CN107278901A - A kind of sterilization method of iris bennet bud - Google Patents
A kind of sterilization method of iris bennet bud Download PDFInfo
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- CN107278901A CN107278901A CN201710644971.3A CN201710644971A CN107278901A CN 107278901 A CN107278901 A CN 107278901A CN 201710644971 A CN201710644971 A CN 201710644971A CN 107278901 A CN107278901 A CN 107278901A
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- bennet
- bud
- thimerosals
- bennet bud
- thimerosal
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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- Developmental Biology & Embryology (AREA)
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Abstract
The present invention is a kind of sterilization method of iris bennet bud, and its step is as follows：The complete Phalaenopsis plants of choosing stalk bud, retain the bennet part with bennet bud；From bottom to top wiped with alcohol swab 35 times；And be classified according to old tender degree, it is divided into two kinds of presbyopic stalk and tender pedicel；Afterwards prepare three kinds of thimerosals, by presbyopic stalk and tender pedicel be respectively put into two empty aseptic bottles, by No. 1 thimerosal pour into equipped with presbyopic obstruct bottle in, No. 2 thimerosals are poured into the bottle equipped with tender pedicel, after pour out thimerosal；Peel off for the first time sterilize after presbyopic stalk and tender pedicel bennet bud on bract, the bennet bud after bract will be divested and be immersed in No. 3 thimerosals, after pour out thimerosal；Then bennet bud is inoculated into the blake bottle equipped with bennet bud inducement cultivation base；The blake bottle being inoculated with is put into the induction that bennet bud is carried out in culturing room.Bennet bud pollution rate is 2 7%, and the survival rate of bennet bud is 95 99%, and the cost of thimerosal is low.
Description
Technical field
Sterilization method, particularly a kind of sterilization method of iris bennet bud are cultivated the present invention relates to a kind of bennet bud.
Background technology
The current most domestic iris tissue culture producer is to butterfly with liquor natrii hypochloritis or mercuric chloride solution
Phalaenopsis bennet bud carries out disinfection processing, and mercuric chloride solution is very good to the Disinfection Effect of bennet bud, but it contain to environment and
Soil has the mercury of severe contamination, thus many responsible producers all abandon without.Liquor natrii hypochloritis's Disinfection Effect is good, easy
Cleaning, it is pollution-free, be the main thimerosal of domestic iris tissue culture producer, iris bennet bud carried out with sodium hypochlorite at present
The pollution rate of sterilization is 20% or so, and sodium hypochlorite is chemical reagent, buy it need special chemical reagent shop and
Price is higher.
The content of the invention
The technical problems to be solved by the invention are in view of the shortcomings of the prior art the sides there is provided a kind of raw material is cheap and easy to get
Method is simple, the sterilization method for the iris bennet bud that pollution rate is low, Disinfection Effect is good.
The technical problems to be solved by the invention are realized by following technical scheme, and the present invention is a kind of butterfly
Orchid obstructs the sterilization method of bud, is characterized in, its step is as follows：
(1) bennet is chosen：Choose at least bloom three, virus-free performance, the iris that inflorescence is normal, bennet bud is complete plant
Strain, is cut with the instrument after sterilization at the base portion 8cm-12cm away from bennet, and cuts off inflorescence above, is only retained and is carried bennet bud
Bennet part；
(2) bennet is slightly sterilized：Bennet with bennet bud is from bottom to top wiped 3-5 times with 70%-75% alcohol swab；
(3) bennet bud is classified：Bennet bud is cut respectively on aseptic operating platform, and is classified according to old tender degree,
It is divided into two kinds of presbyopic stalk and tender pedicel；Above bennet bud at 1.5cm-2cm, below bennet bud during clip bennet bud
Clip at 2cm-3cm；Tender pedicel is the bennet near from top；
(4) thimerosal prepares：Three kinds of thimerosals are prepared respectively, and No. 1 thimerosal is 1 by volume ratio：1.5-2.5 84 sterilization
Liquid and sterilized water composition；No. 2 thimerosals are made up of volume ratio for 1: 2.8-3.5 84 thimerosals and sterilized water；No. 3 thimerosals by
84 thimerosals and the sterilized water composition that volume ratio is 1: 8.5-9.5；Three kinds of thimerosals are all separately added into 1-3 drops after preparing and told
Temperature；
(5) sterilize for the first time：Presbyopic stalk and tender pedicel are respectively put into two empty aseptic bottles, No. 1 thimerosal is fallen
Enter in the bottle obstructed equipped with presbyopic, No. 2 thimerosals are poured into the bottle equipped with tender pedicel, presbyopic stalk and tender pedicel are immersed in sterilization
In liquid, fully shake, after 20-30 minutes, pour out thimerosal；
(6) bract is divested：The bract on presbyopic stalk and tender pedicel bennet bud after sterilizing for the first time is peelled off, must not when divesting
Hurt bennet and bennet bud；
(7) second of sterilization：The bennet bud after bract will be divested to be fitted into aseptic bottle, pouring into No. 3 thimerosals makes bennet bud
It is immersed in thimerosal, after 3-8 minutes, pours out thimerosal；
(8) it is inoculated with：Bennet bud after secondary sterilization is taken out from aseptic bottle, retain above sprout 0.8cm-1.2cm its
Remaining to cut off, upper clip direction is vertical with bennet direction；Retaining 1.2cm-1.8cm below sprout, remaining cuts off, lower clip direction with
Bennet bud, in the same side, is then inoculated into equipped with bennet bud inducement cultivation base by bennet direction angle at 45 °, lower clip with bennet bud
Blake bottle in；
(9) cultivate：The blake bottle being inoculated with is put into the Fiber differentiation that bennet bud is carried out in culturing room, cultivated 45-60 days
Grow up to intact plant, condition of culture is：26 DEG C -30 DEG C of temperature, light intensity 1000-1400LX, the daily 12-16 of light application time is small
When.
In the sterilization method technical scheme of invention described above iris bennet bud：In above-mentioned steps (8), the flower
Stalk bud inducement cultivation base is to spend No. 1 2-3.5g/L+ coconut milk 50-100ml/L+6-BA3-5PPM+NAA0.2-0.5PPM of treasured.
6-BA is 6- benzyl aminoadenines；NAA is methyl α-naphthyl acetate.
It is that commercially available U.S. HYPONEX spends precious No. 1 composite fertilizer (N: P: K=7: 6: 19) to spend precious No. 1.
In the sterilization method technical scheme of invention described above iris bennet bud：In step (4), described No. 1 disappears
Venom is made up of volume ratio for 1: 2 84 thimerosals and sterilized water；84 thimerosals and nothing of No. 2 thimerosals by volume ratio for 1: 3
Bacterium water is constituted；No. 3 thimerosals are made up of volume ratio for 1: 9 84 thimerosals and sterilized water.
Compared with prior art, the present invention has its following feature：
(1) it is classified, is then carried out disinfection respectively according to the old tender degree of bennet, it is to avoid presbyopic stalk in the same time
Thorough or tender pedicel does not sterilize excessive situation for sterilization；
(2) sterilized, the outer bract of whole exposed bennet and bennet bud outside is carried out thoroughly twice for the first time
Sterilization, carries out disinfection to bennet sprout for the second time, has accomplished comprehensively, thoroughly；
(3) thimerosal is simple, preparation is easy, cost is low, and the thimerosal sterilized twice is all 84 thimerosals of various concentrations
With a few drop tweens, buying is easily, cost is very low, stability is strong, pollution rate is low, preparation is also very easy to；
(4) Disinfection Effect is good, by thorough disinfection twice, and the pollution rate of bennet bud can be controlled in 2%-7%, greatly
It has compressed production cost and female parent material.
Embodiment
Concrete technical scheme of the invention described further below, in order to which those skilled in the art is further understood that
The present invention, without constituting the limitation to its right.
A kind of embodiment 1, sterilization method of iris bennet bud, its step is as follows：
(1) bennet is chosen：Chosen in greenhouse at least bloom three, virus-free performance, inflorescence be normal, bennet bud is complete
Phalaenopsis plants, are cut with the instrument after sterilization such as knife or scissors at the base portion 10cm away from bennet, and cut off inflorescence above,
Only retain the bennet part with bennet bud；
(2) bennet is slightly sterilized：Bennet with bennet bud is from bottom to top wiped 3-5 times with 70%-75% alcohol swab；
Code name and date are write after being wrapped up after wiping with preservative film, later process is sent to.
(3) bennet bud is classified：Bennet bud is cut respectively on aseptic operating platform, and is classified according to old tender degree,
It is divided into two kinds of presbyopic stalk and tender pedicel；Above bennet bud at 2cm, at 3cm below bennet bud during clip bennet bud
Clip；
(4) thimerosal prepares：Three kinds of thimerosals, 84 thimerosals and nothing of No. 1 thimerosal by volume ratio for 1: 2 are prepared respectively
Bacterium water is constituted；No. 2 thimerosals are made up of volume ratio for 1: 3 84 thimerosals and sterilized water；No. 3 thimerosals are 1: 9 by volume ratio
84 thimerosals and sterilized water composition；Three kinds of thimerosals are all separately added into 2 drop tweens after preparing, tween is drawn by glue head dropper
Instill；
(5) sterilize for the first time：Presbyopic stalk and tender pedicel are respectively put into two empty aseptic bottles, No. 1 thimerosal is fallen
Enter in the bottle obstructed equipped with presbyopic, No. 2 thimerosals are poured into the bottle equipped with tender pedicel, presbyopic stalk and tender pedicel are immersed in sterilization
In liquid, fully shake, after 25 minutes, pour out thimerosal；
(6) bract is divested：Presbyopic stalk and tender pedicel bennet after sterilizing for the first time are lightly peelled off with scalpel and tweezers
Bract on bud, must not hurt bennet and bennet bud when divesting；
(7) second of sterilization：The bennet bud after bract will be divested to be fitted into aseptic bottle, pouring into No. 3 thimerosals makes bennet bud
It is immersed in thimerosal, after 5 minutes, pours out thimerosal；
(8) it is inoculated with：Bennet bud after secondary sterilization is taken out from aseptic bottle, retaining 1cm above sprout, remaining cuts off, on
Clip direction is vertical with bennet direction；Retaining 1.5cm below sprout, remaining cuts off, lower clip direction and bennet direction angle at 45 °,
Bennet bud, in same direction, is then inoculated into the blake bottle equipped with bennet bud inducement cultivation base by lower clip with bennet bud；
(9) cultivate：The blake bottle being inoculated with is put into the Fiber differentiation that bennet bud is carried out in culturing room, 5 days or so
See that sprout is expanded, see within 30 days or so sprout elongation or grow up to seedling, 45-60 days or so can long great achievement intact plant, training
Foster condition is：26 DEG C -30 DEG C of temperature, light intensity 1000-1400LX, light application time is daily 12-16 hours.
In embodiment 2, the sterilization method of iris bennet bud described in embodiment 1, the bennet bud inducement cultivation base is flower
No. 1 2g/L+ coconut milk 50ml/L+6-BA 3PPM+NAA 0.2PPM of treasured.
In embodiment 3, the sterilization method of iris bennet bud described in embodiment 1, the bennet bud inducement cultivation base is flower
No. 1 3.5g/L+ coconut milk 100ml/L+6-BA 5PPM+NAA0.5PPM of treasured.
In embodiment 4, the sterilization method of iris bennet bud described in embodiment 1, the bennet bud inducement cultivation base is flower
No. 1 2.8g/L+ coconut milk 75ml/L+6-BA 4PPM+NAA 0.3PPM of treasured.
A kind of embodiment 5, sterilization method of iris bennet bud, its step is as follows：
(1) bennet is chosen：Chosen in greenhouse at least bloom three, virus-free performance, inflorescence be normal, bennet bud is complete
Phalaenopsis plants, are cut with the instrument after sterilization such as knife or scissors at the base portion 8cm away from bennet, and cut off inflorescence above,
Only retain the bennet part with bennet bud；
(2) bennet is slightly sterilized：Bennet with bennet bud is from bottom to top wiped 4 times with 70%-75% alcohol swab；Wipe
Code name and date are write after being wrapped up after wiping with preservative film, later process is sent to.
(3) bennet bud is classified：Bennet bud is cut respectively on aseptic operating platform, and is classified according to old tender degree,
It is divided into two kinds of presbyopic stalk and tender pedicel；Above bennet bud at 2cm, at 2cm below bennet bud during clip bennet bud
Clip；
(4) thimerosal prepares：Respectively prepare three kinds of thimerosals, No. 1 thimerosal by volume ratio for 1: 1.5 84 thimerosals and
Sterilized water is constituted；No. 2 thimerosals are made up of volume ratio for 1: 2.5 84 thimerosals and sterilized water；No. 3 thimerosals are by volume ratio
1: 8.5 84 thimerosals and sterilized water composition；Three kinds of thimerosals are all separately added into 3 drop tweens after preparing, tween is dripped by glue head
Pipe, which is drawn, to be instilled；
(5) sterilize for the first time：Presbyopic stalk and tender pedicel are respectively put into two empty aseptic bottles, No. 1 thimerosal is fallen
Enter in the bottle obstructed equipped with presbyopic, No. 2 thimerosals are poured into the bottle equipped with tender pedicel, presbyopic stalk and tender pedicel are immersed in sterilization
In liquid, fully shake, after 20 minutes, pour out thimerosal；
(6) bract is divested：Presbyopic stalk and tender pedicel bennet after sterilizing for the first time are lightly peelled off with scalpel and tweezers
Bract on bud, must not hurt bennet and bennet bud when divesting；
(7) second of sterilization：The bennet bud after bract will be divested to be fitted into aseptic bottle, pouring into No. 3 thimerosals makes bennet bud
It is immersed in thimerosal, after 8 minutes, pours out thimerosal；
(8) it is inoculated with：Bennet bud after secondary sterilization is taken out from aseptic bottle, retaining 1.2cm above sprout, remaining cuts off,
Upper clip direction is vertical with bennet direction；Retaining 1.8cm below sprout, remaining cuts off, and lower clip direction and bennet direction are at 45 °
Bennet bud, in same direction, is then inoculated into the blake bottle equipped with bennet bud inducement cultivation base by angle, lower clip with bennet bud；
The bennet bud inducement cultivation base is to spend No. 1 2.5g/L+ coconut milk 70ml/L+6-BA 4PPM+NAA 0.35PPM of treasured.
(9) cultivate：The blake bottle being inoculated with is put into the Fiber differentiation that bennet bud is carried out in culturing room, 5 days or so
See that sprout is expanded, see within 30 days or so sprout elongation or grow up to seedling, 45-60 days or so can long great achievement intact plant, training
Foster condition is：26 DEG C -30 DEG C of temperature, light intensity 1000-1400LX, light application time is daily 12-16 hours.
When carrying out the sterilization of iris bennet bud using the sterilization breeding method of the embodiment of the present invention 1 and 5 and cultivate, bennet bud
Pollution rate is 2-7%, and the survival rate of bennet bud is 95-99%.And the cost of thimerosal be other thimerosals such as mercury chloride into
This is low by 1/10, and the influence to environment is also low compared with mercury chloride.
Claims (3)
1. a kind of sterilization method of iris bennet bud, it is characterised in that its step is as follows：
(1) bennet is chosen：Choose at least bloom three, virus-free performance, the Phalaenopsis plants that inflorescence is normal, bennet bud is complete,
Cut with the instrument after sterilization at the base portion 8cm-12cm away from bennet, and cut off inflorescence above, only retained with bennet bud
Bennet part；
(2) bennet is slightly sterilized：Bennet with bennet bud is from bottom to top wiped 3-5 times with 70%-75% alcohol swab；
(3) bennet bud is classified：Bennet bud is cut respectively on aseptic operating platform, and is classified according to old tender degree, is divided into
Presbyopic is obstructed and two kinds of tender pedicel；In the 2cm- at 1.5cm-2cm, below bennet bud above bennet bud during clip bennet bud
Clip at 3cm；
(4) thimerosal prepares：Respectively prepare three kinds of thimerosals, No. 1 thimerosal by volume ratio for 1: 1.5-2.5 84 thimerosals and
Sterilized water is constituted；No. 2 thimerosals are made up of volume ratio for 1: 2.8-3.5 84 thimerosals and sterilized water；No. 3 thimerosals are by volume
Than 84 thimerosals and the sterilized water composition for 1: 8.5-9.5；Three kinds of thimerosals are all separately added into 1-3 drop tweens after preparing；
(5) sterilize for the first time：Presbyopic stalk and tender pedicel are respectively put into two empty aseptic bottles, No. 1 thimerosal is poured into dress
Have in the bottle that presbyopic is obstructed, No. 2 thimerosals are poured into the bottle equipped with tender pedicel, presbyopic stalk and tender pedicel are immersed in thimerosal
In, fully shake, after 20-30 minutes, pour out thimerosal；
(6) bract is divested：The bract on presbyopic stalk and tender pedicel bennet bud after sterilizing for the first time is peelled off, must not be hurt when divesting
Bennet and bennet bud；
(7) second of sterilization：The bennet bud after bract will be divested to be fitted into aseptic bottle, pouring into No. 3 thimerosals soaks bennet bud
In thimerosal, after 3-8 minutes, thimerosal is poured out；
(8) it is inoculated with：Bennet bud after secondary sterilization is taken out from aseptic bottle, retaining 0.8cm-1.2cm above sprout, remaining is cut
Go, upper clip direction is vertical with bennet direction；Retaining 1.2cm-1.8cm below sprout, remaining cuts off, lower clip direction and bennet
Bennet bud, in the same side, is then inoculated into the training equipped with bennet bud inducement cultivation base by direction angle at 45 °, lower clip with bennet bud
Support in bottle；
(9) cultivate：The blake bottle being inoculated with is put into the Fiber differentiation that bennet bud is carried out in culturing room, culture grows up to for 45-60 days
Intact plant, condition of culture is：26 DEG C -30 DEG C of temperature, light intensity 1000-1400LX, light application time is daily 12-16 hours.
2. the sterilization method of iris bennet bud according to claim 1, it is characterised in that：In step (8), the bennet
Bud inducement cultivation base is to spend No. 1 2-3.5g/L+ coconut milk 50-100ml/L+6-BA3-5PPM+NAA0.2-0.5PPM of treasured.
3. the sterilization method of iris bennet bud according to claim 1, it is characterised in that：In step (4), described No. 1
Thimerosal is made up of volume ratio for 1: 2 84 thimerosals and sterilized water；No. 2 thimerosals by volume ratio for 1: 3 84 thimerosals and
Sterilized water is constituted；No. 3 thimerosals are made up of volume ratio for 1: 9 84 thimerosals and sterilized water.
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| CN110896856A (en) * | 2019-11-07 | 2020-03-24 | 郑州师范学院 | Method for inducing complete inflorescence in vitro by axillary bud of inflorescence shaft of phalaenopsis |
| CN113575421A (en) * | 2021-08-27 | 2021-11-02 | 中德润萌生态科技(青岛)有限公司 | Aseptic processing method for tissue culture material of orchidaceae plant |
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| CN113575421A (en) * | 2021-08-27 | 2021-11-02 | 中德润萌生态科技(青岛)有限公司 | Aseptic processing method for tissue culture material of orchidaceae plant |
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