CN112806260A - Tissue culture seedling raising method for baby cabbage with high disease resistance - Google Patents
Tissue culture seedling raising method for baby cabbage with high disease resistance Download PDFInfo
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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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- 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
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Abstract
The invention discloses a tissue culture seedling method of baby cabbage with high disease resistance, which comprises the following steps of 1) taking materials, 2) cultivating, 3) differentiating, 4) rooting and seedling strengthening cultivating, 5) carrying out proliferation cultivating, 6) carrying out rooting cultivating, 7) heeling in, hardening rooted test-tube seedlings in a greenhouse for 5-7 d, washing the rooted test-tube seedlings with water to remove residual culture medium at the root, and heeling in on a heeling root substrate; after 15-20 days, new roots grow out; the invention improves the disease resistance of the baby cabbage, can effectively resist the pathogen invasion of the baby cabbage, enhances the disease resistance advantage of the baby cabbage, effectively improves the survival rate of planting and improves the economic benefit.
Description
Technical Field
The invention relates to the technical field of planting, in particular to a tissue culture seedling method for baby cabbage with high disease resistance.
Background
Baby cabbage is a pocket-sized small-plant Chinese cabbage, and belongs to Brassica subspecies of Brassicaceae. The general growth period is 45-55 days, the height of the commodity ball is 20 cm, the diameter is 8-9 cm, and the net weight of the vegetable is about 150-200 g. The growth temperature is proper at 5-25 ℃, and the temperature is lower than 5 ℃, so that the balls are easily frozen and loosened or cannot be held; above 25 ℃ it is susceptible to viral diseases. The baby vegetable sides are thin, sweet and tender and have delicious taste. However, due to the limitation of local climatic conditions, farmers apply fertilizers blindly, so that the physiological problems of the baby cabbage are frequent, and the quality and the yield of the baby cabbage are influenced; moreover, farmers apply fertilizer blindly, which results in low fertilizer utilization rate and serious soil hardening. An efficient and high-quality cultivation method is urgently needed to improve the local planting technical level. .
Disclosure of Invention
The invention provides a tissue culture seedling method of baby cabbage with high disease resistance.
The scheme of the invention is as follows:
a tissue culture seedling method of giant salamander with high disease resistance comprises the following steps:
1) taking materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into (1.0-1.4) cm x (1.0-1.4) cm square blocks;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
As a preferred technical scheme, the detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing and detoxification are taken out and placed in a soaking solution at the temperature of 18-20 ℃ for soaking for 40-50 min.
Preferably, the washing solution comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the adding amount ratio is 95: 1:1:0.8: 0.6.
according to a preferable technical scheme, the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis chinensis extracting solution, ruyi grass and distilled water, wherein the feeding ratio is 15-20: 2-3: 20-30: 1-3: 1-3: 2-4: 40 to 50.
As a preferable technical scheme, the infectious bacterium liquid in the step 2) is prepared by placing the baby cabbage leaves which are just infected with pathogenic bacteria in a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass of the diseased leaves, soaking at 30-35 ℃ for 30-45 min, filtering to remove impurities, adding 20 times of water to dilute the filtrate, adding 1 time of treating agent with the mass fraction of the diseased leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterium liquid.
As a preferred technical scheme, the infectious germs comprise downy mildew and soft rot.
According to a preferable technical scheme, the treating agent comprises folium artemisiae argyi extract, honeysuckle extract, fructus forsythiae extract, herba houttuyniae extract and distilled water.
According to a preferable technical scheme, the adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the forsythia extract, the houttuynia cordata extract to distilled water is 2-3: 2-5: 1-3: 3-7: 85-96.
As a preferred technical scheme, the minimal medium comprises the following components in the following concentration: MS, 0.3mg/L BA, 0.3mg/L KT.
The differentiation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA and 1500mg/L colchicine.
The rooting and seedling strengthening culture medium comprises the following components in the following concentration: MS, 0.1mg/L IAA and 100g/L banana puree.
The proliferation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA.
The rooting medium comprises the following components in the following concentrations: MS, 0.1mg/L NAA.
The temporary planting substrate is prepared by perlite, peat, humus and soil according to the volume ratio of 2: 3: 1.
Due to the adoption of the technical scheme, the tissue culture seedling method for the baby cabbage with high disease resistance comprises the steps of 1) obtaining materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the filter paper is sucked to be dry, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into small blocks of (1.0-1.4) cm x (1.0-1.4) cm; 2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus; 3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds; 4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system; 5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity; 6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution; 7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
The invention has the following advantages:
the disease resistance of the baby cabbage is improved, the pathogen invasion of the baby cabbage can be effectively resisted, the disease resistance advantage of the baby cabbage is enhanced, the planting survival rate is effectively improved, and the economic benefit is improved.
After experimental detection, the cabbage seedling has obvious advantages in disease resistance, cold resistance, physiological shape and yield compared with other varieties, the yield of the cabbage is increased, and the quality of the cabbage is also improved.
Detailed Description
In order to make up for the defects, the invention provides a tissue culture seedling method of baby cabbage with high disease resistance, which aims to solve the problems in the background technology.
A tissue culture seedling method of giant salamander with high disease resistance comprises the following steps:
1) taking materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into (1.0-1.4) cm x (1.0-1.4) cm square blocks;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
The detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing liquid is taken out and is soaked in a soaking solution at the temperature of 18-20 ℃ for 40-50 min.
Preferably, the washing solution comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the adding amount ratio is 95: 1:1:0.8: 0.6.
the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis chinensis extracting solution, ruyi grass and distilled water, wherein the feed ratio is 15-20: 2-3: 20-30: 1-3: 1-3: 2-4: 40 to 50.
And 2) placing the Mesorethrus mollissima leaves just infected with pathogenic bacteria into a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass fraction of the pathogenic leaves, soaking at 30-35 ℃ for 30-45 min, filtering to remove impurities, adding 20 times of water into the filtrate for dilution, adding 1 time of treating agent with the mass fraction of the pathogenic leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterial liquid.
The infectious germs comprise downy mildew and soft rot.
The treating agent comprises folium Artemisiae Argyi extract, flos Lonicerae extract, fructus forsythiae extract, herba Houttuyniae extract and distilled water.
The adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the fructus forsythiae extract, the houttuynia cordata extract to the distilled water is 2-3: 2-5: 1-3: 3-7: 85-96.
The minimal medium comprises the following components in the following concentrations: MS, 0.3mg/L BA, 0.3mg/L KT.
The differentiation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA and 1500mg/L colchicine.
The rooting and seedling strengthening culture medium comprises the following components in the following concentration: MS, 0.1mg/L IAA and 100g/L banana puree.
The proliferation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA.
The rooting medium comprises the following components in the following concentrations: MS, 0.1mg/L NAA.
The temporary planting substrate is prepared by perlite, peat, humus and soil according to the volume ratio of 2: 3: 1.
In order to make the technical means, the creation characteristics, the achievement purposes and the effects of the invention easy to understand, the invention is further described with the specific embodiments.
Example 1:
1) taking materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into (1.0-1.4) cm x (1.0-1.4) cm square blocks;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
The detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing liquid is taken out and is soaked in a soaking solution at the temperature of 18-20 ℃ for 40-50 min.
Preferably, the washing solution comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the adding amount ratio is 95: 1:1:0.8: 0.6.
the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis root extracting solution, ruyi grass and distilled water, wherein the feed ratio is 15: 2: 20:1: 1: 2: 40.
and 2) placing the Mesorethrus mollissima leaves just infected with pathogenic bacteria into a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass fraction of the pathogenic leaves, soaking at 30-35 ℃ for 30-45 min, filtering to remove impurities, adding 20 times of water into the filtrate for dilution, adding 1 time of treating agent with the mass fraction of the pathogenic leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterial liquid.
The infectious germs comprise downy mildew and soft rot.
The treating agent comprises folium Artemisiae Argyi extract, flos Lonicerae extract, fructus forsythiae extract, herba Houttuyniae extract and distilled water.
The adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the fructus forsythiae extract, the houttuynia cordata extract to the distilled water is 2: 2:1:3: 85.
the minimal medium comprises the following components in the following concentrations: MS, 0.3mg/L BA, 0.3mg/L KT.
The differentiation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA and 1500mg/L colchicine.
The rooting and seedling strengthening culture medium comprises the following components in the following concentration: MS, 0.1mg/L IAA and 100g/L banana puree.
The proliferation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA.
The rooting medium comprises the following components in the following concentrations: MS, 0.1mg/L NAA.
The temporary planting substrate is prepared by perlite, peat, humus and soil according to the volume ratio of 2: 3: 1.
Example 2:
1) taking materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into (1.0-1.4) cm x (1.0-1.4) cm square blocks;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
The detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing liquid is taken out and is soaked in a soaking solution at the temperature of 18-20 ℃ for 40-50 min.
Preferably, the washing solution comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the adding amount ratio is 95: 1:1:0.8: 0.6.
the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis root extracting solution, ruyi grass and distilled water, wherein the feed ratio is 20: 3: 30:3: 3: 4: 50.
and 2) placing the Mesorethrus mollissima leaves just infected with pathogenic bacteria into a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass fraction of the pathogenic leaves, soaking at 30-35 ℃ for 30-45 min, filtering to remove impurities, adding 20 times of water into the filtrate for dilution, adding 1 time of treating agent with the mass fraction of the pathogenic leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterial liquid.
The infectious germs comprise downy mildew and soft rot.
The treating agent comprises folium Artemisiae Argyi extract, flos Lonicerae extract, fructus forsythiae extract, herba Houttuyniae extract and distilled water.
The adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the fructus forsythiae extract, the houttuynia cordata extract to the distilled water is 3: 5:3:7: 96.
the minimal medium comprises the following components in the following concentrations: MS, 0.3mg/L BA, 0.3mg/L KT.
The differentiation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA and 1500mg/L colchicine.
The rooting and seedling strengthening culture medium comprises the following components in the following concentration: MS, 0.1mg/L IAA and 100g/L banana puree.
The proliferation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA.
The rooting medium comprises the following components in the following concentrations: MS, 0.1mg/L NAA.
The temporary planting substrate is prepared by perlite, peat, humus and soil according to the volume ratio of 2: 3: 1.
Example 3:
1) taking materials, washing well-grown baby cabbage leaves with running water for 40min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into small squares of (1.0-1.4) cm x (1.0-1.4) cm;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.6ml for the first time, spraying 2ml for the second time, culturing in a culture chamber, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove the residual culture medium at the roots, and heeling in the rooting matrix; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
The detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing liquid is taken out and is soaked in a soaking solution at the temperature of 18-20 ℃ for 40-50 min.
Preferably, the washing solution comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the adding amount ratio is 95: 1:1:0.8: 0.6.
the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis root extracting solution, ruyi grass and distilled water, wherein the feed ratio is 17: 2: 25:2: 2: 3: 45.
and 2) placing the Mesorethrus mollissima leaves just infected with pathogenic bacteria into a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass fraction of the diseased leaves, soaking at 30-35 ℃ for 35min, filtering to remove impurities, adding 20 times of water into the filtrate for dilution, adding 1 time of treating agent with the mass fraction of the diseased leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterial liquid.
The infectious germs comprise downy mildew and soft rot.
The treating agent comprises folium Artemisiae Argyi extract, flos Lonicerae extract, fructus forsythiae extract, herba Houttuyniae extract and distilled water.
The adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the fructus forsythiae extract, the houttuynia cordata extract to the distilled water is 3: 3:2:4: 87.
the minimal medium comprises the following components in the following concentrations: MS, 0.3mg/L BA, 0.3mg/L KT.
The differentiation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA and 1500mg/L colchicine.
The rooting and seedling strengthening culture medium comprises the following components in the following concentration: MS, 0.1mg/L IAA and 100g/L banana puree.
The proliferation medium included the following components at the following concentrations: MS, 0.4mg/L BA, 0.1mg/L IAA.
The rooting medium comprises the following components in the following concentrations: MS, 0.1mg/L NAA.
The temporary planting substrate is prepared by perlite, peat, humus and soil according to the volume ratio of 2: 3: 1.
The seedling plant of the embodiment 3 is planted, the yield is improved by more than 7 percent compared with the conventional yield, and the economic benefit is improved.
Identifying disease resistance of plants:
60 seedlings of example 3 were used as a test group, 60 seedlings were obtained by a conventional method as a control group, 30 seedlings of each of the test group and the control group were inoculated to downy mildew, and 30 seedlings of each of the test group and the control group were inoculated to soft rot, and the cultivation was continued.
The incidence of downy mildew in the test group is 8%, the incidence of soft rot is 6%, while the incidence of downy mildew in the control group is 35% and the incidence of soft rot is 48%.
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (8)
1. A tissue culture seedling method of giant salamander with high disease resistance is characterized by comprising the following steps:
1) taking materials, washing well-grown baby cabbage leaves with running water for 30-60 min, after the leaves are sucked dry by filter paper, disinfecting with 85% alcohol for 2-3S, performing detoxification treatment, washing with distilled water, and cutting into (1.0-1.4) cm x (1.0-1.4) cm square blocks;
2) culturing, namely transferring the treated baby cabbage tissue into a basic culture medium, spraying infectious bacteria liquid twice at intervals of 24 hours, spraying 0.5-0.8 ml for the first time and 2-3 ml for the second time, culturing in a culture room, culturing for 20-25 days in a dark room at the room temperature of 15-20 ℃, and generating callus;
3) the callus is cut into powder, inoculated on a differentiation culture medium, cultured for 20-25 days under the conditions of illumination for 12h/d, light intensity 2600Lx and room temperature of 20-22 ℃, and induced to differentiate to form adventitious buds;
4) rooting and strengthening seedling culture, namely vertically inoculating adventitious buds to a rooting and strengthening seedling culture medium; culturing for 25-30 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃, and culturing the giant salamander rooting test-tube plantlet with large plants and developed and strong root system;
5) performing proliferation culture, namely cutting off root hairs of the baby cabbage rooted test-tube plantlets, vertically inoculating the root hairs to a proliferation culture medium, and culturing the baby cabbage for 20-24 d under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃; culturing the adventitious buds with required quantity;
6) rooting culture, namely vertically inoculating the adventitious buds obtained in the step 5) on a rooting culture medium, and culturing for 15-20 days under the conditions of illumination for 12h/d, light intensity of 2000-2400 Lx and room temperature of 20-22 ℃ to induce a root system with developed coarse and developed length for preventing pollution;
7) temporary planting: hardening the rooted test-tube plantlets in a greenhouse for 5-7 days, washing the test-tube plantlets with water to remove residual culture medium at the roots, and heeling in on a substrate; after 15-20 days, new roots grow out; obtaining the baby cabbage rooted seedlings.
2. The tissue culture seedling method for baby cabbage with high disease resistance of claim 1, wherein the tissue culture seedling method comprises the following steps: the detoxification treatment in the step 1) comprises washing and soaking, wherein washing and detoxification are carried out by using a washing liquid, and then the washing liquid is taken out and is soaked in a soaking solution at the temperature of 18-20 ℃ for 40-50 min.
3. The tissue culture seedling method for baby cabbage with high disease resistance of claim 2, wherein the tissue culture seedling method comprises the following steps: the flushing fluid comprises sterile water, 0.1% of sodium hypochlorite, 0.1% of mercuric chloride solution, ginkgo extract and gleditsia sinensis extract, and the dosage ratio is 95: 1:1:0.8: 0.6.
4. the tissue culture seedling method for baby cabbage with high disease resistance of claim 2, wherein the tissue culture seedling method comprises the following steps: the soaking solution comprises xanthan gum, potassium citrate, modified polylactic acid, plant growth factors, a coptis chinensis extracting solution, ruyi grass and distilled water, wherein the feed ratio is 15-20: 2-3: 20-30: 1-3: 1-3: 2-4: 40 to 50.
5. The tissue culture seedling method for baby cabbage with high disease resistance of claim 1, wherein the tissue culture seedling method comprises the following steps: and 2) placing the Mesorethrus mollissima leaves just infected with pathogenic bacteria into a container, adding 1 time of glucose solution with the mass fraction of 2% of the mass fraction of the pathogenic leaves, soaking at 30-35 ℃ for 30-45 min, filtering to remove impurities, adding 20 times of water into the filtrate for dilution, adding 1 time of treating agent with the mass fraction of the pathogenic leaves, heating to 48-51 ℃, preserving heat for 20min, and filtering to obtain the bacterial liquid.
6. The tissue culture seedling method for baby cabbage with high disease resistance of claim 5, wherein the tissue culture seedling method comprises the following steps: the infectious germs comprise downy mildew and soft rot.
7. The tissue culture seedling method for baby cabbage with high disease resistance of claim 5, wherein the tissue culture seedling method comprises the following steps: the treating agent comprises folium Artemisiae Argyi extract, flos Lonicerae extract, fructus forsythiae extract, herba Houttuyniae extract and distilled water.
8. The tissue culture seedling method for baby cabbage with high disease resistance of claim 7, wherein the tissue culture seedling method comprises the following steps: the adding amount ratio of the folium artemisiae argyi extract, the honeysuckle extract, the fructus forsythiae extract, the houttuynia cordata extract to the distilled water is 2-3: 2-5: 1-3: 3-7: 85-96.
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| CN114342810A (en) * | 2022-02-23 | 2022-04-15 | 内蒙古科学技术研究院 | Tissue culture method of rhizoma atractylodis with high disease resistance |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107094620A (en) * | 2017-04-10 | 2017-08-29 | 周兴虎 | A kind of method of raising Piao Shu seedling resistances against diseases |
| CN111567402A (en) * | 2020-06-17 | 2020-08-25 | 南通科技职业学院 | Method for cultivating efficient tetraploid pakchoi |
| CN114342810A (en) * | 2022-02-23 | 2022-04-15 | 内蒙古科学技术研究院 | Tissue culture method of rhizoma atractylodis with high disease resistance |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107094620A (en) * | 2017-04-10 | 2017-08-29 | 周兴虎 | A kind of method of raising Piao Shu seedling resistances against diseases |
| CN111567402A (en) * | 2020-06-17 | 2020-08-25 | 南通科技职业学院 | Method for cultivating efficient tetraploid pakchoi |
| CN114342810A (en) * | 2022-02-23 | 2022-04-15 | 内蒙古科学技术研究院 | Tissue culture method of rhizoma atractylodis with high disease resistance |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114342810A (en) * | 2022-02-23 | 2022-04-15 | 内蒙古科学技术研究院 | Tissue culture method of rhizoma atractylodis with high disease resistance |
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