CN108848994A - A kind of method of suitable grape micrografting seedling Liquid Culture - Google Patents
A kind of method of suitable grape micrografting seedling Liquid Culture Download PDFInfo
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- CN108848994A CN108848994A CN201810853915.5A CN201810853915A CN108848994A CN 108848994 A CN108848994 A CN 108848994A CN 201810853915 A CN201810853915 A CN 201810853915A CN 108848994 A CN108848994 A CN 108848994A
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- grafting
- culture
- scion
- seedling
- grape
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G2/00—Vegetative propagation
- A01G2/30—Grafting
- A01G2/35—Cutting; Inserting
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G31/00—Soilless cultivation, e.g. hydroponics
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
Abstract
The present invention relates to a kind of methods of suitable grape micrografting seedling Liquid Culture, its main feature is that, include the following steps：(1) to cultivate 30-40 days stock variety tissue-cultured seedling as material；(2) to cultivate 30-40 days scion variety tissue-cultured seedling as material；(3) aseptically, by the notch of scion beveled end insertion stock, grafting is formed；(4) U-shaped card is made to be fixedly clamped the graft union of grafting with tweezers compressing U-shaped card both ends；(5) the miniature grafting grafted is inserted on ready chalina, then the chalina for being inserted with miniature grafting is put into the culture bottle for filling 50ml fluid nutrient medium and is cultivated.The method of the present invention has the beneficial effect that：1. filtering out a kind of Liquid Culture based formulas for being suitble to multiple grape Stockscion combination cultures, using the grape grafting rooting rate of the culture medium culture up to 100%, and the root system turned out is more flourishing compared with solid culture.
Description
Technical field
The present invention relates to a kind of methods of suitable grape micrografting seedling Liquid Culture.
Background technique
Micrografting technology be a kind of tissue cultures combined with grafting, by the stock of different cultivars and scion tissue-cultured seedling in group
Knit the technology grafted under condition of culture.Shoot-tip Grafting skill is applied at first from Du Linbosi the 1950s using ivy
Since art, miniature graft technology obtains development at full speed.The U.S., France, Italy, Spain, South Africa, Israel, Japan and
It is Chinese successive in succession in fruits such as citrus, apple, peach, cherry, apricot, Lee, strawberry, grape, avocado, almond, jujube, Kuerle delicious pears
It is applied when setting micrografting.This technology is now mainly used for fruit-tree grafting seedling fast breeding, detoxic seedling is cultivated, graft compatibility mirror
Fixed, viral diagnosis and fruit-tree grafting seedling resistance mechanism study etc..
Currently, the mode that fruit tree micrografting seedling is cultivated is mainly based on solid culture, and research is concentrated mainly on culture medium
Formula, grafting method, anvil fringe compatibility etc., it is less to the specific method research of micrografting, and in specific operation process,
Since anvil fringe combines, insecure or stock, scion is dealt with improperly leads to graft easy failure.For solid culture, liquid
Culture is more conducive to the growth of micrografting seedling root system, and is more suitable for (adding PEG, ABA in the medium by short time environment stress
Deng) carry out the degeneration-resistant mechanism study of grafting.Although there is only a few research to carry out the Liquid Culture of fruit tree micrografting seedling,
The supporter of its Liquid Culture is mainly paper bridge, is easy to cause grafting to cultivate failure because of paper bridge damage during the cultivation process.
At home, the cultivation of grape micrografting seedling has relevant research report, but graft survival rate is relatively low (being lower than 70%), and has no
There is the relevant report of grape micrografting Liquid Culture.In view of the above problems, the present invention mainly takes root to grape grafting cultivation
Culture medium, stock, scion processing mode, grafting fixed form and Liquid Culture supporter etc. are innovated, and establish one
The method of the perfect suitable grape micrografting seedling Liquid Culture of kind, this method can be applied to multiple and different grape Stockscion combinations
Grafting is cultivated, and the grape seedlings grafted by this method, well developed root system, graft survival rate is more than 85%, is far longer than existing
The survival rate that grape micrografting seedling is cultivated.
Summary of the invention
The object of the present invention is to provide a kind of methods of suitable grape micrografting seedling Liquid Culture, and this method is easy to operate,
Grape micrografting shoot survival percent height, well developed root system, can be applied not only to the factorial production of grape grafting, are also convenient for grape
The degeneration-resistant Journal of Sex Research of grafting.
A kind of method of suitable grape micrografting seedling Liquid Culture, its special feature is that, include the following steps：
(1) using the stock variety tissue-cultured seedling for cultivating 30-40 days as material, remove the tiny part in upper end and lower end lignifying
Part, leaving and taking length is the middle stem section of 2~3cm and rugosity greater than 1mm as stock, weeds out institute in stem section with scalpel
Some blades and axillary bud；
(2) using the scion variety tissue-cultured seedling for cultivating 30-40 days as material, take the stem section with full axillary bud as scion,
Remove blade, an axillary bud is only stayed on each stem section top, it is desirable that scion length is greater than 1cm, and rugosity is greater than 1mm；
(3) aseptically, with scalpel, a 0.4-0.6cm is cut in ready stock upper end in step (1)
Notch, by scion lower end ready in step (2) chamfer an inclined-plane, bevel angle≤30 degree, then by scion inclined-plane
The notch of end insertion stock, forms grafting；
(4) aluminium foil with a thickness of 1-1.2mm is selected, aluminium foil is cut into the small item of rectangle of long 2cm wide 0.5cm, then
It is folded in half into U-shaped card, U-shaped card will be prepared is placed in closed container resistant to high temperature and carry out high pressure sterilization, with U-shaped after sterilizing
Clamp folder in the graft union of middle grafting, makes U-shaped card be fixedly clamped and transfers with tweezers compressing U-shaped card both ends in step (3)
The graft union of seedlings picking；
(5) mature chalina is chosen, rear deseeding is dried, the square tiles that side length is 4-5cm is then cut into, put
Enter in closed container resistant to high temperature, add a small amount of distilled water, high pressure sterilization is spare after sealing；
(6) on ready chalina, miniature transfer will then will be inserted in the miniature grafting inserting step (4) grafted
The chalina of seedlings picking is put into the culture bottle for filling 50ml fluid nutrient medium and cultivates.
Culture bottle diameter 6cm, height 12cm, fluid nutrient medium group become in step (6)：1/2B5A great number of elements+B5Molysite+
B5Microelement+B5Organic+0.2~0.4mgL of indolebutyric acid-1+ sucrose 30gL-1, pH6.0.
Condition of culture is to cultivate preceding 5 days low light cultures in step (6), it is desirable that intensity of illumination 1000Lx, strong optical culture later,
It is required that intensity of illumination is greater than 2500Lx, cultivation temperature is 25 ± 2 DEG C, can seedling after culture 40d.
The method of the present invention is a kind of method of suitable grape micrografting seedling Liquid Culture, and its advantages are：1. filtering out
A kind of Liquid Culture based formulas being suitble to multiple grape Stockscion combination cultures grafts seedling rooting using the grape of the culture medium culture
Rate is up to 100%, and the root system turned out is more flourishing compared with solid culture；It is made into 2. selecting with a thickness of the aluminium foil of 1-1.2mm for material
" U " clamp, it is more convenient compared with other materials in fixing grafting mouth, it is easy to operate, be to select aluminium foil to do material to use height first
Pressure sterilizing, sterilizes more thorough, and the aluminium foil of followed by selection 1-1.2mm thickness can both guarantee to have in fixing grafting mouth enough
Dynamics guarantees that the material of grafting is not easily to fall off, and has good plasticity, can arbitrarily change with the rugosity of grafting twig
Shape；3. chalina is selected to do the supporter of grafting Liquid Culture, high pressure sterilization can be not only used, is sterilized more thorough, and
It is porous, it is flexible, convenient for the insertion of grafting, and it is unlikely to deform damage, operated more simple and efficient.
Specific embodiment
The method of the present invention includes following steps：
(1) preparation and packing of culture medium：
According to culture medium prescription：1/2B5(a large amount of)+B5(molysite)+B5(micro)+B5(organic)+indolebutyric acid 0.2~
0.4mg·L-1+ sucrose 30gL-1PH6.0 prepares culture medium, and is sub-packed in the glass culture of diameter 6-8cm, height 12-15cm
In bottle, the culture medium of every bottle of packing 40-50mL, cooling is spare after high pressure sterilization.
(2) preparation of aluminium foil clamp and chalina supporter
Selecting with a thickness of the aluminium foil of 1-1.2mm is material, is cut into long 2cm, the small item of the rectangle of wide 0.5cm, doubling
At U-shaped card, U-shaped card is placed in closed container resistant to high temperature, it is then spare after high pressure sterilization；Choose mature sponge gourd
Flesh dries rear deseeding, is cut into the square tiles (depending on the diameter of culture bottle) that side length is 4-5cm, is set
In closed container resistant to high temperature, appropriate distilled water is added, high pressure sterilization is spare after sealing.
(3) grape rootstock, scion preparation and grafting
1. the preparation of stock：Using the stock variety tissue-cultured seedling for cultivating 30-40d as material, remove the tiny part in upper end and
It leaves and takes the middle stem section (rugosity is greater than 1mm) that length is 2~3cm and is used as stock, and rejected with scalpel in lower end lignifying part
Fall blade and axillary bud all in stem section；
2. the preparation of scion：Using the scion variety tissue-cultured seedling for cultivating 30-40d as material, the stem section with full axillary bud is taken,
Remove blade, an axillary bud, stem section length 1-2cm are only stayed in each stem section top, and rugosity is greater than 1mm；
3. grafting：Aseptically, cutting for a 0.4-0.6cm is cut in ready stock upper end with scalpel
Mouthful, an inclined-plane is chamfer in ready scion lower end, then scion beveled end is inserted into cutting for stock by bevel angle≤30 degree
Mouthful, form grafting.
(4) grafting is fixed：
It is clipped in the graft union of grafting with the U-typed aluminium foil clamp after sterilizing, U-shaped card both ends are gently oppressed with tweezers,
U-shaped card is set to be fixedly secured graft union.
(5) Liquid Culture：
The miniature grafting grafted is inserted on spare chalina, then puts the chalina for being inserted with miniature grafting
Enter to be ready to fill and normally be cultivated in the culture bottle of fluid nutrient medium.Condition of culture：3-5 days low light culture (light before cultivating
It is less than 1000Lx according to intensity), later period strong optical culture (intensity of illumination is greater than 2000Lx, is less than 3000Lx), cultivation temperature is 25 ± 2
DEG C, it can seedling after culture 30-40d.
Embodiment 1：
(1) stock variety：V. amurensis, 110R (110Richter), 140R (140Ruggeri), 5BB (Kober 5BB),
SO4(Selection Oppenheim 4)。
(2) scion variety：PINOT NOIR, Cabernet Sauvignon.
(3) Stockscion combination：PINOT NOIR-V. amurensis, Cabernet Sauvignon-V. amurensis, Cabernet Sauvignon -110R, Cabernet Sauvignon -140R, red rosy clouds
Pearl -5BB, Cabernet Sauvignon-SO4.
(2) specific steps：
1. the preparation and packing of culture medium：According to culture medium prescription：1/2B5(a large amount of)+B5(molysite)+B5(micro)+B5
(organic)+indolebutyric acid+sucrose 30gL-1PH6.0 prepares culture medium, and is sub-packed in the glass culture of diameter 6cm, height 12cm
In bottle, the culture medium of every bottle of packing 50mL, cooling is spare after high pressure sterilization.
2. the preparation of aluminium foil clamp and chalina supporter：Selecting with a thickness of the aluminium foil of 1.2mm is material, is cut into
Long 2cm, the small item of the rectangle of wide 0.5cm are folded in half into U-shaped card, spare after high pressure sterilization；Mature chalina is chosen, after drying
Deseeding is cut into the square tiles that side length is 4cm, is put into vial, adds appropriate distilled water, high pressure is gone out after sealing
Bacterium is spare.
3. the preparation and grafting of grape rootstock, scion：
The preparation of stock：Using stock varieties tissue-cultured seedling such as V. amurensis, 110R, 140R, 5BB, SO4 for cultivating 35d as material,
Remove the tiny part in upper end and lower end lignifying part, leaves and takes the middle stem section (rugosity is greater than 1mm) that length is 2~3cm and make
For stock, and blade and axillary bud all in stem section are weeded out with scalpel；
The preparation of scion：Using scion varieties tissue-cultured seedling such as the PINOT NOIRs, Cabernet Sauvignon, Chardonnay of cultivating 35d as material, band is taken
There is the stem section of full axillary bud, remove blade, an axillary bud is only stayed on each stem section top, and stem section length is greater than 1cm, and rugosity is greater than
1mm；
Grafting：According to the requirement of different Stockscion combinations, aseptically, cut with scalpel in ready stock upper end
The notch for opening 0.5cm or so, ready scion lower end chamfer an inclined-plane, 30 degree of bevel angle, then by scion
The notch of beveled end insertion stock.
4. grafting is fixed：
It is clipped in the graft union of grafting with the U-typed aluminium foil clamp after sterilizing, gently oppresses U-shaped card with tweezers, makes U-shaped
Clamp is fixedly secured graft union.
5. Liquid Culture：The tissue-cultured seedling grafted is inserted on spare chalina, it is then whole to be put into ready liquid
It is normally cultivated on body culture medium.Condition of culture：Cultivate preceding 5 days low light cultures (intensity of illumination 1000Lx), later period strong light training
It supports (intensity of illumination is greater than 2500Lx), cultivation temperature is 25 ± 2 DEG C, can seedling after culture 40d.(aforesaid liquid culture medium tool
Body ingredient see the table below)
By the implementation of this example, the miniature grafting shoot survival percent of grape reaches 95%, and it is fast to take root, and shifts to an earlier date compared with solid culture
2 days；Rooting rate reaches 92%, and 3.6 roots of mean elements, root growth is vigorous, and root volume improves 15% compared with solid culture.
Claims (3)
1. a kind of method of suitable grape micrografting seedling Liquid Culture, which is characterized in that include the following steps：
(1) using the stock variety tissue-cultured seedling for cultivating 30-40 days as material, remove the tiny part in upper end and lower end lignifying portion
Point, leaving and taking length, the middle stem section greater than 1mm is weeded out in stem section with scalpel and is owned as stock for 2~3cm and rugosity
Blade and axillary bud；
(2) it using the scion variety tissue-cultured seedling for cultivating 30-40 days as material, takes the stem section with full axillary bud as scion, removes
An axillary bud is only stayed on blade, each stem section top, it is desirable that scion length is greater than 1cm, and rugosity is greater than 1mm；
(3) aseptically, with scalpel, cutting for a 0.4-0.6cm is cut in ready stock upper end in step (1)
Mouthful, scion lower end ready in step (2) is chamfer into an inclined-plane, then bevel angle≤30 degree insert scion beveled end
Enter the notch of stock, forms grafting；
(4) aluminium foil with a thickness of 1-1.2mm is selected, aluminium foil is cut into the small item of rectangle of long 2cm wide 0.5cm, then doubling
It at U-shaped card, U-shaped card will be prepared is placed in closed container resistant to high temperature and carry out high pressure sterilization, with the U-shaped card after sterilizing
Folder in step (3) in grafting graft union on, so that U-shaped card is fixedly clamped grafting with tweezers compressing U-shaped card both ends
Graft union；
(5) mature chalina is chosen, rear deseeding is dried, the square tiles that side length is 4-5cm is then cut into, is put into resistance to
In the closed container of high temperature, a small amount of distilled water is added, high pressure sterilization is spare after sealing；
(6) on ready chalina, miniature grafting will then will be inserted in the miniature grafting inserting step (4) grafted
Chalina be put into the culture bottle for filling 50ml fluid nutrient medium and cultivate.
2. a kind of method of suitable grape micrografting seedling Liquid Culture as described in claim 1, it is characterised in that：Step (6)
Middle culture bottle diameter 6cm, height 12cm, fluid nutrient medium group become：1/2B5A great number of elements+B5Molysite+B5Microelement+B5Have
Machine+0.2~0.4mgL of indolebutyric acid-1+ sucrose 30gL-1, pH6.0.
3. a kind of method of suitable grape micrografting seedling Liquid Culture as described in claim 1, it is characterised in that：Step (6)
Middle condition of culture is to cultivate preceding 5 days low light cultures, it is desirable that intensity of illumination 1000Lx, later strong optical culture, it is desirable that intensity of illumination is big
In 2500Lx, cultivation temperature is 25 ± 2 DEG C, can seedling after culture 40d.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201810347558 | 2018-04-18 | ||
| CN2018103475585 | 2018-04-18 |
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| CN108848994A true CN108848994A (en) | 2018-11-23 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115136805A (en) * | 2022-08-09 | 2022-10-04 | 西北农林科技大学 | Double-stock micro-grafting method for grapes |
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|---|---|---|---|---|
| CN103782811A (en) * | 2014-02-26 | 2014-05-14 | 武汉市农业科学研究所 | Watermelon tissue culture seedling test tube micro-grafting method |
| CN105993633A (en) * | 2016-07-21 | 2016-10-12 | 江门市新会区林业科学研究所 | Xinhui mandarin orange virus-free test tube micro-grafting method |
| CN106417025A (en) * | 2016-10-14 | 2017-02-22 | 山西农业大学 | Test tube micro-grafting method for seedless grape embryo-rescued malformed plantlets |
-
2018
- 2018-07-30 CN CN201810853915.5A patent/CN108848994A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103782811A (en) * | 2014-02-26 | 2014-05-14 | 武汉市农业科学研究所 | Watermelon tissue culture seedling test tube micro-grafting method |
| CN105993633A (en) * | 2016-07-21 | 2016-10-12 | 江门市新会区林业科学研究所 | Xinhui mandarin orange virus-free test tube micro-grafting method |
| CN106417025A (en) * | 2016-10-14 | 2017-02-22 | 山西农业大学 | Test tube micro-grafting method for seedless grape embryo-rescued malformed plantlets |
Non-Patent Citations (2)
| Title |
|---|
| 刘玉晗: "田间成年"碧秀"苦瓜优株离体快繁体系研究", 《中国期刊网全文数据库农业科技辑》 * |
| 葡萄试管苗嫁接方法研究: "赵改荣等", 《果树科学》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115136805A (en) * | 2022-08-09 | 2022-10-04 | 西北农林科技大学 | Double-stock micro-grafting method for grapes |
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