CN113711908A - Method for improving survival rate of double haploids - Google Patents
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- 238000000034 method Methods 0.000 title claims abstract description 37
- 230000004083 survival effect Effects 0.000 title claims abstract description 34
- 240000008042 Zea mays Species 0.000 claims abstract description 50
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims abstract description 50
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 39
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims abstract description 34
- 235000005822 corn Nutrition 0.000 claims abstract description 34
- 239000000645 desinfectant Substances 0.000 claims abstract description 22
- 235000016383 Zea mays subsp huehuetenangensis Nutrition 0.000 claims abstract description 16
- 210000000349 chromosome Anatomy 0.000 claims abstract description 16
- 235000009973 maize Nutrition 0.000 claims abstract description 16
- 238000002360 preparation method Methods 0.000 claims abstract description 16
- 238000012258 culturing Methods 0.000 claims abstract description 13
- 241000196324 Embryophyta Species 0.000 claims abstract description 12
- 238000012216 screening Methods 0.000 claims abstract description 9
- 235000015097 nutrients Nutrition 0.000 claims abstract description 8
- 239000002689 soil Substances 0.000 claims abstract description 8
- 238000009331 sowing Methods 0.000 claims abstract description 8
- OSVXSBDYLRYLIG-UHFFFAOYSA-N dioxidochlorine(.) Chemical compound O=Cl=O OSVXSBDYLRYLIG-UHFFFAOYSA-N 0.000 claims description 60
- IAKHMKGGTNLKSZ-INIZCTEOSA-N (S)-colchicine Chemical compound C1([C@@H](NC(C)=O)CC2)=CC(=O)C(OC)=CC=C1C1=C2C=C(OC)C(OC)=C1OC IAKHMKGGTNLKSZ-INIZCTEOSA-N 0.000 claims description 32
- 239000004155 Chlorine dioxide Substances 0.000 claims description 30
- 235000019398 chlorine dioxide Nutrition 0.000 claims description 30
- 229960001338 colchicine Drugs 0.000 claims description 16
- 239000012153 distilled water Substances 0.000 claims description 15
- 238000007789 sealing Methods 0.000 claims description 14
- 238000005406 washing Methods 0.000 claims description 14
- 239000000243 solution Substances 0.000 claims description 13
- 239000003814 drug Substances 0.000 claims description 10
- 241001057636 Dracaena deremensis Species 0.000 claims description 7
- ATJFFYVFTNAWJD-UHFFFAOYSA-N Tin Chemical compound [Sn] ATJFFYVFTNAWJD-UHFFFAOYSA-N 0.000 claims description 7
- 210000002257 embryonic structure Anatomy 0.000 claims description 7
- 239000011259 mixed solution Substances 0.000 claims description 7
- 239000000203 mixture Substances 0.000 claims description 7
- 210000002706 plastid Anatomy 0.000 claims description 7
- 230000010152 pollination Effects 0.000 claims description 7
- 238000012545 processing Methods 0.000 claims description 7
- 241000894007 species Species 0.000 claims description 7
- 238000012360 testing method Methods 0.000 claims description 7
- 238000009395 breeding Methods 0.000 abstract description 19
- 230000001488 breeding effect Effects 0.000 abstract description 18
- 230000035784 germination Effects 0.000 abstract description 5
- 230000009286 beneficial effect Effects 0.000 abstract description 4
- 241000894006 Bacteria Species 0.000 abstract description 3
- 238000005516 engineering process Methods 0.000 description 4
- 229940079593 drug Drugs 0.000 description 3
- 241000233866 Fungi Species 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 230000002363 herbicidal effect Effects 0.000 description 1
- 239000004009 herbicide Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000008186 parthenogenesis Effects 0.000 description 1
- 238000011012 sanitization Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
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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
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/06—Processes for producing mutations, e.g. treatment with chemicals or with radiation
- A01H1/08—Methods for producing changes in chromosome number
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- Developmental Biology & Embryology (AREA)
- Environmental Sciences (AREA)
- Pretreatment Of Seeds And Plants (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a method for improving the survival rate of a doubled haploid, which comprises the following steps: step one, screening corn haploid seeds; step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent; step three, preparation of a disinfectant; step four, putting the stacked fan-shaped filter paper into the prepared solution, taking out the filter paper after a proper amount of water is contained, and putting the filter paper on a tray; step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained; and seventhly, transplanting and managing the maize seedlings with doubled chromosomes. The method for survival rate of the doubled haploid sprouts of the corn can obviously improve the survival rate of the doubled haploid sprouts in the doubling process of the corn haploid, is beneficial to breeding of doubled haploid plants, solves the technical problem that the survival rate of the doubled haploid sprouts is influenced by growth bacteria in the monoploid germination process in the haploid breeding process, and greatly improves the survival rate of the doubled haploid.
Description
Technical Field
The invention relates to the technical field of breeding, in particular to a method for improving the survival rate of doubled haploids.
Background
China is the second largest corn producing country and consuming country, and the improvement of the corn yield and quality requires continuous improvement and development of breeding means. The breeding of the inbred line with high yield, high resistance and high combining ability is a basic and core link in the corn breeding. Haploid breeding refers to a method of utilizing parthenogenesis of a plant to form but replace the haploid, and doubling the chromosome of the plant in a natural or artificial manner to form a homozygous diploid (i.e., a pure line), and then breeding a new variety (line) from the homozygous diploid.
The cultivation of excellent pure lines is a core link of corn breeding. Using traditional breeding methods, it is generally necessary to self-breed 4-6 generations to make the line Write homozygous. And theoretically, even though selfing n generations, the genotype cannot reach a complete homozygous state of 100%. Therefore, it usually takes several decades to breed a variety, consuming a lot of manpower, material resources and financial resources, and therefore shortening the breeding process. The improvement of the breeding efficiency is a target which is always pursued by breeders, and the haploid breeding technology can obtain a homozygous diploid by controlling the hybrid separation and 2 generations, so that the breeding of a selfing line is accelerated, and the breeding period is obviously shortened.
The haploid doubling method is commonly used and comprises a natural doubling method and a drug doubling method. The drug doubling is a currently commonly used haploid doubling method, and the mainly used drugs are colchicine and herbicide. In the colchicine bud soaking method, the growth of fungus is serious in the process of haploid bud germination, so that the survival rate of the doubled haploid is low, and therefore, the improvement of the survival rate of the doubled haploid has important significance for doubling the haploid.
Disclosure of Invention
The present invention aims to provide a method for increasing the survival rate of doubled haploids, so as to solve the problems in the background technology.
In order to achieve the purpose, the invention provides the following technical scheme:
a method for improving the survival rate of double haploids comprises the following steps: step one, screening corn haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.1-0.5% of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is shaken up and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds clean, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tin foil paper, and putting the seeds in an incubator for culture;
step five, processing corn haploid seeds:
(1) cutting off the coleoptile tip of the haploid bud of the corn;
(2) treating the sprouts with colchicine mixture;
pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
As a further scheme of the invention, in the step one, the screening of the corn haploid seeds is to select seeds of purple top colorless embryos to obtain haploid seeds;
as a further aspect of the invention, in step two, the sanitizing agent is formulated to have a concentration of chlorine dioxide of about 0.1 to 0.5% by mass, preferably about 0.4% and 0.5% by mass.
As a further scheme of the invention, in the fourth step, the culture temperature in the incubator is 25-26 ℃ and the culture time is 4 days.
Compared with the prior art, the invention has the beneficial effects that: the method for survival rate of doubled haploid sprouts of corn provided by the invention can obviously improve the survival rate of doubled haploid sprouts in the doubling process of corn haploid, is beneficial to breeding of doubled haploid plants, and solves the technical problem that the survival rate of haploid sprouts is influenced by growth bacteria in the monoploid germination process in the haploid breeding process, thereby greatly improving the survival rate of doubled haploid and making the large-scale application of haploid doubling technology more likely.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1 a method for increasing the survival rate of doubled haploids, comprising the steps of: step one, screening corn haploid seeds:
selecting seeds with purple tops and colorless embryos to obtain haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.5 percent of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is uniformly shaken and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds cleanly, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tinfoil paper, putting the seeds in an incubator, and culturing for 4 days at the temperature of 25-26 ℃;
step five, processing corn haploid seeds:
(1) coleoptile tip for cutting haploid young bud of corn
(2) Treating the sprouts with colchicine mixture
Pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
Example 2 a method for increasing the survival rate of doubled haploids, comprising the steps of: step one, screening corn haploid seeds:
selecting seeds with purple tops and colorless embryos to obtain haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.4 percent of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is uniformly shaken and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds cleanly, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tinfoil paper, putting the seeds in an incubator, and culturing for 4 days at the temperature of 25-26 ℃;
step five, processing corn haploid seeds:
(1) coleoptile tip for cutting haploid young bud of corn
(2) Treating the sprouts with colchicine mixture
Pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
Example 3 a method for increasing the survival rate of doubled haploids, comprising the steps of: step one, screening corn haploid seeds:
selecting seeds with purple tops and colorless embryos to obtain haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.3 percent of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is uniformly shaken and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds cleanly, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tinfoil paper, putting the seeds in an incubator, and culturing for 4 days at the temperature of 25-26 ℃;
step five, processing corn haploid seeds:
(1) coleoptile tip for cutting haploid young bud of corn
(2) Treating the sprouts with colchicine mixture
Pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
Example 4 a method for increasing the survival rate of doubled haploids, comprising the steps of: step one, screening corn haploid seeds:
selecting seeds with purple tops and colorless embryos to obtain haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.2 percent of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is uniformly shaken and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds cleanly, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tinfoil paper, putting the seeds in an incubator, and culturing for 4 days at the temperature of 25-26 ℃;
step five, processing corn haploid seeds:
(1) coleoptile tip for cutting haploid young bud of corn
(2) Treating the sprouts with colchicine mixture
Pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
Example 5 a method for increasing the survival rate of doubled haploids, comprising the steps of: step one, screening corn haploid seeds:
selecting seeds with purple tops and colorless embryos to obtain haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.1 percent of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is uniformly shaken and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds cleanly, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tinfoil paper, putting the seeds in an incubator, and culturing for 4 days at the temperature of 25-26 ℃;
step five, processing corn haploid seeds:
(1) coleoptile tip for cutting haploid young bud of corn
(2) Treating the sprouts with colchicine mixture
Pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
Control group: the procedure is exactly the same as in the example except that distilled water is used instead of the disinfectant in the third step.
The results of the survival rate of the haploids germinated by the haploid seeds after the disinfectant solution with different concentrations and the water treatment are as follows:
table 1 shows the survival rate of the haploids germinated on the seeds treated by disinfectant solutions of different concentrations
| Experimental group | Total number of haploid seeds | Concentration of disinfectant solution% | Survival rate of sprout% |
| Example 1 | 600 | 0.5 | 93 |
| Example 2 | 600 | 0.4 | 92 |
| Example 3 | 600 | 0.3 | 80 |
| Example 4 | 600 | 0.2 | 70 |
| Example 5 | 600 | 0.1 | 65 |
| Control group | 600 | Water (W) | 58 |
As can be seen from Table 1, the disinfectant solution has the effect of inhibiting the growth of fungi in the germination process of the corn haploid seeds, so that the survival rate of the sprouts is increased, and the survival rate of the doubled haploid is increased. The disinfectant solutions vary in concentration to inhibit fungal growth, with concentrations of 0.5% or 0.4% being most effective. The method for survival rate of doubled haploid sprouts of corn provided by the invention can obviously improve the survival rate of doubled haploid sprouts in the doubling process of corn haploid, is beneficial to breeding of doubled haploid plants, and solves the technical problem that the survival rate of haploid sprouts is influenced by growth bacteria in the monoploid germination process in the haploid breeding process, thereby greatly improving the survival rate of doubled haploid and making the large-scale application of haploid doubling technology more likely.
It will be evident to those skilled in the art that the invention is not limited to the details of the foregoing illustrative embodiments, and that the present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof. The present embodiments are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims rather than by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.
Furthermore, it should be understood that although the present description refers to embodiments, not every embodiment may contain only a single embodiment, and such description is for clarity only, and those skilled in the art should integrate the description, and the embodiments may be combined as appropriate to form other embodiments understood by those skilled in the art.
Claims (4)
1. A method for improving the survival rate of double haploids is characterized by comprising the following steps: step one, screening corn haploid seeds;
step two, preparation of the germinating paper: folding the purchased filter paper into a fan shape with proper size, wherein the size is consistent;
step three, preparation of the disinfectant: the mass concentration of the prepared disinfectant is about 0.1-0.5% of chlorine dioxide, the dosage of the medicament and distilled water is calculated according to the above concentration, the chlorine dioxide is firstly added into a beaker, the distilled water is added into the beaker until the chlorine dioxide is completely dissolved, and the chlorine dioxide is shaken up and then stored in a plastic container;
fourthly, putting the stacked fan-shaped filter paper into the prepared solution, taking out the stacked fan-shaped filter paper after a proper amount of water is removed, putting the stacked fan-shaped filter paper on a tray, washing haploid seeds clean, uniformly putting the seeds at the bottom of the fan-shaped filter paper with a proper amount of water at intervals of 1 cm-1.5 cm, sealing the seeds with tin foil paper, and putting the seeds in an incubator for culture;
step five, processing corn haploid seeds:
(1) cutting off the coleoptile tip of the haploid bud of the corn;
(2) treating the sprouts with colchicine mixture;
pouring the prepared colchicine mixed solution into a container, completely immersing all cut buds, sealing the opening of the container, and treating for 12 hours at 23 ℃ in a dark place to obtain maize buds with doubled chromosomes;
(3) washing the soaked haploid seeds with clear water for 1-2 hours to obtain pretreated haploid seeds;
step six, sowing the obtained pretreated haploid seeds in nutrient soil, watering, and culturing in a sun-shading mode until three leaves and one heart are obtained;
seventhly, transplanting and managing the maize seedlings with doubled chromosomes:
transplanting the Wang Lai seedlings with doubled chromoplasts cultivated in the sixth step to a testing station of Hebei Ji crop species company Limited responsibility, shading the seedlings for 5 days by using a shading net, finely managing the seedlings in the field to ensure the normal growth of the seedlings, removing non-haploid plants in the seedling stage, and carrying out artificial pollination to obtain the haploid corn plants.
2. The method for improving doubled haploid survival rate of claim 1, wherein in step one, the selection of maize haploid seeds is to select seeds with purple top colorless embryos to obtain haploid seeds.
3. The method for improving doubled haploid survival rate of claim 1, wherein in step two, the disinfectant is formulated with chlorine dioxide at a mass concentration of about 0.1-0.5%, preferably at a mass concentration of about 0.4% and 0.5%.
4. The method for improving doubled haploid survival rate of claim 1, wherein in step four, the cultivation temperature in the incubator is 25-26 degrees, and the cultivation time is 4 days.
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| CN112568077A (en) * | 2020-12-14 | 2021-03-30 | 吉林省农业科学院 | Method for improving doubling efficiency of corn haploid |
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| 程锦等: "气体二氧化氯对玉米表面细菌杀菌", 《粮食与油脂》 * |
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Application publication date: 20211130 |