CN102726300A - Anther culture method of cauliflower - Google Patents

Anther culture method of cauliflower Download PDF

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Publication number
CN102726300A
CN102726300A CN2012102318490A CN201210231849A CN102726300A CN 102726300 A CN102726300 A CN 102726300A CN 2012102318490 A CN2012102318490 A CN 2012102318490A CN 201210231849 A CN201210231849 A CN 201210231849A CN 102726300 A CN102726300 A CN 102726300A
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CN
China
Prior art keywords
culture
callus
flower pesticide
cauliflower
anther
Prior art date
Application number
CN2012102318490A
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Chinese (zh)
Inventor
张绪璋
汤绍康
张丽
吴枝泉
吴雨英
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福建农林大学
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Application filed by 福建农林大学 filed Critical 福建农林大学
Priority to CN2012102318490A priority Critical patent/CN102726300A/en
Publication of CN102726300A publication Critical patent/CN102726300A/en

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Abstract

The invention relates to an anther culture method of cauliflower, belonging to biology technical field. The method includes taking anther of cauliflower as material, inoculating under sterile condition, performing anther induced callus culture, performing callus differentiation culture, and performing differentiated seedling rooting culture, to realize culture of new cauliflower seedlings. Anther culture can keep stable parent purity, and accelerate selection effect of hybrid progeny. The method has good application value for accelerating culture of excellent cauliflower variety.

Description

A kind of anther culture method of cauliflower

Technical field

The invention belongs to biological technical field, be specifically related to a kind of anther culture method of cauliflower.

Background technology

Cauliflower ( Brassica oleracea Var. botrytis) have another name called cauliflower, be one of China's autumn and winter main breed.Because of its edible part raw fiber is few, be of high nutritive value, be well received by consumers.Utilize different ecological type kind to hybridize, can cultivate the cauliflower variety that makes new advances, have very strong hybrid vigour.But cauliflower belongs to the Cruciferae cross pollinated plant, and inbreeding depression is serious, and the self incompatible line seed selection is difficulty, and kind relies on colony's heterozygosity to keep stable.And cultivation Hybrid difficulty is big.The purity that can protect special parent through anther culture is stable, accelerates hybrid generation's selection effect.Has good using value for accelerating to cultivate good cabbage kind.

Summary of the invention

The object of the present invention is to provide a kind of anther culture method of cauliflower.

The anther culture method of a kind of cauliflower provided by the invention, described cultural method may further comprise the steps: (1) flower pesticide collection and disinfecting; (2) anther callus is cultivated; (3) callus differentiation culture; (4) the differentiation seedling rooting is cultivated; (5) refining seedling, heel in, transplant.

Described cultural method specifically may further comprise the steps:

(1) flower pesticide collection and disinfecting

1. flower pesticide collection: getting length in the fine morning is the full bud of 3mm flower pesticide, confirms that through microscopy the pollen development of this moment is in monokaryotic stage or monokaryon later stage;

2. flower pesticide is disinfected: bud with 0.1% mercuric chloride solution sterilization 300 minutes, is used aseptic water washing 5 times through 70% after alcohol-pickled 30 seconds then;

(2) anther callus is cultivated

1. evoked callus medium: basal medium E3+BA0.5mg/L+ NAA0.1 mg/L,

The medium compound method is pressed routine operation.

2. inoculation: the bud that the cancellation poison is crossed, under aseptic condition, remove petal with dissecting needle or tweezers, clamp filigree with tweezers, take out flower pesticide, flower pesticide directly is inoculated on the evoked callus medium equably, connects 3 flower pesticide in every 200ml tissue culture bottle;

3. cultivate: rearmounted 25 ± 1 ℃ of dark the cultivations 7 days of inoculation, then every day illumination cultivation 12 hours, intensity of illumination 800 luxs, relative moisture is 65%;

(3) callus differentiation culture

1. differential medium: MS+0.5-1mg/L 6-BA+0.5 mg/L NAA;

2. the callus lines growth increases, and transfers green gradually to, changes differential medium then over to and cultivates;

3. condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1000~1500LX;

(4) the differentiation seedling rooting is cultivated

1. root media: 1/2MS+0.1 mg/L NAA+ activated carbon 0.2%;

2. work as seedling and grow tall 3~4 centimetres, when growing 3 leaves, insert root media and cultivate;

3. condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1500~2000LX;

(5) heel in and grow seedlings

1. refine seedling: when the seedling base portion sends out roots, when reaching 2cm, length opens bottle cap, 25 ℃ of environmental temperatures, humidity is controlled at 85%, intensity of illumination 2000LX, refining seedling 3~5 days;

2. heel in: behind the refining seedling, move on in the plastics shallow bid and grow seedlings, heel in 25 ℃ of back environmental temperatures, humidity is controlled at 90%, and intensity of illumination 2000LX moves field production after becoming to live.

Cultivate through 3~5 seville orange flower medicines, the cauliflower offspring is tended towards stability, can be used as cauliflower breeding cross parent's material.

Advantage of the present invention: the purity that can protect special parent through anther culture is stable, accelerates hybrid generation's selection effect.Has good using value for accelerating to cultivate good cabbage kind.

Description of drawings

Fig. 1 is flower pesticide callus figure.

Fig. 2 is divided into seedling for callus.

Fig. 3 is a culture of rootage.

Fig. 4 is for heeling in seedling.

Embodiment

Embodiment 1

1, flower pesticide collection and disinfecting

(1) flower pesticide collection: the fine morning get length be the full bud of 3mm left and right sides flower pesticide as experiment material, confirm that through microscopy the pollen development of this moment is in monokaryotic stage or monokaryon later stage.

(2) flower pesticide is disinfected: bud through 70% spill smart soak 30 seconds after, sterilized 300 minutes with 0.1% mercuric chloride solution, use aseptic water washing then 5 times.

2, inoculated and cultured anther callus

(1) evoked callus medium: E3*+BA0.5mg/L+ NAA0.1 mg/L.

The medium compound method is pressed routine operation.

(2) inoculation: the bud that the cancellation poison is crossed, under aseptic condition, remove petal with dissecting needle or tweezers, clamp filigree with tweezers, take out flower pesticide.Flower pesticide directly is inoculated in equably on the evoked callus medium and connects 3 flower pesticide in (rejecting filigree, flat grain flower pesticide) every bottle (200 tissue culture bottle).

(3) cultivate: the dark cultivation 7 days of rearmounted 25 ± 1 ℃ of degree of inoculation, then every day illumination cultivation 12 hours, intensity of illumination 800 luxs, relative moisture is about 65%.Cultivate 30~40 days visible anther callus, as shown in Figure 1.

3, callus differentiation culture

(1) differential medium: MS+0.5-1mg/L 6-BA+0.5 mg/L NAA.

(2) the callus lines growth increases, and transfers green gradually to, changes differential medium then over to, 2 callus lines of every bottle graft.

(3) condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1000~1500LX, to cultivate 15-20 days, callus is divided into seedling, and is as shown in Figure 2.

4, culture of rootage

(1) root media: 1/2MS+0.1 mg/L NAA+ activated carbon 0.2%.

(2) grow tall 3~4 centimetres when seedling, when growing 3 leaves, change root media over to and cultivate.

(3) condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1500~2000LX, cultivated 10-15 days, well developed root system, as shown in Figure 3.

5, heel in and grow seedlings

(1) refining seedling: when the seedling base portion sends out roots, when reaching 2cm, length opens bottle cap, 25 ℃ of environmental temperatures, humidity is controlled at about 85%, the about 2000LX of intensity of illumination.Refining seedling 3~5 days.

(2) heel in: behind the refining seedling, move on in the plastics shallow bid and grow seedlings, the soil of growing seedlings will pass through and disinfect, and heels in 25 ℃ of back environmental temperatures, and humidity is controlled at 90%, and intensity of illumination 2000LX obtains heeling in seedling after one week, as shown in Figure 4.Move field production after becoming to live.

6, cultivate the cauliflower hybrid strain: cultivate through 3~5 seville orange flower medicines, the cauliflower offspring is tended towards stability, can be used as cauliflower breeding cross parent's material.

Attach the E3 medium:

Claims (2)

1. the anther culture method of a cauliflower, it is characterized in that: described cultural method may further comprise the steps: (1) flower pesticide collection and disinfecting; (2) anther callus is cultivated; (3) callus differentiation culture; (4) the differentiation seedling rooting is cultivated; (5) refining seedling, heel in, transplant.
2. the anther culture method of cauliflower according to claim 1 is characterized in that, described cultural method specifically may further comprise the steps:
(1) flower pesticide collection and disinfecting
1. flower pesticide collection: getting length in the fine morning is the full bud of 3mm flower pesticide, confirms that through microscopy the pollen development of this moment is in monokaryotic stage or monokaryon later stage;
2. flower pesticide is disinfected: bud with 0.1% mercuric chloride solution sterilization 300 minutes, is used aseptic water washing 5 times through 70% after alcohol-pickled 30 seconds then;
(2) anther callus is cultivated
1. evoked callus medium: basal medium E3+BA0.5mg/L+ NAA0.1 mg/L,
The medium compound method is pressed routine operation;
2. inoculation: the bud that the cancellation poison is crossed, under aseptic condition, remove petal with dissecting needle or tweezers, clamp filigree with tweezers, take out flower pesticide, flower pesticide directly is inoculated on the evoked callus medium equably, connects 3 flower pesticide in every 200ml tissue culture bottle;
3. cultivate: rearmounted 25 ± 1 ℃ of dark the cultivations 7 days of inoculation, then every day illumination cultivation 12 hours, intensity of illumination 800 luxs, relative moisture is 65%;
(3) callus differentiation culture
1. differential medium: MS+0.5-1mg/L 6-BA+0.5 mg/L NAA;
2. the callus lines growth increases, and transfers green gradually to, changes differential medium then over to and cultivates;
3. condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1000~1500LX;
(4) the differentiation seedling rooting is cultivated
1. root media: 1/2MS+0.1 mg/L NAA+ activated carbon 0.2%;
2. work as seedling and grow tall 3~4 centimetres, when growing 3 leaves, insert root media and cultivate;
3. condition of culture: 25 ℃ of temperature, humidity 70%, intensity of illumination 1500~2000LX;
(5) heel in and grow seedlings
1. refine seedling: when the seedling base portion sends out roots, when reaching 2cm, length opens bottle cap, 25 ℃ of environmental temperatures, humidity is controlled at 85%, intensity of illumination 2000LX, refining seedling 3~5 days;
2. heel in: behind the refining seedling, move on in the plastics shallow bid and grow seedlings, heel in 25 ℃ of back environmental temperatures, humidity is controlled at 90%, and intensity of illumination 2000LX moves field production after becoming to live.
CN2012102318490A 2012-07-06 2012-07-06 Anther culture method of cauliflower CN102726300A (en)

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CN2012102318490A CN102726300A (en) 2012-07-06 2012-07-06 Anther culture method of cauliflower

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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH09206069A (en) * 1996-02-02 1997-08-12 Suzuki Motor Corp Cell culture using bag of semipermeable membrane
CN1180744A (en) * 1996-10-25 1998-05-06 辽宁师范大学生物工程研究所 Method of tissue culture quick breeding for white cauliflower and green cauliflower cross breed new strain test-tube seeding
CN1552198A (en) * 2003-06-05 2004-12-08 天津科润农业科技股份有限公司 Method for culturing cauliflower regenerative tree by free microspore culture technology
US20050183150A1 (en) * 2004-02-13 2005-08-18 Rebecca Torisky Method for regenerating and transforming St. Augustinegrass from embryogenic callus
WO2008042392A2 (en) * 2006-10-03 2008-04-10 Seminis Vegetable Seeds, Inc. Brilliant white cauliflower
CN101810136A (en) * 2010-05-07 2010-08-25 福州市蔬菜科学研究所 Cauliflower cytoplasmic male sterile line breeding method and application of male sterile line

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH09206069A (en) * 1996-02-02 1997-08-12 Suzuki Motor Corp Cell culture using bag of semipermeable membrane
CN1180744A (en) * 1996-10-25 1998-05-06 辽宁师范大学生物工程研究所 Method of tissue culture quick breeding for white cauliflower and green cauliflower cross breed new strain test-tube seeding
CN1552198A (en) * 2003-06-05 2004-12-08 天津科润农业科技股份有限公司 Method for culturing cauliflower regenerative tree by free microspore culture technology
US20050183150A1 (en) * 2004-02-13 2005-08-18 Rebecca Torisky Method for regenerating and transforming St. Augustinegrass from embryogenic callus
WO2008042392A2 (en) * 2006-10-03 2008-04-10 Seminis Vegetable Seeds, Inc. Brilliant white cauliflower
CN101810136A (en) * 2010-05-07 2010-08-25 福州市蔬菜科学研究所 Cauliflower cytoplasmic male sterile line breeding method and application of male sterile line

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
《河南科技大学学报:自然科学版》 20060831 张绪璋等 花椰菜花药培养试验研究 第69页1材料与方法、第71页2.4 2 第27卷, 第4期 *
张绪璋等: "花椰菜花药培养试验研究", 《河南科技大学学报:自然科学版》 *

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Application publication date: 20121017