CN104106464A - Explant disinfection method in plant tissue culture - Google Patents
Explant disinfection method in plant tissue culture Download PDFInfo
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- CN104106464A CN104106464A CN201410261359.4A CN201410261359A CN104106464A CN 104106464 A CN104106464 A CN 104106464A CN 201410261359 A CN201410261359 A CN 201410261359A CN 104106464 A CN104106464 A CN 104106464A
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Abstract
An explant disinfection method in plant tissue culture is disclosed. An explant material is washed by tap water, the cleaned explant material is disinfected by ethylene oxide, and then the explant material is treated by a disinfectant under ultrasonic conditions by utilization of the disinfectant so that disinfection is completed. The method has characteristics of easy disinfection, good disinfection effects, low toxicity, and difficult damaging for the material.
Description
Technical field
The present invention relates to field of plant tissue culture technique, relate in particular to the method for explant sterilization in a kind of Plant Tissue Breeding.
Background technology
The explant major part that Plant Tissue Breeding is used is taken from field, adheres to a large amount of microorganisms on surface, and this is the large obstacle that tissue is cultivated.Therefore before material inoculated and cultured, must disinfect, sterilization requires on the one hand the various microorganism killings on material surface, can not damage again or slight damage organization material and do not affect its growth only simultaneously.Therefore, disinfecting of explant is the important ring in Plant Tissue Breeding work.
Existing sterilization method, Disinfection Effect is bad, and the microorganism on material can not be killed to greatest extent, and, existing sterilization method, explant is vulnerable to infringement, greatly reduces the survival rate of Plant Tissue Breeding.
Summary of the invention
The object of this part is to summarize some aspects of embodiments of the invention and briefly introduces some preferred embodiments.In this part and the application's specification digest and denomination of invention, may do a little simplification or omit to avoid the making object of this part, specification digest and denomination of invention fuzzy, and this simplification or omit can not be for limiting the scope of the invention.
Problem in view of existing in explant sterilization method in above-mentioned and/or existing Plant Tissue Breeding, has proposed the present invention.
Therefore, the present invention wherein object is to provide a kind of method that in Plant Tissue Breeding, explant is sterilized, this sterilization method, and its sterilization is easy, Disinfection Effect is good, and toxicity is little, does not damage material.
For solving the problems of the technologies described above, according to an aspect of the present invention, the invention provides following technical scheme: a kind of method of explant sterilization in Plant Tissue Breeding, adopt material running water to clean explant material, then adopt oxirane to carry out disinfection to the explant material after cleaning, finally adopt thimerosal lower the explant material liquid that carries out disinfection to be processed ultrasonic, complete sterilization.
As preferred embodiment of the present invention, described thimerosal is the mixed solution of mercuric chloride and tween disinfectant.
As preferred embodiment of the present invention, it specifically comprises the steps:
Step 1: utilize running water to rinse explant material 30～50min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 2～4 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 0.025～1% mercuric chloride and 0.1～0.2% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 5～15min, then use sterile water wash explant material 5～7 times.
As preferred embodiment of the present invention, in described step 1, the time that running water rinses explant material is 50min.
As preferred embodiment of the present invention, in described step 2, the time that adopts oxirane to carry out disinfection to the explant material after cleaning is 3 hours.
As preferred embodiment of the present invention, in described step 3, pretreated explant material is immersed in the mixed solution that contains 0.025% mercuric chloride and 0.1% tween disinfectant, and the ultrasonic oscillation 10min of use.
As preferred embodiment of the present invention, in described step 3, adopting the number of times of sterile water wash explant material is 6 times.
As preferred embodiment of the present invention, the power of described Vltrasonic device is 90W.
The method that the invention provides explant sterilization in a kind of Plant Tissue Breeding, its sterilization is easy, bactericidal effect good, and toxicity is little, and explant material is easy damaged not.
Embodiment
For above-mentioned purpose of the present invention, feature and advantage can be become apparent more, below in conjunction with accompanying drawing, the specific embodiment of the present invention is described in detail.
A lot of details have been set forth in the following description so that fully understand the present invention, but the present invention can also adopt other to be different from alternate manner described here and implement, those skilled in the art can do similar popularization without prejudice to intension of the present invention in the situation that, so the present invention is not subject to the restriction of following public specific embodiment.
The invention provides the method for explant sterilization in a kind of Plant Tissue Breeding, adopt material running water to clean explant material, then adopt oxirane to carry out disinfection to the explant material after cleaning, finally adopt thimerosal lower the explant material liquid that carries out disinfection to be processed ultrasonic, complete sterilization.
Below adopt embodiment to describe embodiments of the present invention in detail, to the present invention, how application technology means solve technical problem whereby, and the implementation procedure of reaching technique effect can fully understand and implement according to this.
It should be noted that the mixed solution that the thimerosal that the present invention uses is mercuric chloride and tween disinfectant.
Embodiment 1
Explant material is cut into segment or fritter or sheet or thread, then carries out disinfection according to the following steps:
Step 1: utilize running water to rinse explant material 30min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 2 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 0.025% mercuric chloride and 0.1% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 5min, then use sterile water wash explant material 5 times, complete sterilization.
Embodiment 2
Explant material is cut into segment or fritter or sheet or thread, then carries out disinfection according to the following steps:
Step 1: utilize running water to rinse explant material 50min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 3 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 0.025% mercuric chloride and 0.1% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 10min, then use sterile water wash explant material 6 times, complete sterilization.
Embodiment 3
Explant material is cut into segment or fritter or sheet or thread, then carries out disinfection according to the following steps:
Step 1: utilize running water to rinse explant material 40min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 4 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 1% mercuric chloride and 0.2% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 15min, then use sterile water wash explant material 7 times, complete sterilization.
Embodiment 4
Explant material is cut into segment or fritter or sheet or thread, then carries out disinfection according to the following steps:
Step 1: utilize running water to rinse explant material 50min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 2 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 0.025% mercuric chloride and 0.1% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 10min, then use sterile water wash explant material 5 times, complete sterilization.
Sterilization effect
Select the sterilization method of the embodiment of the present invention 1～4 and traditional sterilizing methods to organize culture experiment to explant, its aseptic rate contrast table is in Table 1:
Table 1 sterilization method of the present invention and the traditional method for disinfection killing effect to bacterium and fungi
| Sequence number | Sterilization complexity | Aseptic rate (%) |
| Embodiment 1 | Easily | 96 |
| Embodiment 2 | The easiest | 100 |
| Embodiment 3 | The easiest | 100 |
| Embodiment 4 | Easily | 99 |
| Tradition method for disinfection | Be difficult for | 20.1 |
It should be noted that, the present invention does not limit the kind of explant and position.
As can be seen here, the method for explant sterilization in Plant Tissue Breeding of the present invention, its sterilization is easy, bactericidal effect good, and toxicity is little, not easy damaged explant material.
It should be noted that, above embodiment is only unrestricted in order to technical scheme of the present invention to be described, although the present invention is had been described in detail with reference to preferred embodiment, those of ordinary skill in the art is to be understood that, can modify or be equal to replacement technical scheme of the present invention, and not departing from the spirit and scope of technical solution of the present invention, it all should be encompassed in the middle of claim scope of the present invention.
Claims (8)
1. the method that in a Plant Tissue Breeding, explant is sterilized, it is characterized in that: adopt material running water to clean explant material, then adopt oxirane to carry out disinfection to the explant material after cleaning, finally adopt thimerosal lower the explant material liquid that carries out disinfection to be processed ultrasonic, complete sterilization.
2. the method for explant sterilization in Plant Tissue Breeding according to claim 1, is characterized in that: described thimerosal is the mixed solution of mercuric chloride and tween disinfectant.
3. the method for explant sterilization in Plant Tissue Breeding according to claim 2, is characterized in that: it specifically comprises the steps:
Step 1: utilize running water to rinse explant material 30～50min;
Step 2: at room temperature, the explant material after utilizing oxirane to above-mentioned cleaning carries out disinfection 2～4 hours, completes pretreatment;
Step 3: above-mentioned pretreated explant material is immersed in the mixed solution that contains 0.025～1% mercuric chloride and 0.1～0.2% tween disinfectant, and with Vltrasonic device ultrasonic oscillation 5～15min, then use sterile water wash explant material 5～7 times.
4. the method that in Plant Tissue Breeding according to claim 3, explant is sterilized, is characterized in that:
In described step 1, the time that running water rinses explant material is 50min.
5. the method for explant sterilization in Plant Tissue Breeding according to claim 3, is characterized in that: in described step 2, the time that adopts oxirane to carry out disinfection to the explant material after cleaning is 3 hours.
6. the method that in Plant Tissue Breeding according to claim 3, explant is sterilized, it is characterized in that: in described step 3, pretreated explant material is immersed in the mixed solution that contains 0.025% mercuric chloride and 0.1% tween disinfectant, and the ultrasonic oscillation 10min of use.
7. the method for explant sterilization in Plant Tissue Breeding according to claim 3, is characterized in that: in described step 3, adopting the number of times of sterile water wash explant material is 6 times.
8. the method for explant sterilization in Plant Tissue Breeding according to claim 3, is characterized in that: the power of described Vltrasonic device is 90W.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201410261359.4A CN104106464B (en) | 2014-06-12 | 2014-06-12 | The method of explant sterilization in a kind of Plant Tissue Breeding |
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| CN201410261359.4A CN104106464B (en) | 2014-06-12 | 2014-06-12 | The method of explant sterilization in a kind of Plant Tissue Breeding |
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| CN104106464B CN104106464B (en) | 2016-03-30 |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105850730A (en) * | 2016-04-06 | 2016-08-17 | 南京晓庄学院 | Plant material surface disinfection method |
| CN106613951A (en) * | 2016-10-21 | 2017-05-10 | 定州市绿谷农业科技发展有限公司 | Method for disinfection of explant in tissue culture |
| CN106888976A (en) * | 2017-04-01 | 2017-06-27 | 中国农业科学院草原研究所 | Plant Tissue Breeding Explant sterilizing device |
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| CN102823497A (en) * | 2012-09-19 | 2012-12-19 | 广东省林业科学研究院 | Clonal tissue culture breeding method of Liquidambar formosana Hance |
| CN103120126A (en) * | 2013-02-04 | 2013-05-29 | 南京工业大学 | Method for carrying out tissue cultivation propagation of anoectochilus roxburghii by intermittent submerged bioreactor |
| CN103125389A (en) * | 2013-03-04 | 2013-06-05 | 北京农业生物技术研究中心 | Method for increasing cycle conversion utilization of Zoysia japonica high frequency regeneration series |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102823497A (en) * | 2012-09-19 | 2012-12-19 | 广东省林业科学研究院 | Clonal tissue culture breeding method of Liquidambar formosana Hance |
| CN103120126A (en) * | 2013-02-04 | 2013-05-29 | 南京工业大学 | Method for carrying out tissue cultivation propagation of anoectochilus roxburghii by intermittent submerged bioreactor |
| CN103125389A (en) * | 2013-03-04 | 2013-06-05 | 北京农业生物技术研究中心 | Method for increasing cycle conversion utilization of Zoysia japonica high frequency regeneration series |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105850730A (en) * | 2016-04-06 | 2016-08-17 | 南京晓庄学院 | Plant material surface disinfection method |
| CN106613951A (en) * | 2016-10-21 | 2017-05-10 | 定州市绿谷农业科技发展有限公司 | Method for disinfection of explant in tissue culture |
| CN106888976A (en) * | 2017-04-01 | 2017-06-27 | 中国农业科学院草原研究所 | Plant Tissue Breeding Explant sterilizing device |
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| CN104106464B (en) | 2016-03-30 |
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