blbizzia falcata tissue cultures explant according to any one of claim 1-4, it is characterized in that, step (4) described raise seedling culture medium is composed of the following components: 1/2MS medium, 0.2mg/L indolebutyric acid, 0.8mg/L methyl α-naphthyl acetate, 25g/L sucrose, 7g/L carragheen; The pH of described raise seedling culture medium is 5.8.
7. the breeding method of the blbizzia falcata tissue cultures explant according to any one of claim 1-4, it is characterized in that, the time of step (5) described cultivation is 7 days, and condition of culture is: temperature 25 ~ 28 DEG C, illumination every day 16 hours, the intensity of illumination is 1000lx.
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Citations (3)
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| CN103098653A (en) * | 2013-02-06 | 2013-05-15 | 广东省林业科学研究院 | Grafting method of albizia falcataria |
| CN103444549A (en) * | 2013-09-17 | 2013-12-18 | 南京通泽农业科技有限公司 | Rapid albizia chinensis propagation method |
| CN104186183A (en) * | 2014-09-12 | 2014-12-10 | 南京通泽农业科技有限公司 | Cutting propagation method for albizia chinensis |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103098653A (en) * | 2013-02-06 | 2013-05-15 | 广东省林业科学研究院 | Grafting method of albizia falcataria |
| CN103444549A (en) * | 2013-09-17 | 2013-12-18 | 南京通泽农业科技有限公司 | Rapid albizia chinensis propagation method |
| CN104186183A (en) * | 2014-09-12 | 2014-12-10 | 南京通泽农业科技有限公司 | Cutting propagation method for albizia chinensis |
Non-Patent Citations (4)
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| 刘君昂: "红汁乳菇菌种分离培养及其对马尾松苗木生长效应研究", 《林业科学研究》 * |
| 奚伟鹏等: "南洋楹快速繁殖技术", 《广西林业科学》 * |
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