CN101731144B - Method for culturing tomato tissues in test tube - Google Patents
Method for culturing tomato tissues in test tube Download PDFInfo
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- CN101731144B CN101731144B CN2008102259405A CN200810225940A CN101731144B CN 101731144 B CN101731144 B CN 101731144B CN 2008102259405 A CN2008102259405 A CN 2008102259405A CN 200810225940 A CN200810225940 A CN 200810225940A CN 101731144 B CN101731144 B CN 101731144B
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- tomato
- test tube
- tissue culture
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- callus
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Abstract
The invention belongs to the field of biotechnology, and relates to a method for realizing blossoming and fruiting of tomatoes in a test tube by using an induction, differentiation and rooting culture medium used by a tomato tissue culture method. The method comprises the following steps: induction of callus culture, differentiation to become seedling, rooting, differentiation of flower buds, blossoming and fruiting. The whole process of the method is completed in the test tube, which means that the method can enable the tomato to differentiate and proliferate in the test tube, and enable the tomato to blossom and fruit in the test tube. The invention perfects the tomato tissue culture technique, and can enable the tomato to differentiate and proliferate in the test tube and enable the tomato to blossom and fruit in the test tube. Since the culture technique in the test tube is very difficult, the invention embodies a high level of tissue culture.
Description
Technical field
The invention belongs to biological technical field, relate to that a kind of tomato method for tissue culture uses induce, differentiation, root media, utilize this medium tomato to be implemented in the method for yielding positive results in the test tube.
Background technology
Tomato belongs to Solanaceae, tomato genus, tomato species (Latin title Lycopersicon esculantumMill).Tomato is a kind of important economic crops, is the main vegetables wide, that consumption is many that distribute in the world wide, and tomato is again a kind of important experimental material in the bioscience research simultaneously, is research fruit development and ripe model plant.Realize that through method for tissue culture tomato yields positive results in test tube, can a new method be provided for the research of tomato bioscience.For further exploring flowering of plant result's principle mechanisms, theoretical foundation is provided, also can new approach be provided for the miniature cultivation of gardening plant.。
Tomato method for tissue culture of the prior art is mainly used in the basic operation of the biotechnology in the fast numerous and tomato transgenosis of improved seeds; But there are not the research and development of the method for tissue culture that tomato yields positive results in test tube; Because tomato in vitro yields positive results to be different from the soil and yields positive results; Because in the test tube be the microenvironment of an airtight relatively artificial control; Difficult point is for nutrient component in illumination, temperature controlling and the medium and hormone combination, satisfies the demand that tomato can yield positive results in vitro to reach.Conventional tomato tissue culture mainly is for the differentiation and proliferation ability of the regulating tomato tissue reproduction coefficient with raising tomato test-tube plantlet; Or regulate and control for the needs of transgenic experiments; After the differentiation of tomato test-tube plantlet, just forward in the root media, just transplant to soil after taking root.The inventor is on the basis of tomato tissue culture mature technology, and hormone kind in the medium and concentration, nutrient component are improved.Hormone regulation in this invention mainly realizes reaching the purpose of blossoming and bearing fruit by nourishing and growing to the reproductive growth conversion in test tube for the tomato test-tube plantlet, therefore differs widely in the kind of hormone and the tomato tissue culture of concentration and nutrient component and routine.Owing to in vitro blossom and bear fruit and do not receive season limit, can induce at any time, tissue cultured test-tube becomes flower to have into colored rate height, good reproducibility, advantage such as consistent, can be used in a large number therefore that bow structure is real to be studied.
Summary of the invention
The present invention is in order to fill up the blank of the method that method for tissue culture realization tomato yields positive results in the prior art in test tube; Invented a kind of method of in test tube, carrying out the tomato tissue culture, realized that the tomato test-tube plantlet reaches the effect and the purpose of blossoming and bearing fruit by nourishing and growing to the reproductive growth conversion in test tube.
Content of the present invention is:
A kind of method of in test tube, carrying out the tomato tissue culture, said method comprise tomato evoked callus incubation step, break up young stem step, and cultivation is taken root and yielded positive results step; The whole process of said method is all accomplished in test tube, and promptly this method can not only make the tomato differentiation and proliferation in test tube, but also in test tube, accomplishes the group training process of blossoming and bearing fruit.
In the group training of reality, the step of in test tube, carrying out the tomato tissue culture is:
(1) described tomato evoked callus incubation step is: get the tomato young leaflet tablet, with stopping in the Tween-20 that is placed on 1%-10% behind the running water cleaning down 2-10 minute, with the distilled water rinsing of autoclave sterilization 3-4 time, use 0.1% mercuric chloride (HgCl then
2) solution surface sterilization 2-10 minute, use distilled water rinsing 4-5 time of autoclave sterilization again; Leaf tissue be cut into big or small areal extent be 1mm * 1mm～10mm * 10mm as explant, be inoculated into MS minimal medium+6-benzyl aminoadenine (6-BA) 0.1-10mgL
-1+ heteroauxin (IAA) 0.01-1mgL
-1+ agar 4-10g L
-1In the medium of+sucrose 20-50g, callus induction;
(2) the young stem step of described differentiation is: forward callus to the MS minimal medium, callus is divided into young stem;
(3) cultivation is taken root, and the step of yielding positive results is: young stem is inoculated into MS minimal medium+indolebutyric acid IBA0.1-10mgL
-1+ agar 4-10g L
-1In the matrix of+sucrose 20-50g, its pH value is 5.0-6.7, and control tissue culture room cultivation temperature is 23-25 ℃, and illumination is white fluorescent, and luminous intensity is decided to be 65 μ E/m
2/ s 12-20 hour photoperiod, accomplishes in 12-4 hour dark cycle and takes root, yields positive results.
In concrete practice, said step of in test tube, carrying out the tomato tissue culture is:
In described (1) tomato evoked callus incubation step: be placed on behind the described use running water cleaning down that the time of staying is 5 minutes in 5% the Tween-20; With 0.1% mercuric chloride (HgCl
2) the solution surface sterilization time is 4 minutes; Described leaf tissue be cut into magnitude range be about 5mm * 5mm as explant, be inoculated into medium MS minimal medium+6-benzyl aminoadenine (6-BA) 2mgL
-1+ heteroauxin (IAA) 0.2mgL
-1+ agar 6g L
-1In the medium of+sucrose 30g, callus induction;
Cultivation is taken root in described (3), and the medium of the young stem inoculation in the step of yielding positive results is MS minimal medium+indolebutyric acid IBAlmgL
-1+ agar 8g L
-1+ sucrose 30g; Described pH value is 5.8; Described tissue culture room cultivation temperature is 23-25 ℃, and described illumination is white fluorescent, and light intensity is decided to be 65 μ E/m
2/ s, the described photoperiod is 16 hours, described 8 hours dark cycles.
The technology of tomato tissue culture that the present invention is perfect; Promptly can not only in test tube, make the tomato differentiation and proliferation; But also can in test tube, blossom and bear fruit; Because the technology of in test tube, cultivating is difficult, the floristics that in test tube, can blossom and bear fruit in the world at present seldom, this invention has embodied the higher level of tissue culture.
Embodiment
Instance 1
As explant, cleaning down under the running water is placed in 5% the Tween-20 5 minutes then with the blade of tomato variety MicroTom field growing, with the distilled water rinsing of autoclave sterilization 3-4 time, 0.1% mercuric chloride (HgCl
2) surface sterilizing 4 minutes, with the distilled water rinsing of autoclave sterilization 4-5 time.
Leaf tissue be cut into size for about 5mm * 5mm as explant, be inoculated into medium: MS minimal medium+6-benzyl aminoadenine (6-BA) 2mgL
-1+ heteroauxin (IAA) 0.2mgL
-1+ agar 6g L
-1+ sucrose 30g, callus induction 25-30 days.
Forward callus to the MS minimal medium, cultivated 7-10 days, callus is divided into young stem.Young stem is inoculated into MS minimal medium+indolebutyric acid (IBA) 1mgL
-1+ agar 8g L
-1+ sucrose 30g, pH value 5.8,23-25 ℃ of tissue culture room cultivation temperature, white fluorescent light intensity 65 μ E/m
2/ s, 16 hours photoperiod, 8 hours dark cycle.Take root, form bud after 7 days, bloomed in 15 days, 30 days fruit annesls, 45 days fruit maturations.
Instance 2
As explant, cleaning down under the running water is placed in 5% the Tween-20 5 minutes then with the blade of tomato variety Bener Best field growing, with the distilled water rinsing of autoclave sterilization 3-4 time, 0.1% mercuric chloride (HgCl
2) surface sterilizing 4 minutes, with the distilled water rinsing of autoclave sterilization 4-5 time.
Leaf tissue be cut into size for about 5mm * 5mm as explant, be inoculated into medium MS minimal medium+6-benzyl aminoadenine (6-BA) 2mgL
-1+ heteroauxin (IAA) 0.2mgL
-1+ agar 6g L
-1+ sucrose 30g, callus induction 25-30 days.
Forward callus to the MS minimal medium, cultivated 14-20 days, callus is divided into young stem.Young stem is inoculated into MS minimal medium+indolebutyric acid (IBA) 1mgL
-1+ agar 8g L
-1+ sucrose 30g, pH value 5.8,23-25 ℃ of tissue culture room cultivation temperature, illumination is white fluorescent, light intensity 65 μ E/m
2/ s, 16 hours photoperiod, 8 hours dark cycle.Take root after 7 days, formed bud in 14 days, bloomed 40 days fruit annesls, 60 days fruit maturations in 20 days.
Claims (2)
1. a method of in test tube, carrying out the tomato tissue culture is characterized in that;
Said method comprises tomato evoked callus incubation step, breaks up young stem step, and cultivation is taken root and yielded positive results step;
The whole process of said method is all accomplished in test tube, and promptly this method can not only make the tomato differentiation and proliferation in test tube, but also in test tube, accomplishes the group training process of blossoming and bearing fruit;
The tomato method for tissue culture comprises the steps:
(1) described tomato evoked callus incubation step is: get the tomato young leaflet tablet, with stopping in the Tween-20 that is placed on 1%-10% behind the running water cleaning down 2-10 minute, with the distilled water rinsing of autoclave sterilization 3-4 time, use 0.1% mercuric chloride (HgCl then
2) solution surface sterilization 2-10 minute, use distilled water rinsing 4-5 time of autoclave sterilization again; Leaf tissue be cut into big or small areal extent be 1mm * 1mm～10mm * 10mm as explant, be inoculated into MS minimal medium+6-benzyl aminoadenine (6-BA) 0.1-10mgL
-1+ heteroauxin (IAA) 0.01-1mgL
-1+ agar 4-10gL
-1In the medium of+sucrose 20-50g, callus induction;
(2) the young stem step of described differentiation is: forward callus to the MS minimal medium, callus is divided into young stem;
(3) cultivation is taken root, and the step of yielding positive results is: young stem is inoculated into MS minimal medium+indolebutyric acid IBA0.1-10mgL
-1+ agar 4-10gL
-1In the medium of+sucrose 20-50g, its pH value is 5.0-6.7, and control tissue culture room cultivation temperature is 23-25 ℃, and illumination is white fluorescent, and luminous intensity is decided to be 65 μ E/m
2/ s 12-20 hour photoperiod, accomplishes in 12-4 hour dark cycle and takes root, yields positive results.
2. a kind of method of in test tube, carrying out the tomato tissue culture according to claim 1 is characterized in that:
In described (1) tomato evoked callus incubation step: be placed on behind the described use running water cleaning down that the time of staying is 5 minutes in 5% the Tween-20; With 0.1% mercuric chloride (HgCl
2) the solution surface sterilization time is 4 minutes; Described leaf tissue be cut into magnitude range be about 5mm * 5mm as explant, be inoculated into medium MS minimal medium+6-benzyl aminoadenine (6-BA) 2mgL
-1+ heteroauxin (IAA) 0.2mgL
-1+ agar 6gL
-1In the medium of+sucrose 30g, callus induction;
Cultivation is taken root in described (3), and the medium of the young stem inoculation in the step of yielding positive results is MS minimal medium+indolebutyric acid IBA1mgL
-1+ agar 8gL
-1+ sucrose 30g; Described pH value is 5.8; Described tissue culture room cultivation temperature is 23-25 ℃, and described illumination is white fluorescent, and light intensity is decided to be 65 μ E/m
2/ s, the described photoperiod is 16 hours, described 8 hours dark cycles.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2008102259405A CN101731144B (en) | 2008-11-07 | 2008-11-07 | Method for culturing tomato tissues in test tube |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2008102259405A CN101731144B (en) | 2008-11-07 | 2008-11-07 | Method for culturing tomato tissues in test tube |
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| Publication Number | Publication Date |
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| CN101731144A CN101731144A (en) | 2010-06-16 |
| CN101731144B true CN101731144B (en) | 2012-07-18 |
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| CN2008102259405A Expired - Fee Related CN101731144B (en) | 2008-11-07 | 2008-11-07 | Method for culturing tomato tissues in test tube |
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Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102187811B (en) * | 2011-03-21 | 2012-09-05 | 厦门华侨亚热带植物引种园 | Tomato culture medium and tomato culture method |
| CN102283117A (en) * | 2011-06-28 | 2011-12-21 | 淮海工学院 | Sterile culture podding and maturing technique for soybean tissue culture seedlings and transformation seedlings |
| CN103314850B (en) * | 2013-06-02 | 2015-11-04 | 周口师范学院 | A kind of method building base eggplant regenerating system and genetic conversion system in wild-type tomato |
| CN103314849B (en) * | 2013-06-02 | 2016-05-04 | 周口师范学院 | Base eggplant body embryo generation abductive approach in a kind of wild-type tomato |
| CN105145360A (en) * | 2015-09-17 | 2015-12-16 | 福建省农业科学院农业工程技术研究所 | Rapid propagation method for tomato test tube seedling capable of improving proliferation rate |
| CN108184671B (en) * | 2018-03-06 | 2021-08-03 | 山东寿光蔬菜种业集团有限公司 | Method for quickly propagating tomatoes and improving hardening survival rate of tomatoes |
| CN108552058A (en) * | 2018-05-04 | 2018-09-21 | 孟静 | A kind of tomato tissue culture inducing culture and method for tissue culture and application |
| CN110839530A (en) * | 2019-11-25 | 2020-02-28 | 航天神舟生物科技集团有限公司 | Method for inducing flowering of Chinese rose in test tube |
| CN111919753B (en) * | 2020-09-08 | 2022-05-06 | 安徽农业大学 | Culture medium for tomato tissue culture and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101189956A (en) * | 2006-11-23 | 2008-06-04 | 上海市农业科学院园艺研究所 | Method for using tissue culture to separate salt-tolerant tomato |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101189956A (en) * | 2006-11-23 | 2008-06-04 | 上海市农业科学院园艺研究所 | Method for using tissue culture to separate salt-tolerant tomato |
Non-Patent Citations (2)
| Title |
|---|
| 姚宗国等."五彩番茄试管内外快繁技术的研究".《北方园艺》.2002,(第5期),第64-66页. |
| 赵玉萍."试管番茄".《种子世界》.1992,(第4期),第40页. |
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| CN101731144A (en) | 2010-06-16 |
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