CN105580733B - A kind of morning glory fast propagating culture medium - Google Patents
A kind of morning glory fast propagating culture medium Download PDFInfo
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- CN105580733B CN105580733B CN201510990256.6A CN201510990256A CN105580733B CN 105580733 B CN105580733 B CN 105580733B CN 201510990256 A CN201510990256 A CN 201510990256A CN 105580733 B CN105580733 B CN 105580733B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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Abstract
The present invention relates to a kind of morning glory fast propagating culture medium, including three kinds of callus inducing medium, callus differential medium and root media.The quick breeding that the culture medium of the present invention is specifically adapted to morning glory uses; callus inducing medium can quickly form callus with induced tissue; callus differential medium is used to promote callus to break up; improve growth coefficient; root media can then greatly promote the rooting rate of tissue, and then improve the survival rate of morning glory；Three culture mediums synergy of the present invention, the quick breeding for facilitating morning glory to organize, lays the foundation for the research of morning glory flower color gene engineering.
Description
Technical field
The invention belongs to field of plant tissue culture, is related to a kind of morning glory fast propagating culture medium.
Background technology
Morning glory is the summer and autumn common tendril showy flowers of herbaceous plants, can make to shelter from heat or light before garth and room window, small-sized frame, hedge wall
Beautification, is transplanted in which can also make；Except cultivating in addition to viewing and admiring, seed is conventional Chinese medicine, ugly ox of name (Yunnan), black ugly, semen pharbitidis, two
Ugly (black, the sub- mixing of white race), is used as medicine multi-purpose black ugly, the less use of semen pharbitidis.Morning glory has water pouring diuresis, by phlegm, the effect of desinsection.Flower
Color is one of most important characteristics of plants ' aestheticses, and one of highest priority of flowers improvement.Conventional hybridization, mutation breeding and fixed
There is the drawbacks of very big to selection technique, it is difficult to have breakthrough on new pattern is created to make traditional breeding technology.Genetic engineering it is emerging
Rise and provide brand-new thinking to improve and modifying pattern, other original characters can be retained with directed modification pattern, by leading
Enter foreign gene and change the method for flower color, succeeded in various plants.As flower color gene engineering research
Classical material, the molecular biology research of petunia carry out it is more early, relatively deeply, be the separation of flower color gene, conversion it is main
One of model plant, but morning glory research in this respect or space state, tissue cultures are the bases of transgenosis.
Plant Tissue Breeding (broad sense) is called cultured in vitro, refer to isolated from plant the tissue to suit the requirements, organ or
Cell, protoplast etc., by sterile working, cultivated under manual control condition to obtain the intact plant of regeneration or life
The technology of other products of the production with economic value.Plant Tissue Breeding (narrow sense) refers to plant parts tissue, as forming layer,
Parenchymal tissue, mesophyll tissue, endosperm etc. carry out culture and obtain regeneration plant, also refer to produce from each organ in incubation and are cured
The culture of injured tissue, callus again by being differentiated to form aftergrowth again.
Various culture mediums are used in Plant Tissue Breeding.Culture medium is for microorganism, plant tissue and animal tissue's life
The nutriment of long and maintenance artificial preparation, typically all containing carbohydrate, nitrogen substance, inorganic salts (including trace element)
And vitamin and water etc..Different culture media can add some compounds that itself can not be synthesized, that is, grow according to being actually needed
The factor.
But at present, be not specifically adapted to the culture mediums of morning glory tissue cultures also, had a strong impact on to morning glory this
The research of the classical material of kind.
The content of the invention
The first object of the present invention is to provide a kind of morning glory fast propagating culture medium, is specifically adapted to morning glory tissue training
Support, on the one hand can carry out the quick breeding of rare kind, base is established in the on the other hand research for morning glory flower color gene engineering
Plinth.
The second object of the present invention is to provide the purposes of above-mentioned morning glory fast propagating culture medium.
The present invention is achieved through the following technical solutions：
First, a kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium
With three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, 6-benzyl aminopurine 0.8-1.4 mg/litres, methyl α-naphthyl acetate 1.5-
2.5 mg/litres, 30 g/l of sucrose, 7-8 g/l of plant agar；
Callus differential medium：1 liter of MS culture mediums, 6-benzyl aminopurine 0.5-2.0 mg/litres, methyl α-naphthyl acetate 0.1-
0.5 mg/litre, 30 g/l of sucrose, 7-8 g/l of plant agar；
Root media：500 milliliters of MS culture mediums, heteroauxin 0.5-1.5 mg/litres, 20 g/l of sucrose, plant fine jade
7-8 g/l of fat, 500 milliliters of water.
2nd, according to a kind of above-mentioned morning glory fast propagating culture medium, including callus inducing medium, callus
Three kinds of differential medium and root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, 6-benzyl aminopurine 1.0-1.2 mg/litres, methyl α-naphthyl acetate 1.5-
2.0 mg/litres, 30 g/l of sucrose, 7-8 g/l of plant agar；
Callus differential medium：1 liter of MS culture mediums, 6-benzyl aminopurine 1.0-1.5 mg/litres, methyl α-naphthyl acetate 0.1-
0.3 mg/litre, 30 g/l of sucrose, 7-8 g/l of plant agar；
Root media：500 milliliters of MS culture mediums, heteroauxin 0.75-1.25 mg/litres, 20 g/l of sucrose, plant
7-8 g/l of agar, 500 milliliters of water.
Minimal medium of the MS culture mediums as the technical program, it is the common knowledge of those skilled in the art, it is contained
Component and content such as table 1：
The MS culture mediums of table 1
3rd, application of the above-mentioned culture medium in morning glory quickly breeds culture.
Using the good effect of above-mentioned technical proposal：The culture medium of the present invention is specifically adapted to the quick breeding to morning glory
Use, callus inducing medium can quickly form callus with induced tissue, and callus differential medium is used to promote
Enter callus differentiation, improve growth coefficient, root media can then greatly promote the rooting rate of tissue, and then improve and lead a cow
Colored survival rate；Three culture mediums synergy of the present invention, the quick breeding for facilitating morning glory to organize, is morning glory pattern base
Because the research of engineering lays the foundation.
Embodiment
Technical scheme is described further below by embodiment and test example, but should not be construed as pair
The limitation of the present invention：
Embodiment 1
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.8,1.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.5,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.5,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 2
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.8,1.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.5,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.75,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 3
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.8,2.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.5,0.5 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.0,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 4
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.8,2.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 0.1,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.25,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 5
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,1.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.5,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 6
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,1.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,0.5 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.5,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 7
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,2.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.5,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.75,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 8
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,2.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.5,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.0,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 9
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.2,1.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.5,0.5 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.25,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 10
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.2,1.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 2.0,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.5,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 11
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.2,2.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 2.0,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.5,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 12
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.2,2.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 2.0,0.5 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 0.75,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 13
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.4,1.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.0,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 14
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.4,1.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.0,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.25,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 15
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.4,2.0 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.5,0.1 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 7 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.5,20 g/l of sucrose, plant agar 7
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Embodiment 16
A kind of morning glory fast propagating culture medium, including callus inducing medium, callus differential medium and
Three kinds of root media, wherein,
Callus inducing medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.4,2.5 milligrams of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Callus differential medium：1 liter of MS culture mediums, the mg/litre of 6-benzyl aminopurine 1.5,0.3 milligram of methyl α-naphthyl acetate/
Liter, 30 g/l of sucrose, 8 g/l of plant agar.During preparation, MS culture mediums are first measured, add 6-benzyl aminopurine, methyl α-naphthyl acetate,
Sucrose is added, after sucrose dissolving, adjusts pH to 5.8, adds plant agar, heating melts plant agar, and HTHP goes out
Bacterium (121 DEG C, 25 minutes), it is standby to be cooled to solid.
Root media：500 milliliters of MS culture mediums, the mg/litre of heteroauxin 1.0,20 g/l of sucrose, plant agar 8
G/l, 500 milliliters of water.During preparation, MS culture mediums are first measured, add heteroauxin, sucrose is added, adds water, treat that sucrose dissolves
Afterwards, pH to 5.8 is adjusted, adds plant agar, heating melts plant agar, and autoclave sterilization (121 DEG C, 25 minutes) is cold
But it is standby into solid.
Test example 1
This test example illustrates with optimization of the culture medium to the quick reproductive process and various parameters of morning glory.
1st, seed disinfection
The full flower seed of leading a cow of health is taken, first rinses 10min with running water, then is rinsed 5 times with sterile distilled water, by it
It is put in sterilized triangular flask, first with the Ethanol Treatment 5min that percent by volume is 75%, is then with mass percent
0.1% mercury chloride processing 10min, is finally rinsed 5 times with sterile distilled water, standby.
2nd, the acquisition of morning glory aseptic seedling
Seed after sterilization is inoculated into MS culture mediums, (25 ± 2) DEG C light culture 5 days, then cultivated under light, is cultivated
Obtain within 14 days the high aseptic seedlings of 4-5cm.
3rd, the induction of callus
By the hypocotyl of aseptic seedling be cut into 0.4-0.6cm length segment be put into culture callus inducing medium in train
Support, callus inducing medium refers to added with 0.5-1.5mg/L 6-benzyl aminopurines, the MS of 1.0-2.5mg/L methyl α-naphthyl acetates
Solid medium (embodiment 1-16).When cultivating 5d, hypocotyl starts to expand, and when 15d is arrived in culture, forms diameter 1.0-
1.2cm callus.Experimental result such as table 2：
The callus inducing medium experimental result of table 2
| 6-BenzylAdenine phosphate | NaphthaleneAcetic acid | Callus induction rate | |
| Embodiment 1 | 0.8 | 1.0 | 81% |
| Embodiment 2 | 0.8 | 1.5 | 85% |
| Embodiment 3 | 0.8 | 2.0 | 86% |
| Embodiment 4 | 0.8 | 2.5 | 80% |
| Embodiment 5 | 1.0 | 1.0 | 83% |
| Embodiment 6 | 1.0 | 1.5 | 99% |
| Embodiment 7 | 1.0 | 2.0 | 99% |
| Embodiment 8 | 1.0 | 2.5 | 76% |
| Embodiment 9 | 1.2 | 1.0 | 82% |
| Embodiment 10 | 1.2 | 1.5 | 99% |
| Embodiment 11 | 1.2 | 2.0 | 99% |
| Embodiment 12 | 1.2 | 2.5 | 75% |
| Embodiment 13 | 1.4 | 1.0 | 80% |
| Embodiment 14 | 1.4 | 1.5 | 73% |
| Embodiment 15 | 1.4 | 2.0 | 72% |
| Embodiment 16 | 1.4 | 2.5 | 69% |
It is as shown in the table, and callus induction rate reaches as high as 99%.
4. callus differentiation culture
4-5 rear stable callus of subculture is inoculated into callus differential medium and cultivated, callus point
Change culture medium and refer to that the MS solid mediums added with 0.5-2.0mg/L 6-benzyl aminopurines, 0.1-0.5mg/L methyl α-naphthyl acetates are (real
Apply a 1-12).Experimental result such as table 3：
The callus differential medium experimental result of table 3
| 6-BenzylAdenine phosphate | NaphthaleneAcetic acid | Phenylacetic acid % | Growth coefficient | |
| Embodiment 1 | 0.5 | 0.1 | 33 | 1.2 |
| Embodiment 2 | 0.5 | 0.3 | 41 | 1.5 |
| Embodiment 3 | 0.5 | 0.5 | 40 | 1.6 |
| Embodiment 4 | 1.0 | 0.1 | 88 | 4.0 |
| Embodiment 5 | 1.0 | 0.3 | 89 | 5.8 |
| Embodiment 6 | 1.0 | 0.5 | 68 | 2.4 |
| Embodiment 7 | 1.5 | 0.1 | 89 | 6.0 |
| Embodiment 8 | 1.5 | 0.3 | 88 | 5.5 |
| Embodiment 9 | 1.5 | 0.5 | 70 | 3.0 |
| Embodiment 10 | 2.0 | 0.1 | 61 | 2.4 |
| Embodiment 11 | 2.0 | 0.3 | 65 | 2.5 |
| Embodiment 12 | 2.0 | 0.5 | 52 | 1.8 |
It is as shown in the table, and phenylacetic acid is up to 89%, value-added coefficient 4-6.
5th, culture of rootage
By the tissue-cultured seedling culture that callus differentiates to 4-5cm it is high when, be inoculated in root media and cultivated, it is raw
Root culture medium refers to the 1/2MS solid mediums (embodiment 1-5) of the heteroauxin added with 0.5-1.5mg/L.Experimental result
Such as table 4：
The root media experimental result of table 4
| Heteroauxin | Rooting rate % | Take root and count (bar) | |
| Embodiment 1 | 0.5 | 86 | 5.5 |
| Embodiment 2 | 0.75 | 100 | 8.0 |
| Embodiment 3 | 1.0 | 100 | 9.5 |
| Embodiment 4 | 1.25 | 100 | 10 |
| Embodiment 5 | 1.5 | 91 | 6.0 |
It is as shown in the table, and rooting rate reaches 100%, and number of taking root is 8-10 bars.
6th, tissue culture transplantation of seedlings
When plant root long length is to 1.5-2cm, opens sealed membrane and carry out hardening 2-3 days, be transplanted into sand: the weight ratio of vermiculite
For 1: 1, through excess temperature be 121 DEG C and matrix that pressure is 100kPa sterilizings in, cover within first 5-6 days sealed membrane, keep humidity
For 80%-90%, grown 15-18 days under the conditions of 23-27 DEG C, pour a MS culture medium inorganic salt solution, Zhi Houyi within during which every 3 days
Grown into field soil, substantially increase the survival rate of morning glory.
The culture medium of the present invention is specifically adapted to quick breeding to morning glory and used, and callus inducing medium can be with
Induced tissue quickly forms callus, and callus differential medium is used to promote callus to break up, and improves growth coefficient,
Root media can then greatly promote the rooting rate of tissue, and then improve the survival rate of morning glory；Three cultures of the present invention
Base acts synergistically, and the quick breeding for facilitating morning glory to organize, is laid the foundation for the research of morning glory flower color gene engineering.
Claims (3)
- A kind of 1. morning glory fast propagating culture medium, it is characterised in that：Break up including callus inducing medium, callus Three kinds of culture medium and root media, wherein,Callus inducing medium formula：1 liter of MS culture mediums, 6-benzyl aminopurine 0.8-1.4 mg/litres, methyl α-naphthyl acetate 1.5- 2.5 mg/litres, 30 g/l of sucrose, 7-8 g/l of plant agar；Callus differential medium formula：1 liter of MS culture mediums, 6-benzyl aminopurine 0.5-2.0 mg/litres, methyl α-naphthyl acetate 0.1- 0.5 mg/litre, 30 g/l of sucrose, 7-8 g/l of plant agar；Prescription of rooting medium：500 milliliters of MS culture mediums, heteroauxin 0.5-1.5 mg/litres, 20 g/l of sucrose, plant fine jade 7-8 g/l of fat, 500 milliliters of water.
- A kind of 2. morning glory fast propagating culture medium according to claim 1, it is characterised in that：Including callus induction Three kinds of culture medium, callus differential medium and root media, wherein,Callus inducing medium formula：1 liter of MS culture mediums, 6-benzyl aminopurine 1.0-1.2 mg/litres, methyl α-naphthyl acetate 1.5- 2.0 mg/litres, 30 g/l of sucrose, 7-8 g/l of plant agar；Callus differential medium formula：1 liter of MS culture mediums, 6-benzyl aminopurine 1.0-1.5 mg/litres, methyl α-naphthyl acetate 0.1- 0.3 mg/litre, 30 g/l of sucrose, 7-8 g/l of plant agar；Prescription of rooting medium：500 milliliters of MS culture mediums, heteroauxin 0.75-1.25 mg/litres, 20 g/l of sucrose, plant 7-8 g/l of agar, 500 milliliters of water.
- 3. application of the culture medium in morning glory quickly breeds culture described in claim 1 or 2.
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