CN113068612A - Method for improving tissue culture germination rate of strawberry seeds - Google Patents
Method for improving tissue culture germination rate of strawberry seeds Download PDFInfo
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- CN113068612A CN113068612A CN202110385161.7A CN202110385161A CN113068612A CN 113068612 A CN113068612 A CN 113068612A CN 202110385161 A CN202110385161 A CN 202110385161A CN 113068612 A CN113068612 A CN 113068612A
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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
- A01H4/001—Culture apparatus for tissue culture
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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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- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a method for improving tissue culture germination rate of strawberry seeds, belonging to the field of tissue culture of wild strawberry seeds. The invention can improve the tissue culture germination rate of wild strawberry seeds from 0 to more than 70 percent, realizes high germination rate, the natural germination rate of the strawberry seeds is only 30 to 40 percent, and the method greatly exceeds the natural germination rate of the strawberry seeds. Through disinfecting and disinfecting the seed, can effectually guarantee the growing environment of seed, realized awakening up wild strawberry seed tissue at the in-process that disinfects simultaneously to through carrying out physical method to the seed coat and handling, broken the physiological dormancy of seed, great improvement the germination percentage of strawberry seed.
Description
Technical Field
The invention relates to the field of tissue culture of wild strawberry seeds, in particular to a method for improving tissue culture germination rate of strawberry seeds.
Background
Strawberry belongs to the genus strawberry of the family Rosaceae, is a perennial evergreen herbaceous plant, and belongs to berry fruit trees in horticulture. The strawberry is fresh, tender and succulent, sweet and sour, unique in flavor and rich in nutrition, and compared with other fruit trees, the strawberry cultivation method is short in cultivation period, suitable for facility cultivation and high in economic benefit. The continuous improvement and updating of strawberry varieties are the foundation of the development of the world strawberry industry, and the strawberry varieties bred at home and abroad are mainly obtained by breeding the seedling progeny obtained from the hybrid seeds through the conventional cross breeding means. Sowing seeds and obtaining seedlings are one of the important links in the current strawberry breeding work. The wild strawberry has partial excellent characteristics, and the wild strawberry needs to be cultivated, but in the cultivation process, the germination rate of the wild strawberry seeds seriously influences the cultivation research, and the natural germination rate of the existing wild strawberry is only 5-10%, so that a method for improving the germination rate of the wild strawberry seeds needs to be researched.
Disclosure of Invention
The invention aims to provide a method for improving tissue culture germination rate of strawberry seeds, and solves the technical problem of low germination rate of wild strawberry seeds in the prior art.
A method for improving tissue culture germination rate of strawberry seeds comprises the following steps:
step 1: placing wild strawberry seeds in an open container, pouring distilled water to submerge the seeds, and placing the container in a refrigerator at 4 ℃ for standing for 2 d;
step 2: preparing a culture medium, placing the culture medium in a sterilization pot at 118 ℃ for sterilization for 30min for later use, wrapping fine iron sand cloth, tweezers, a culture dish and filter paper with newspaper tightly, and sterilizing together with the culture medium;
and step 3: filtering out the soaked wild strawberry seeds from the container, placing on a filter paper to absorb water, placing on a super-clean workbench together with gauze, tweezers, a culture dish, the filter paper, a sodium hypochlorite solution and instruments required by a culture medium, and sterilizing for 30min by opening an ultraviolet lamp;
and 4, step 4: putting wild strawberry seeds into a culture dish, pouring a sterilizing liquid to perform secondary sterilization treatment on the wild strawberry seeds, repeatedly cleaning the wild strawberry seeds subjected to secondary sterilization with sterile water, and putting the wild strawberry seeds on filter paper to absorb water;
and 5: treating the skin of the wild strawberry seeds until the skin becomes one fifth of the original skin;
step 6: and (3) inoculating the treated wild strawberry seeds into an MS culture medium by using forceps, sealing the culture medium, and then putting the culture medium into a culture room or an illumination incubator for culture.
Further, in the step 2, the culture medium comprises 33g/LNH4NO3、7.4g/L MgSO4、2.1g/LZnSO4、1g/L CoCl20.83g/LKI, 0.3g/L nicotinic acid, 0.1g/L thiamine hydrochloride, 2g/L glycine, 5g sucrose and 0.57g/L kanamycin.
Further, in the step 4, the sterilizing solution is sodium hypochlorite with the concentration of 1%, the sodium hypochlorite is soaked for 20min, the sodium hypochlorite is repeatedly washed for 4 to 5 times by using sterile water, and then the sodium hypochlorite is placed on filter paper to suck water.
Further, in the step 4, the sterilizing solution is a mixed solution of 70% ethanol, 1% mercury and water, soaking for 20-25min, then repeatedly cleaning with clear water until the mercury on the surface is completely cleaned, and then placing on filter paper to suck water.
Further, in the step 5, the seed coat treatment process includes uniformly spreading wild strawberry seeds on a flat sand paper in a single layer, covering another sand paper, covering a layer of sand above the sand paper, wherein the pressure of the sand on the wild strawberry seeds is 0.1-0.08N, fixing four sides of the upper sand paper, dragging 8-10 of the sand paper at the bottom back and forth, rolling the wild strawberry seeds between the two sand papers, fixing four sides of the sand paper at the bottom back and forth, dragging 5-8 of the upper sand paper back and forth, moving the wild strawberry seeds between the two sand cloths, pulling the upper sand cloth by two hands, pressing the upper sand cloth downwards, and dragging the wild strawberry seeds back and forth for 14-17 times, wherein the pressure is 0.1-0.2N.
Further, in the step 5, wild strawberry seeds are soaked in sulfuric acid with the concentration of 18.4mol/L, the ratio of the wild strawberry seeds to the sulfuric acid is 1:0.4, the soaking time is 1-4min, and during soaking, a glass rod is used for stirring.
By adopting the technical scheme, the invention has the following technical effects:
the invention can improve the tissue culture germination rate of wild strawberry seeds from 0 to more than 70 percent, realizes high germination rate, the natural germination rate of the strawberry seeds is only 30 to 40 percent, and the method greatly exceeds the natural germination rate of the strawberry seeds. Through disinfecting and disinfecting the seed, can effectually guarantee the growing environment of seed, realized awakening up wild strawberry seed tissue at the in-process that disinfects simultaneously to through carrying out physical method to the seed coat and handling, broken the physiological dormancy of seed, great improvement the germination percentage of strawberry seed.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, preferred embodiments are given and the present invention is described in further detail. It should be noted, however, that the numerous details set forth in the description are merely for the purpose of providing the reader with a thorough understanding of one or more aspects of the present invention, which may be practiced without these specific details.
Example 1:
a method for improving tissue culture germination rate of strawberry seeds comprises the following steps:
step 1: placing wild strawberry seeds in an open container, pouring distilled water to submerge the seeds, and placing the container in a refrigerator at 4 ℃ for standing for 2 d.
Step 2: preparing a culture medium, placing the culture medium in a sterilization pot at 118 ℃ for sterilization for 30min for standby, wrapping fine iron sand cloth, tweezers, a culture dish and filter paper with newspaper tightly, and sterilizing together with the culture medium.
The culture medium comprises 33g/LNH4NO3、7.4g/L MgSO4、2.1g/L ZnSO4、1g/L CoCl20.83g/LKI, 0.3g/L nicotinic acid, 0.1g/L thiamine hydrochloride, 2g/L glycine, 5g sucrose and 0.57g/L kanamycin.
And step 3: filtering out soaked wild strawberry seeds from the container, placing on filter paper to absorb water, placing on a super clean bench together with gauze, tweezers, culture dish, filter paper, sodium hypochlorite solution, and culture medium, and sterilizing with ultraviolet lamp for 30 min.
And 4, step 4: putting wild strawberry seeds into a culture dish, adding sodium hypochlorite with the concentration of 1%, soaking for 20min, repeatedly washing with sterile water for 4-5 times, and then placing on filter paper to suck water.
And 5: the method comprises the steps of uniformly spreading wild strawberry seeds on a flat abrasive paper in a single layer, covering another piece of abrasive paper, covering a layer of sand above the abrasive paper, wherein the pressure of the sand on the wild strawberry seeds is 0.1-0.08N, fixing the four sides of the upper piece of abrasive paper, dragging 8-10 of the bottom abrasive paper back and forth, rolling the wild strawberry seeds between the two pieces of abrasive paper, fixing the four sides of the bottom abrasive paper, dragging 5-8 of the upper piece of abrasive paper back and forth, moving the wild strawberry seeds between the two pieces of abrasive paper, pulling the upper piece of abrasive paper by two hands, pressing the upper piece of abrasive paper downwards, controlling the pressure to be 0.1-0.2N, and dragging the wild strawberry seeds back and forth 14-17 times. Until the seed coat becomes one fifth of the original one.
Step 6: and (3) inoculating the treated wild strawberry seeds into an MS culture medium by using forceps, sealing the culture medium, and then putting the culture medium into a culture room or an illumination incubator for culture.
After 8-10 days of culture, the statistical germination percentage is 70.3%.
Example 2:
a method for improving tissue culture germination rate of strawberry seeds comprises the following steps:
step 1: placing wild strawberry seeds in an open container, pouring distilled water to submerge the seeds, and placing the container in a refrigerator at 4 ℃ for standing for 2 d.
Step 2: preparing a culture medium, placing the culture medium in a sterilization pot at 118 ℃ for sterilization for 30min for standby, wrapping fine iron sand cloth, tweezers, a culture dish and filter paper with newspaper tightly, and sterilizing together with the culture medium.
The culture medium comprises 33g/LNH4NO3、7.4g/L MgSO4、2.1g/L ZnSO4、1g/L CoCl20.83g/LKI, 0.3g/L nicotinic acid, 0.1g/L thiamine hydrochloride, 2g/L glycine, 5g sucrose and 0.57g/L kanamycin.
And step 3: filtering out soaked wild strawberry seeds from the container, placing on filter paper to absorb water, placing on a super clean bench together with gauze, tweezers, culture dish, filter paper, sodium hypochlorite solution, and culture medium, and sterilizing with ultraviolet lamp for 30 min.
And 4, step 4: putting wild strawberry seeds into a culture dish, pouring a mixed solution of 70% ethanol, 1% mercury and water, soaking for 20-25min, then repeatedly cleaning with clear water until mercury on the surface is completely cleaned, and then placing on filter paper to suck water.
And 5: treating the skins of wild strawberry seeds, wherein the wild strawberry seeds are soaked in sulfuric acid with the concentration of 18.4mol/L, the ratio of the wild strawberry seeds to the sulfuric acid is 1:0.4, the soaking time is 1-4min, and during soaking, a glass rod is used for stirring until the skins are one fifth of the original skins.
Step 6: and (3) inoculating the treated wild strawberry seeds into an MS culture medium by using forceps, sealing the culture medium, and then putting the culture medium into a culture room or an illumination incubator for culture.
After 8-10 days of culture, the statistical germination percentage is 73.4%.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that those skilled in the art can make various improvements and modifications without departing from the principle of the present invention, and these improvements and modifications should also be construed as the protection scope of the present invention.
Claims (6)
1. A method for improving tissue culture germination rate of strawberry seeds is characterized by comprising the following steps: the method comprises the following steps:
step 1: placing wild strawberry seeds in an open container, pouring distilled water to submerge the seeds, and placing the container in a refrigerator at 4 ℃ for standing for 2 d;
step 2: preparing an MS culture medium, placing the culture medium in a sterilization pot at 118 ℃ for sterilization for 30min for later use, wrapping fine iron sand cloth, tweezers, a culture dish and filter paper with newspaper tightly, and sterilizing together with the culture medium;
and step 3: filtering out the soaked wild strawberry seeds from the container, placing on a filter paper to absorb water, placing on a super-clean workbench together with gauze, tweezers, a culture dish, the filter paper, a sodium hypochlorite solution and instruments required by a culture medium, and sterilizing for 30min by opening an ultraviolet lamp;
and 4, step 4: putting wild strawberry seeds into a culture dish, pouring a sterilizing liquid to perform secondary sterilization treatment on the wild strawberry seeds, repeatedly cleaning the wild strawberry seeds subjected to secondary sterilization with sterile water, and putting the wild strawberry seeds on filter paper to absorb water;
and 5: treating the skin of the wild strawberry seeds until the skin becomes one fifth of the original skin;
step 6: and (3) inoculating the treated wild strawberry seeds into an MS culture medium by using forceps, sealing the culture medium, and then putting the culture medium into a culture room or an illumination incubator for culture.
2. The method for improving tissue culture germination rate of strawberry seeds according to claim 1, wherein: in the step 2, the culture medium comprises 33g/LNH4NO3、7.4g/L MgSO4、2.1g/LZnSO4、1g/L CoCl20.83g/LKI, 0.3g/L nicotinic acid, 0.1g/L thiamine hydrochloride, 2g/L glycine, 5g sucrose and 0.57g/L kanamycin.
3. The method for improving tissue culture germination rate of strawberry seeds according to claim 2, wherein: in the step 4, the sterilizing solution is sodium hypochlorite with the concentration of 1%, the sodium hypochlorite is soaked for 20min, and the sodium hypochlorite is repeatedly washed for 4-5 times by using sterile water and then placed on filter paper to absorb water.
4. The method for improving tissue culture germination rate of strawberry seeds according to claim 2, wherein: in the step 4, the sterilizing solution is a mixed solution of 70% ethanol, 1% mercury and water, soaking for 20-25min, then repeatedly cleaning with clear water until the mercury on the surface is completely cleaned, and then placing on filter paper to absorb water.
5. The method for improving tissue culture germination rate of strawberry seeds according to claim 4, wherein: in the step 5, the physical treatment process of the seed coat comprises the steps of uniformly spreading wild strawberry seeds on a flat sand paper in a single layer, covering another piece of sand paper, covering a layer of sand above the sand paper, wherein the pressure of the sand on the wild strawberry seeds is 0.1-0.08N, fixing the four sides of the upper piece of sand paper, dragging 8-10 of the sand paper at the bottom back and forth, rolling the wild strawberry seeds between the two pieces of sand paper, fixing the four sides of the bottom sand paper, dragging 5-8 of the upper piece of sand paper back and forth, moving the wild strawberry seeds between the two pieces of sand paper, pulling the upper piece of sand paper by two hands, pressing the sand paper downwards, wherein the pressure is 0.1-0.2N, and dragging the sand paper back and forth for 14-17 times.
6. The method for improving tissue culture germination rate of strawberry seeds according to claim 4, wherein: in the step 5, wild strawberry seeds are soaked in sulfuric acid with the concentration of 18.4mol/L, the ratio of the wild strawberry seeds to the sulfuric acid is 1:0.4, the soaking time is 1-4min, and a glass rod is used for stirring during soaking.
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| CN202110385161.7A CN113068612A (en) | 2021-04-09 | 2021-04-09 | Method for improving tissue culture germination rate of strawberry seeds |
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| CN202110385161.7A CN113068612A (en) | 2021-04-09 | 2021-04-09 | Method for improving tissue culture germination rate of strawberry seeds |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102626049A (en) * | 2012-04-02 | 2012-08-08 | 常州亚环环保科技有限公司 | Cultivation method of black strawberries |
| CN104195171A (en) * | 2014-09-16 | 2014-12-10 | 江苏农林职业技术学院 | Efficient, rapid and stable genetic transformation method for strawberries |
| US20150140599A1 (en) * | 2006-12-11 | 2015-05-21 | Japan Science And Technology Agency | Plant growth regulator and use thereof |
| CN105191633A (en) * | 2015-09-29 | 2015-12-30 | 淮北市四季蔬菜种植有限公司 | Seedling culturing method capable of raising emergence rate of strawberry seeds |
-
2021
- 2021-04-09 CN CN202110385161.7A patent/CN113068612A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150140599A1 (en) * | 2006-12-11 | 2015-05-21 | Japan Science And Technology Agency | Plant growth regulator and use thereof |
| CN102626049A (en) * | 2012-04-02 | 2012-08-08 | 常州亚环环保科技有限公司 | Cultivation method of black strawberries |
| CN104195171A (en) * | 2014-09-16 | 2014-12-10 | 江苏农林职业技术学院 | Efficient, rapid and stable genetic transformation method for strawberries |
| CN105191633A (en) * | 2015-09-29 | 2015-12-30 | 淮北市四季蔬菜种植有限公司 | Seedling culturing method capable of raising emergence rate of strawberry seeds |
Non-Patent Citations (4)
| Title |
|---|
| 李好琢等: ""提高草莓种子离体培养发芽率的研究"", 《北方果树》 * |
| 王小敏: "《植物生命活动规律及其机理研究》", 31 March 2019, 电子科技大学出版社 * |
| 陈世昌等: "《植物组织培养 第3版》", 30 June 2016, 重庆大学出版社 * |
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