CN108450339A - A kind of tissue culture and rapid proliferation method of Hairy Bittercress - Google Patents
A kind of tissue culture and rapid proliferation method of Hairy Bittercress Download PDFInfo
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- CN108450339A CN108450339A CN201810454045.4A CN201810454045A CN108450339A CN 108450339 A CN108450339 A CN 108450339A CN 201810454045 A CN201810454045 A CN 201810454045A CN 108450339 A CN108450339 A CN 108450339A
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- culture
- tissue culture
- hairy bittercress
- agar
- seedling
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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
-
- 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
Abstract
The invention discloses a kind of tissue culture and rapid proliferation method of Hairy Bittercress, method includes the following steps：A, the materials and disinfection of pretreatment and explant；B, aseptic explant is inoculated into inducing culture, obtains callus；C, the induction of Multiple Buds；D, obtained lateral bud of growing thickly is inoculated into strong seedling culture base, to obtain healthy and strong test tube seedling；E, culture of rootage；F, it tames naturally；G, tissue culture transplantation of seedlings.The present invention can greatly reduce the probability of progeny variation in Hairy Bittercress tissue culture procedures by this method; be conducive to keep the excellent shape of parent; the Plant Tissue Breeding of use is convenient for artificial control condition of culture; avoid influence of the environment to culture rooting rate; its production cost is low simultaneously; free from environmental pollution, quantity proliferation is fast, can accomplish scale production.
Description
Technical field
The present invention relates to Hairy Bittercress Cultivating techniques field, specially a kind of tissue culture and rapid proliferation side of Hairy Bittercress
Method.
Background technology
Tall bottle with spout Hairy Bittercress is the distinctive wild vegetable in China, is distributed in one band of Huping mountain nature reserve of Hunan Hubei boundary, is the world
The upper currently the only pure natural Organic Selenium for being dissolvable in water water, can arrange in pairs or groups with any edible raw material, can process selenium-enriched food,
Selenium-rich beverage more can have powerful market prospects and economic value, due to wild as the selenium element additive of health products
Tall bottle with spout Hairy Bittercress low output, cannot be satisfied huge social demand, for this purpose, it is proposed that a kind of tissue cultures of Hairy Bittercress with it is fast
Fast propagation method.
Invention content
The purpose of the present invention is to provide a kind of tissue culture and rapid proliferation methods of Hairy Bittercress, to solve above-mentioned background
The problem of being proposed in technology.
To achieve the above object, the present invention provides the following technical solutions：A kind of tissue culture and rapid proliferation of Hairy Bittercress
Method, method include the following steps：
A, the materials and disinfection of pretreatment and explant；
B, aseptic explant is inoculated into inducing culture, obtains callus；
C, the induction of Multiple Buds；
D, obtained lateral bud of growing thickly is inoculated into strong seedling culture base, to obtain healthy and strong test tube seedling；
E, culture of rootage；
F, it tames naturally；
G, tissue culture transplantation of seedlings.
Preferably, the step A is：Take the tall bottle with spout Hairy Bittercress root that the good paniculate diameter of growth conditions is about 2mm
Shape stem, is placed in clean plate to dry in the air to put to stem apex and stem section and slightly softens, and tall bottle with spout Hairy Bittercress root shape is gently wiped with alcohol swab
The dust on stem surface, then be placed in sterile glass vials, it is sterilized with 75% alcohol and 0.1% mercuric chloride on superclean bench,
And with 75% 20~50s of alcohol disinfecting aseptic water washing is used later 4~5 times with 0.1% mercuric chloride, 20~40min of disinfection,
It is about 1cm spare to be finally cut into length.
Preferably, the inducing culture in the step B is by brown sugar, 6- benzyls aminoadenine, protein hydrolysate, agar, sugarcane
Sugar, distilled water and hormone 2,4-D compositions, ingredient and content are：12~18g/L of brown sugar, 2~6mg/L of 6- benzyl aminoadenines,
3~7mg/L of protein hydrolysate, 6~8g/L of agar, sucrose 20~30 and distilled water 900~950, hormone 2,4-D 2.4~
2.6mg/L。
Preferably, the step C is：Callus is positioned over culturing room, first darkroom turns illumination cultivation after cultivating 7 days,
12 hour/day of photoperiod, 1200~1500lx of illuminance, indoor temperature control in 25 ± 2 DEG C, visible white bud point after 7 days
Turn green, sprout extends after 30 days, cuts sprout after 35 days and is transferred to proliferated culture medium and obtains lateral bud of growing thickly.
Preferably, the proliferated culture medium is by 6-benzyladenine, agar powder, calcium chloride dihydrate, distilled water, white sugar and ZT
Composition, ingredient and content are：0.1~0.5mg/l of 6-benzyladenine, 7~8g/L of agar powder, calcium chloride dihydrate 0.45g/L,
Distilled water 900~950,0.5~1mg/L of white sugar 20g/L, ZT.
Preferably, the strong seedling culture base in the step D by 1/2 improvement MS, CuSO4 powder, indolebutyric acid, activated carbon,
NiSO4 powder, agar and distilled water composition, ingredient and content are：1/2 0.2~0.3mg/L of improvement MS, CuSO4 powder
0.75~0.95mg/L, 1.0~2.0g/L of indolebutyric acid, 0.34~0.57mg/L of activated carbon 100ml/L, NiSO4 powder, agar
20g/L, distilled water 900~950.
Preferably, the step E is：Healthy and strong test tube seedling is cut from base portion, culture to bud in root media is inserted into and gives birth to
Root obtains rooted plantlet.
Preferably, the root media is by MS culture mediums, agar, sucrose, vermiculite power, wheat germ powder and distilled water group
At ingredient and content are：0.5~1mg/L of MS culture mediums, agar 5g/L, 30~40g/L of sucrose, 0.6~0.8g/ of vermiculite power
L, 1~2g/L of wheat germ powder, distilled water 700~850.
Preferably, the step F is：After culture of rootage 25~30 days, sees that adventitious root is grown and sprouted with subsequent root restriction
Hair, bottle seedling go in chamber facility the domestication for receiving natural light and temperature, and temperature is within the scope of 15~28 DEG C during domestication, illumination
Within the scope of 3000~5000lx, bottle outlet is transplanted after domestication 20~25 days.
Preferably, the step G is：Wait for that small seedling leaf is unfolded, leaf color is dark green, when 3~5cm of height of seedling, opens bottle cap hardening 2
After~4 days, by seedling plant agar and sundries clean, transplanting to sterilize after vermiculite, peat soil and perlite mixing
In matrix, sprinkle profoundly water, cover film, keep 70% or more humidity, set ventilating and cooling place, 24~26 DEG C, illumination 1500~
It moves under natural conditions and grows after being cultivated 8~12 days under the conditions of 2500lx.
Compared with prior art, beneficial effects of the present invention are as follows：
The present invention can greatly reduce the probability of progeny variation in Hairy Bittercress tissue culture procedures by this method, be conducive to protect
The excellent shape of parent is held, the Plant Tissue Breeding of use avoids environment and take root to culture convenient for artificial control condition of culture
The influence of rate, while its production cost is low, free from environmental pollution, quantity proliferation is fast, can accomplish scale production.
Description of the drawings
Fig. 1 is the method for the present invention flow diagram.
Specific implementation mode
Following will be combined with the drawings in the embodiments of the present invention, and technical solution in the embodiment of the present invention carries out clear, complete
Site preparation describes, it is clear that described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.It is based on
Embodiment in the present invention, it is obtained by those of ordinary skill in the art without making creative efforts every other
Embodiment shall fall within the protection scope of the present invention.
Referring to Fig. 1, a kind of tissue culture and rapid proliferation method of Hairy Bittercress, method include the following steps：
A, the materials and disinfection of pretreatment and explant；
B, aseptic explant is inoculated into inducing culture, obtains callus；
C, the induction of Multiple Buds；
D, obtained lateral bud of growing thickly is inoculated into strong seedling culture base, to obtain healthy and strong test tube seedling；
E, culture of rootage；
F, it tames naturally；
G, tissue culture transplantation of seedlings.
Step A is：The tall bottle with spout Hairy Bittercress root-like stock that the good paniculate diameter of growth conditions is about 2mm is taken, is placed in dry
It dries in the air to put to stem apex and stem section in net plate and slightly soften, the ash on tall bottle with spout Hairy Bittercress root-like stock surface is gently wiped with alcohol swab
Dirt, then be placed in sterile glass vials is sterilized on superclean bench with 75% alcohol and 0.1% mercuric chloride, and with 75% wine
Essence 20~50s of disinfection, 20~40min is sterilized with 0.1% mercuric chloride, uses aseptic water washing later 4~5 times, is finally cut into length
Degree is about 1cm spare.
Inducing culture in step B by brown sugar, 6- benzyls aminoadenine, protein hydrolysate, agar, sucrose, distilled water and
Hormone 2,4-D compositions, ingredient and content are：12~18g/L of brown sugar, 2~6mg/L of 6- benzyl aminoadenines, protein hydrolysate 3~
7mg/L, 6~8g/L of agar, sucrose 20~30 and distilled water 900~950,2.4~2.6mg/L of hormone 2,4-D.
Step C is：Callus is positioned over culturing room, first darkroom turns illumination cultivation after cultivating 7 days, and the photoperiod 12 is small
When/day, 1200~1500lx of illuminance, indoor temperature control is in 25 ± 2 DEG C, and visible white bud point turns green after 7 days, after 30 days
Sprout extends, and cuts sprout after 35 days and is transferred to proliferated culture medium and obtains lateral bud of growing thickly.
Proliferated culture medium is made of 6-benzyladenine, agar powder, calcium chloride dihydrate, distilled water, white sugar and ZT, ingredient
And content is：0.1~0.5mg/l of 6-benzyladenine, 7~8g/L of agar powder, calcium chloride dihydrate 0.45g/L, distilled water 900~
950,0.5~1mg/L of white sugar 20g/L, ZT.
Strong seedling culture base in step D is by 1/2 improvement MS, CuSO4 powder, indolebutyric acid, activated carbon, NiSO4 powder, fine jade
Fat and distilled water composition, ingredient and content are：1/2 improvement MS 0.2~0.3mg/L, 0.75~0.95mg/L of CuSO4 powder,
1.0~2.0g/L of indolebutyric acid, 0.34~0.57mg/L of activated carbon 100ml/L, NiSO4 powder, agar 20g/L, distilled water 900
~950.
Step E is：Healthy and strong test tube seedling is cut from base portion, is inserted into root media to cultivate and take root to bud, taken root
Seedling.
Root media is made of MS culture mediums, agar, sucrose, vermiculite power, wheat germ powder and distilled water, ingredient and
Content is：0.5~1mg/L of MS culture mediums, agar 5g/L, 30~40g/L of sucrose, 0.6~0.8g/L of vermiculite power, wheat germ powder
1~2g/L, distilled water 700~850.
Step F is：After culture of rootage 25~30 days, sees that adventitious root is grown and sprouted with subsequent root restriction, bottle seedling is gone to
Receive the domestication of natural light and temperature in chamber facility, during domestication temperature within the scope of 15~28 DEG C, illumination 3000~
Within the scope of 5000lx, bottle outlet is transplanted after domestication 20~25 days.
Step G is：Wait for that small seedling leaf is unfolded, leaf color is dark green, when 3~5cm of height of seedling, opens bottle cap hardening after 2~4 days, will
Agar and sundries on seedling plant are cleaned, and in the mixed-matrix of vermiculite, peat soil and perlite after transplanting to disinfection, are irrigated
Water covers film, keeps 70% or more humidity, sets ventilating and cooling place, trained under the conditions of 24~26 DEG C, 1500~2500lx of illumination
It moves under natural conditions and grows after supporting 8~12 days.
In use, this method can greatly reduce the probability of progeny variation in Hairy Bittercress tissue culture procedures, be conducive to keep
The Plant Tissue Breeding of the excellent shape of parent, use avoids environment to cultivating rooting rate convenient for artificial control condition of culture
Influence, while its production cost is low, free from environmental pollution, and quantity proliferation is fast, can accomplish scale production.
It although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
Understanding without departing from the principles and spirit of the present invention can carry out these embodiments a variety of variations, modification, replace
And modification, the scope of the present invention is defined by the appended.
Claims (10)
1. a kind of tissue culture and rapid proliferation method of Hairy Bittercress, it is characterised in that：Its method includes the following steps：
A, the materials and disinfection of pretreatment and explant；
B, aseptic explant is inoculated into inducing culture, obtains callus；
C, the induction of Multiple Buds；
D, obtained lateral bud of growing thickly is inoculated into strong seedling culture base, to obtain healthy and strong test tube seedling；
E, culture of rootage；
F, it tames naturally；
G, tissue culture transplantation of seedlings.
2. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Suddenly A is：The tall bottle with spout Hairy Bittercress root-like stock that the good paniculate diameter of growth conditions is about 2mm is taken, is placed in clean plate
It dries in the air to put to stem apex and stem section and slightly soften, the dust on tall bottle with spout Hairy Bittercress root-like stock surface is gently wiped with alcohol swab, then be placed in nothing
In bacterium vial, sterilized with 75% alcohol and 0.1% mercuric chloride on superclean bench, and with 75% alcohol disinfecting 20~
50s, 20~40min is sterilized with 0.1% mercuric chloride, uses aseptic water washing later 4~5 times, and it is about 1cm standby to be finally cut into length
With.
3. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Inducing culture in rapid B is by brown sugar, 6- benzyls aminoadenine, protein hydrolysate, agar, sucrose, distilled water and hormone 2,4-D groups
At ingredient and content are：12~18g/L of brown sugar, 2~6mg/L of 6- benzyl aminoadenines, 3~7mg/L of protein hydrolysate, agar 6
~8g/L, sucrose 20~30 and distilled water 900~950,2.4~2.6mg/L of hormone 2,4-D.
4. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Suddenly C is：Callus is positioned over culturing room, first darkroom turns illumination cultivation, 12 hour/day of photoperiod, illuminance after cultivating 7 days
1200~1500lx, indoor temperature control in 25 ± 2 DEG C, and visible white bud point turns green after 7 days, and sprout extends after 30 days, and 35
Sprout is cut after it be transferred to proliferated culture medium obtain lateral bud of growing thickly.
5. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 4, it is characterised in that：The increasing
It grows culture medium to be made of 6-benzyladenine, agar powder, calcium chloride dihydrate, distilled water, white sugar and ZT, ingredient and content are：
0.1~0.5mg/l of 6-benzyladenine, 7~8g/L of agar powder, calcium chloride dihydrate 0.45g/L, distilled water 900~950, white sugar
0.5~1mg/L of 20g/L, ZT.
6. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Strong seedling culture base in rapid D is by 1/2 improvement MS, CuSO4 powder, indolebutyric acid, activated carbon, NiSO4 powder, agar and distilled water
Composition, ingredient and content are：1/2 0.2~0.3mg/L of improvement MS, 0.75~0.95mg/L of CuSO4 powder, indolebutyric acid
1.0~2.0g/L, 0.34~0.57mg/L of activated carbon 100ml/L, NiSO4 powder, agar 20g/L, distilled water 900~950.
7. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Suddenly E is：Healthy and strong test tube seedling is cut from base portion, is inserted into root media to cultivate and take root to bud, obtain rooted plantlet.
8. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 7, it is characterised in that：The life
Root culture medium is made of MS culture mediums, agar, sucrose, vermiculite power, wheat germ powder and distilled water, and ingredient and content are：MS
0.5~1mg/L of culture medium, agar 5g/L, 30~40g/L of sucrose, 0.6~0.8g/L of vermiculite power, 1~2g/L of wheat germ powder,
Distilled water 700~850.
9. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：The step
Suddenly F is：After culture of rootage 25~30 days, sees that adventitious root is grown and sprouted with subsequent root restriction, bottle seedling is gone in chamber facility
Receive the domestication of natural light and temperature, during domestication temperature within the scope of 15~28 DEG C, illumination within the scope of 3000~5000lx,
Bottle outlet is transplanted after domestication 20~25 days.
10. a kind of tissue culture and rapid proliferation method of Hairy Bittercress according to claim 1, it is characterised in that：It is described
Step G is：Wait for that small seedling leaf is unfolded, leaf color is dark green, when 3~5cm of height of seedling, bottle cap hardening is opened after 2~4 days, by seedling plant
On agar and sundries clean, vermiculite of the transplanting to after sterilizing, peat soil and perlite mixed-matrix in, sprinkle profoundly water, cover
Film keeps 70% or more humidity, sets ventilating and cooling place, and 8~12 are cultivated under the conditions of 24~26 DEG C, 1500~2500lx of illumination
It moves under natural conditions and grows after it.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810454045.4A CN108450339A (en) | 2018-05-14 | 2018-05-14 | A kind of tissue culture and rapid proliferation method of Hairy Bittercress |
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|---|---|---|---|
| CN201810454045.4A CN108450339A (en) | 2018-05-14 | 2018-05-14 | A kind of tissue culture and rapid proliferation method of Hairy Bittercress |
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| CN108450339A true CN108450339A (en) | 2018-08-28 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN109652360A (en) * | 2019-01-11 | 2019-04-19 | 成都大学 | A kind of Rhizoma Et Radix Notopterygii cell culture processes obtaining high biological yield |
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| KR101131865B1 (en) * | 2008-05-13 | 2012-04-03 | 한국과학기술원 | Method for modifying the germination of plant seed by controlling the level of SOMNUS or functional equivalents thereof in cell |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109652360A (en) * | 2019-01-11 | 2019-04-19 | 成都大学 | A kind of Rhizoma Et Radix Notopterygii cell culture processes obtaining high biological yield |
| CN109652360B (en) * | 2019-01-11 | 2022-04-15 | 成都大学 | Notopterygium incisum cell culture method for obtaining high biological yield |
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Application publication date: 20180828 |