CN103918553A - Environment-friendly explant sterilizing method and sterilizing solution - Google Patents
Environment-friendly explant sterilizing method and sterilizing solution Download PDFInfo
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- CN103918553A CN103918553A CN201410137112.1A CN201410137112A CN103918553A CN 103918553 A CN103918553 A CN 103918553A CN 201410137112 A CN201410137112 A CN 201410137112A CN 103918553 A CN103918553 A CN 103918553A
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Abstract
The invention discloses an environment-friendly explant sterilizing solution, namely a garlic aqueous solution, which is prepared by peeling the garlic, mashing, and then mixing with sterile distilled water according to a weight ratio (by volume) of (1-9):1. The invention further provides an environment-friendly explant sterilizing method based on the sterilizing solution, wherein plant explant is immersed into the garlic aqueous solution for sterilizing. The new explant sterilizing method, namely the garlic aqueous solution method is moderate for treated plants, high in sterilization rate which can be up to 100%, stable in sterilizing effect, simple to operate, low in cost, and friendly to environment.
Description
Technical field
The present invention relates to the sterilizing of plant explants, more particularly, relate to the improvement of sterilizing methods and sterile solution.
Background technology
Explant sterilization is that Plant Tissue Breeding can a successful important step.Traditionally, when Plant Tissue Breeding, conventional explant sterilization method is: explant soaks and is no more than 30s in 70% or 75% ethanol, and then with clorox (NaOCl) or calcium hypochlorite [Ca (OCl)
2] or mercuric chloride (mercury chloride, HgCI
2) further sterilization, just can receive good explant sterilization effect.The typical concentrations 0.1% of mercuric chloride, processing time 10～15min, bactericidal effect is good, but residual hazard is high.After sterilization, need to use aseptic water washing 3～5 times; Clorox and calcium hypochlorite typical concentrations can be 0.5%～10%, sterilizing time 5～30min, and Comparison of effect on sterilization is good, harmless to plant; But clorox and calcium hypochlorite more or less have certain toxicity.Above-mentioned chemical reagent cost is higher.
In prior art, those skilled in the art select the branch of resting stage as explant mostly, and do not select the young sprout in growth season.Because the sterilizing solvent of prior art is larger to the tender tip infringement of plant, the tender tip cannot be survived.Therefore those skilled in the art can not select the tender tip to do explant.Herein, to be plant growth season push up one section of long young sprout of 5～10cm on the lower side or branch from axis to the described tender tip, therefore, as the tender tip of the part of young sprout front end, if need to be used as explant, need to select gentle sterilizing solvent.
Summary of the invention
The invention provides explant sterilization solution---the garlic that a kind of cost is low, and the method that uses garlic sterilization is provided.The bactericidal action of application garlic, while becoming Plant Tissue Breeding, new, the effective and antihunt means of explant sterilization, do not injure explant.With realize explant sterilization reagent without poisoning, and reached the effect of preserving the ecological environment, the young sprout in the season that is particularly useful for growing uses as explant.
In order to achieve the above object, the invention provides a kind of environmentally friendly explant sterilization solution, utilize the aqueous solution of garlic configuration.
Collocation method is various, as the method that water configuration was dried, pulverized, adds to garlic, the method for soaking in garlic chopping for another example, water.A kind of optimal way provided by the invention is: smash into mud with peeling garlic, with aseptic single steam water by weight volume ratio be configured to the solution of 1～9:1.
The present invention also provides the environmentally friendly explant sterilization method based on above-mentioned sterile solution, and plant explants is immersed in the aqueous solution based on garlic configuration.In addition,, under optimal way, explant first rinses 1 hour under running water.
Explant sterilization new method used in the present invention---garlic aqua-solution method, gentle to handled plant performance, sterilization rate is higher, can reach 100%, and sterilization effect is stable, and simple to operate, cost is low, environmentally friendly.Specifically, advantage of the present invention is as follows:
1. the reagent of explant sterilization (as ethanol, clorox, calcium hypochlorite, mercury chloride etc.) while replacing traditional Plant Tissue Breeding with the garlic aqueous solution, having realized the avirulence of explant sterilization, is a kind of environmentally friendly Plant Tissue Breeding explant sterilization agent.
This kind of explant sterilization agent for sweet cherry, peach, Chinese rose etc. dormancy season leaf bud sterilizing time, aseptic rate reaches 100%.
3., when this kind of explant sterilization agent is used for the sterilizing of the tender tip in the growth season of sweet cherry, peach, Chinese rose etc., aseptic rate reaches 100%.
4. compared with traditional explant sterilization agent, sterilizing cost. in growth season, the sterilizing cost of the garlic aqueous solution is 8.7% of traditional bactericidal agent (70% ethanol+clorox); In dormancy season, the sterilizing cost of the garlic aqueous solution is 17.4% of traditional bactericidal agent (70% ethanol+clorox).
5. operation is simpler.The explant running water of preparing sterilizing can be rinsed after 1h, directly be immersed in the garlic aqueous solution, and do not need other pre-treatment, as the sheet etc. of scaling.
In addition, the tender tip is as explant, and because the ability of anti-external environmental interference is poor, therefore, the sterile solution of prior art is larger to its injury, has therefore limited the possibility that the tender tip uses as explant.For this reason, the invention provides a kind of novel sterile solution---utilize the aqueous solution of garlic configuration.Specifically smash into mud with peeling garlic, with aseptic single steam water by weight volume ratio be configured to the solution of 1～9:1.This sterile solution gentleness, is applicable to the sterilizing of the tender tip of plant as explant.Herein, to be plant growth season push up one section of long young sprout of 5～10cm on the lower side or branch from axis to the described tender tip.
Embodiment
The present invention is based on the sterilizing mechanisms of allicin and garlic oil in garlic, carry out the research of garlic aqueous solution sterilization effect as explant using leaf bud and the tender tip in growth season in dormancy season.Respectively in the time of winter Mo, get the leaf bud of sweet cherry, peach, Chinese rose as explant, carry out garlic aqueous solution sterilization treatment, the aseptic rate of observing, adding up leaf bud, and in growth season, get the tender tip of sweet cherry, peach, Chinese rose as explant, carry out garlic aqueous solution sterilization treatment, observe, add up the aseptic rate of the tender tip.
Concrete technology flow process:
1, get the explant of different times;
2, be immersed in the garlic aqueous solution;
3, be seeded on suitable medium;
4, research sterilization effect.
Embodiment 1: garlic: aseptic single steaming water is 2:1(W/V), peeling garlic reduces to pulp and adds aseptic single water stirring 2min that steams, and then under room temperature, places after 30min, filters rear for subsequent use.
Embodiment 2: garlic: aseptic single steaming water is 1:1(W/V), peeling garlic reduces to pulp and adds aseptic single water stirring 2min that steams, and then under room temperature, places after 30min, filters rear for subsequent use.
Embodiment 3: garlic: aseptic single steaming water is 1:3(W/V), peeling garlic reduces to pulp and adds aseptic single water stirring 2min that steams, and then under room temperature, places after 30min, filters rear for subsequent use.
The occupation mode of above-mentioned garlic is:
In winter end, get the leaf bud of sweet cherry, peach, Chinese rose as explant, taken back indoor, running water rinse 1h.Under aseptic technique, carry out explant sterilization, the leaf bud of sweet cherry, peach, Chinese rose is respectively put into the garlic aqueous solution of embodiment 1, embodiment 2.Middle immersion 30min, rinses 3 times with aseptic single water that steams after taking-up, uses aseptic filter paper suck dry moisture, is inoculated on first culture base (culture medium prescription is: MS+6-BA0.2mg/L+IBA0.05mg/L+ agar 7g/L+ sucrose 30g/L, pH5.8).
In growth season, get sweet cherry, peach, the tender tip in the long top of Chinese rose 2～3cm, directly put into respectively the garlic aqueous solution soaking 30min of embodiment 1, embodiment 2, after taking out, rinse 3 times with aseptic single water that steams, finally use aseptic filter paper suck dry moisture, be inoculated on first culture base (culture medium prescription is: MS+6-BA0.2mg/L+IBA0.05mg/L+ agar 7g/L+ sucrose 30g/L, pH5.8).
3. observe and add up the average aseptic rate (sterilization rate can reach 100%) of the germination rate of leaf bud, the tender tip.
4. on the basis of first culture, proceed subculture cultivation, culture of rootage and rooting culture etc.
Wherein, for the garlic of variable concentrations, according to the kind of concrete plant, appropriateness regulates soak time, as the solution of embodiment 3, needs proper extension soak time.
By above-described embodiment, respectively taking the leaf bud in dormancy season and the tender tip in growth season as explant, successfully set up explant sterilization technical system before the Plant Tissue Breeding inoculations such as sweet cherry, peach, Chinese rose, effect stability.
In addition, adopt the 3:1 garlic aqueous solution following plants explant to be carried out to sterilization treatment, effect: pollute body and be 0, sterilization effect is 100%.These plant explants have: peach, cherry tree, apricot, Japanese plum, Chinese rose, blueberry, raspberry, grape, Chinese white poplar, italian poplar, Chinese scholar tree, Chinese pagoda tree, apple, Moth orchid resting stage leaf bud (sterilizing 30 minutes) and the tender tip in vegetative period (sterilizing 20 minutes).
For different explants, the time of sterilizing is different, and for the leaf bud of resting stage, sterilization time is slightly more longer than the tender tip in growth season, because different Genes For Plant Tolerance kill capabilitys is also different, the anti-kill capability of leaf bud of same plant dormancy phase is stronger.
The above; it is only preferably embodiment of the present invention; but protection scope of the present invention is not limited to this; any be familiar with those skilled in the art the present invention disclose technical scope in; be equal to replacement or changed according to technical scheme of the present invention and inventive concept thereof, within all should being encompassed in protection scope of the present invention.
Claims (4)
1. an environmentally friendly explant sterilization solution, is characterized in that, utilizes the aqueous solution of garlic configuration.
2. environmentally friendly explant sterilization solution according to claim 1, is characterized in that, smashes into mud with peeling garlic, with aseptic single steam water by weight volume ratio be configured to the solution of 1～9:1.
3. the environmentally friendly explant sterilization method based on above-mentioned sterile solution, is characterized in that, plant explants is immersed to sterilizing in the aqueous solution based on garlic configuration.
4. the environmentally friendly explant sterilization method based on above-mentioned sterile solution according to claim 3, is characterized in that, described sterile solution is: peeling garlic is smash into mud, with aseptic single steam water by weight volume ratio be configured to the solution of 1～9:1.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105961205A (en) * | 2016-07-28 | 2016-09-28 | 广西陆川县乌坭坡珍珠番石榴专业合作社 | Tissue culture method for increasing survival rate of psidium guajava L. |
| CN106613967A (en) * | 2016-12-01 | 2017-05-10 | 大连大学 | Explant for detoxification of plant tissue culture and detoxification method of explant |
| CN108575739A (en) * | 2018-01-23 | 2018-09-28 | 昆明学院 | The bacteria inhibiting composition and preparation method thereof of blueberry open tissue culture |
| CN109452330A (en) * | 2017-11-21 | 2019-03-12 | 浙江省嘉兴市农业科学研究院（所） | A kind of Plant Tissue Breeding bacteriostatic agent and its application in roxburgh anoectochilus terminal bud tissue cultures |
| CN110250225A (en) * | 2019-06-28 | 2019-09-20 | 大连大学 | A kind of explant sterilizing reagent and preparation method thereof |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105961205A (en) * | 2016-07-28 | 2016-09-28 | 广西陆川县乌坭坡珍珠番石榴专业合作社 | Tissue culture method for increasing survival rate of psidium guajava L. |
| CN105961205B (en) * | 2016-07-28 | 2017-10-27 | 广西陆川县乌坭坡珍珠番石榴专业合作社 | A kind of method for tissue culture for improving pearl guava survival rate |
| CN106613967A (en) * | 2016-12-01 | 2017-05-10 | 大连大学 | Explant for detoxification of plant tissue culture and detoxification method of explant |
| CN109452330A (en) * | 2017-11-21 | 2019-03-12 | 浙江省嘉兴市农业科学研究院（所） | A kind of Plant Tissue Breeding bacteriostatic agent and its application in roxburgh anoectochilus terminal bud tissue cultures |
| CN109452330B (en) * | 2017-11-21 | 2020-10-20 | 浙江省嘉兴市农业科学研究院（所） | Bacteriostatic agent for plant tissue culture and application of bacteriostatic agent in anoectochilus formosanus tissue culture |
| CN108575739A (en) * | 2018-01-23 | 2018-09-28 | 昆明学院 | The bacteria inhibiting composition and preparation method thereof of blueberry open tissue culture |
| CN108575739B (en) * | 2018-01-23 | 2021-08-03 | 昆明学院 | Bacteriostatic composition for open tissue culture of blueberries and preparation method thereof |
| CN110250225A (en) * | 2019-06-28 | 2019-09-20 | 大连大学 | A kind of explant sterilizing reagent and preparation method thereof |
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Application publication date: 20140716 |