CN1255283A - Technology for culturing improved virus-resistant forestry nursery stock - Google Patents
Technology for culturing improved virus-resistant forestry nursery stock Download PDFInfo
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- CN1255283A CN1255283A CN 98123266 CN98123266A CN1255283A CN 1255283 A CN1255283 A CN 1255283A CN 98123266 CN98123266 CN 98123266 CN 98123266 A CN98123266 A CN 98123266A CN 1255283 A CN1255283 A CN 1255283A
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Abstract
A technology for culturing improved virus-resistant forestry nursery stock includes high-temp ultivation of the nursery stock infected with virus in dark condition at 35-40 deg.C for 40-60 day to exend the non-virous area at seedling top, cutting off the seedling tops of 5-10 mm in length, tissue culture for quick reproduction and shoot tip graft. Its advantages are simple operation, high virus removing rate, high survival rate and retaining all characteristics of excellent nursely stock.
Description
The invention belongs to cultivated plant breeding nursery stock technical field.
The plant (crops, fruit tree, forest, flowers etc.) of human cultivation infects mostly and has accumulated multiple virus in its long-term vegetative propagation process.Bao Dao apple virus has 39 kinds all over the world, 23 kinds in pears virus, 25 kinds in 3 kinds of grape virus-4s, strawberry virus, 39 kinds in peach virus, 28 kinds in Lee's virus, 36 kinds in sweet cherry virus, 21 kinds in sour cherry virus, 19 kinds in apricot virus, 20 kinds in citrus virus, also have banana virus, Potyvirus, tulip virus, sword lily virus, ficus virus etc.These viruses have been disturbed normal physiological metabolism activity in plant cell, cause that the plant corpus growth weakens, early ageing, plant is downgraded, the blade variable color, curl, shrinkage, yellow, the fruit deformity diminishes, and is not painted, sugar content reduces, fallen flowers, shedding make cultivated plant output underproduction 20%-100%, bring enormous economic loss for human agricultural production.In modern high-efficiency high-quality agricultural economy, the cultivated plant virus disease has caused global great attention.The seventies it is found that in the past the branch of cultivated plant is placed under the hot conditions and heat-treated, and can make the passivation of part virus, temporarily lost biologically active, can reduce the extent of injury of virus disease to plant corpus.But this heat treatment method is not thoroughly removed virus in plant corpus, can spread again in plant corpus again through virus after the long period.Particularly concerning those perennial plants, this heat treatment method poor effect.People find that again it is uneven that virus distributes in plant corpus later on.There is not virus in the stem apex of plant and the meristem zone of the tip of a root and this tiny area of elongation zone.So people's appliable plant method for tissue culture, the stem apex that cutting the 0.2-0.5 millimeter does not have a vascular bundle tissue carries out tissue culture, has obtained the nursery stock of virus-free.Need carry out work under the microdissection mirror but the stem apex that this stem apex de-virusing method cuts is very little, concrete operations are very difficult, and Shoot Tip Culture survival rate, virus elimination rate are also very low.After entering the nineties, people combine heat treatment method and Shoot Tip Culture method again and carry out virus-free.Plant stem apex (as potted plant nursery stock or test-tube plantlet) that earlier will detoxification is placed on 35-40 ℃ of condition thermal treatment 1 wheat harvesting period, and at high temperature virus is passivated, and its breeding spread speed in plant corpus is subtracted greatly.The new plant stem apex that grows in this hot environment is taken off virus probably, can cut long stem apex 1-3 millimeter and cultivate, and can obtain the virus-free nursery stock.What cultivate the employing of virus-free breeding nursery stock in producing at present is above-mentioned this method.In the actual production operation, cut bigger stem apex (as 3 millimeters long) and cultivate the survival rate height, but virus elimination rate is very low; It is higher to cut less stem apex (in 1 millimeter) virus elimination rate, but survival rate is very low, and cultivates the detoxification test tube plantlet that survives and also might morph, and changes the proterties of improved seeds.So these problems are perplexing the operate as normal that people cultivate virus-free breeding nursery stock all the time in actual production.
The object of the invention is exactly at above-mentioned situation, adopt complete berlin black dark condition that nursery stock is heat-treated cultivation, expansion at double the virus-free zone of stem apex, cutting stem apex again cultivates, promptly can remove virus fully, improve virus elimination rate, can cut long stem apex again, improve the Shoot Tip Culture survival rate, prevent the variation of breeding nursery stock.The method of a kind of new high-efficiency breeding virus-free breeding nursery stock is provided for actual production.
A kind of method of cultivating virus-free breeding nursery stock of the present invention is: nursery stock that earlier will virus-free is adopted the method for Plant Tissue Breeding, they is cultivated into test-tube plantlet, the seedling in the stage of preferably taking root or the seedling in the bud shape stage of growing thickly in vitro.This eugonic test-tube plantlet is placed on heat-treat under the condition of complete unglazed dark and cultivated 45-60 days.Heat treatment temperature is 35-40 ℃.Potted plant nursery stock (preferably just having passed through the nursery stock of dormant stage) that perhaps will virus-free is placed on to heat-treat under the condition of complete unglazed dark and cultivated 45-50 days, and heat treatment temperature is 35-40 ℃.After nursery stock was handled through 45-60 days berlin black dark heat, the test-tube plantlet stem apex can grow the flaxen almost vaneless newborn stem apex of 1-4 centimeter length; Potted plant nursery stock can grow 5-10 centimetre the almost vaneless newborn stem apex in flaxen top.These newborn stem apexs are carried out vertical paraffin section observe discovery: the meristem zone and the elongation zone of the newborn stem apex of test-tube plantlet, these no microtubule fasolculus section length can reach more than the 5-10 millimeter; The meristematic zone and elongation zone (the no microtubule fasolculus district) length of the newborn stem apex of potted plant nursery stock can reach more than the 10-15 millimeter.Virus is mainly propagated by microtubule fasolculus in plant corpus, microtubule fasolculus can be sent to the virus in the maturation plant body of bottom microtubule fasolculus district and elongation zone intersection, stem apex meristematic zone and elongation zone do not have microtubule fasolculus, so stem apex meristematic zone and elongation zone belong to virus-free zone on the plant corpus.Handle cultivation by the berlin black dark heat, stoped normal photomorphogenesis process in the plant corpus, make the meristematic zone of stem apex and the cell differentiation of elongation zone become the process of microtubule fasolculus to be slowed down, so this virus-free zone of meristematic zone and elongation zone is extended.Add 35-40 ℃ long term thermal and handle, the virus in the plant corpus is passivated, temporarily lose vegetable active, irreproducible and quick propagation.So through the newborn stem apex of thermal treatment nursery stock under the berlin black dark condition, virus-free zone is extended.Characteristics of the present invention are that long term thermal is handled nursery stock under the berlin black dark condition, can greatly enlarge the virus-free zone of newborn stem apex.
A kind of method of cultivating virus-free breeding nursery stock of the present invention is: article one approach according to method for plant tissue culture, at the aseptic condition incision long 5-8 millimeter of newborn stem apex that the berlin black dark heat handles of learning from else's experience, is inoculated on the Shoot Tip Culture base of new preparation.In intensity of illumination 1500-3000 lux, cultivate under the temperature 25-28 ℃ of condition, make the stem apex growth, differentiation.Enlarge enrichment culture repeatedly through several generations, the clone nursery stock of each stem apex is carried out virus detection.Adopt indicator plant detection method, ELISA, the electron microscope detection method, after determining the complete virus-free of this clone nursery stock, appliable plant tissue culture quick propagation culturing method is bred virus-free breeding nursery stock, for production provides virus-free breeding nursery stock in enormous quantities.The second approach through the potted plant nursery stock that the berlin black dark heat is handled, can cut the stem apex of 10-12 millimeters long, grafting to the virus-free stock of preprepared, stock of each stem apex grafting.After the stem apex scion survives growth, each stem apex nursery stock is carried out virus detects, determine that this stem apex nursery stock is virus-free after, be parent with this virus-free breeding nursery stock, a large amount of propagation by grafiting virus-free breeding nursery stocks are carried out in its shoot row culture scion of clip.Characteristics of the present invention are to cut the stem apex through berlin black dark heat processing test-tube plantlet of 5-8 millimeters long, breed virus-free breeding nursery stock in a large number by the tissue culture quick propagation culturing method; Cut the potted plant nursery stock stem apex through the processing of berlin black dark heat of 10-12 millimeters long, breed virus-free breeding nursery stock in a large number by grafting method.
Adopt the inventive method to cultivate virus-free breeding nursery stock, have very advantages of higher of simple, the easy row of method of operating, virus-free rate and survival rate, effect is good in actual production.Take off grapevine fanleaf virus, Potato Leaf Roll Virus (PLRV), stem poxvirus with the inventive method, cork virus, after test-tube plantlet is handled cultivation through the berlin black dark heat, can directly cut the stem apex of 8 millimeters long, the stem apex that cuts 0.3 millimeters long need do not operated under the microdissection mirror, so operation is simple for the inventive method.The stem apex of such 8 millimeters long is inoculated on the medium survival rate all 100%.Detect virus with indicator plant, the virus-free rate is 100% as a result.So the inventive method virus-free rate height, the survival rate height.After the virus-free grape breeding nursery stock that cultivates was cultivated the result aborning, grape fruit increased, and sugariness obviously improves, the 70-80% of output volume increase as a result, and remarkable in economical benefits increases.
Embodiment 1:
To have the potted plant Kuerle delicious pear of pears stone poxvirus winter through the infection of resting stage, be placed under the complete berlin black dark condition, and thermal treatment was cultivated 60 days in 38 ℃ ± 1 ℃ environment.To water regularly for potted plant bergamot pear, prevent to cause nursery stock death because of lack of water.In the greenhouse, sow virus-free birch-leaf pear seed simultaneously, cultivate the birch-leaf pear nutrition pot seedling and make graft stock.Cut bergamot pear stem apex 10 millimeters long that the berlin black dark heat is handled, will notice during operation that the cutter apparatus does not contact with stem apex bottom tissue, in order to avoid cross-infection virus.To birchleaf pear rootstock, the bergamot pear stem apexs begin to sprout, grow the employing cleft graft after 2 weeks with the grafting of bergamot pear stem apex, and the stem apex graft survival rate is 80%, stock of each stem apex grafting, and number in order.Carried out virus after spring and summer, bergamot pear stem apex grafting grew up and detect, can adopt the serology fast detection method, the nursery stock that testing result removes pears stone poxvirus is 95%.The nontoxic Kuerle delicious pear breeding nursery stock that cultivation is obtained is adopted its twig and does scion as adopting the fringe maternal plant, breeds detoxification bergamot pear nursery stock in a large number, provides nontoxic breeding nursery stock for producing cultivation.
Embodiment 2:
Behind the Hami date infective virus, date diminishes, and obvious minus green patch appears in blade, and fruit drop is serious, and output declines to a great extent.With Hami date of infective virus,, turn out the test tube bud of growing thickly by method for plant tissue culture.This Hami date test-tube plantlet is placed under the condition of complete unglazed dark, and thermal treatment was cultivated 50 days in 38 ℃ ± 0.5 ℃ environment.Hami date test-tube plantlet faint yellow on-bladed stem apex of 2-3 centimeter length that regrows out at this time.On the clean work station of desinfection chamber, directly cut the stem apex of 8 millimeters long, the cutter apparatus does not contact stem apex bottom tissue during operation, prevents the new stem-tip tissue of viral cross-infection.Each stem apex is inoculated into respectively in the new medium in the triangular flask, and the stem apex in each triangular flask is numbered.The Shoot Tip Culture base is the 6-base aminopurine+0.02 mg/litre methyl of kinetin+1.0 mg/litre of MS medium+1.0 mg/litre.In intensity of illumination 1500-3000 lux, under the temperature 25-30 ℃ of condition, to cultivate 1 all stem apexs and begin to sprout, grow, the Shoot Tip Culture survival rate is 100%.Through several generations induce enrichment culture, each stem apex can obtain many test tubes bud of growing thickly.Adopt ELISA that test-tube plantlet is carried out virus and detect, the virus-free rate is 100% as a result.By the plant tissue culture method for quickly breeding, produce virus-free breeding jujube seedling in large quantity then, provide nontoxic breeding nursery stock for producing cultivation.
Claims (5)
1. a method of cultivating virus-free breeding nursery stock is characterized in that being placed on earlier under the condition of complete unglazed dark wanting the nursery stock of virus-free, heat-treats and cultivates a period, cuts long stem apex again, turns out virus-free breeding nursery stock.
2. the method for cultivation virus-free breeding nursery stock according to claim 1 is characterized in that: the nursery stock of virus-free can be a test-tube plantlet, also can be potted plant nursery stock.
3. the method for cultivation virus-free breeding nursery stock according to claim 1 is characterized in that: heat treatment temperature is 35-40 ℃, and heat treatment time is 40-60 days.
4. the method for cultivation virus-free breeding nursery stock according to claim 1 is characterized in that: the length that cuts the test-tube plantlet stem apex is the 5-8 millimeter, and the length that cuts potted plant seedling stem apex is the 10-12 millimeter.
5. the method for cultivation virus-free breeding nursery stock according to claim 1, it is characterized in that: the stem apex that cuts can be cultivated the virus-free nursery stock by method for plant tissue culture, also can be by the stem apex grafting is cultivated the virus-free nursery stock to the method on the virus-free stock.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 98123266 CN1255283A (en) | 1998-11-29 | 1998-11-29 | Technology for culturing improved virus-resistant forestry nursery stock |
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| CN 98123266 CN1255283A (en) | 1998-11-29 | 1998-11-29 | Technology for culturing improved virus-resistant forestry nursery stock |
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| CN1255283A true CN1255283A (en) | 2000-06-07 |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102232358A (en) * | 2010-05-04 | 2011-11-09 | 河北省农林科学院昌黎果树研究所 | Detoxification method for excised stem segment of grape |
| CN106613967A (en) * | 2016-12-01 | 2017-05-10 | 大连大学 | Explant for detoxification of plant tissue culture and detoxification method of explant |
| CN107409669A (en) * | 2017-04-22 | 2017-12-01 | 蚌埠市乔峰农业蔬菜专业合作社 | A kind of sweet potato is heat-treated poison-removing method |
| CN111226790A (en) * | 2020-01-07 | 2020-06-05 | 徐世彦 | Detoxification method for apple virus diseases |
-
1998
- 1998-11-29 CN CN 98123266 patent/CN1255283A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102232358A (en) * | 2010-05-04 | 2011-11-09 | 河北省农林科学院昌黎果树研究所 | Detoxification method for excised stem segment of grape |
| CN102232358B (en) * | 2010-05-04 | 2012-09-05 | 河北省农林科学院昌黎果树研究所 | Detoxification method for excised stem segment of grape |
| CN106613967A (en) * | 2016-12-01 | 2017-05-10 | 大连大学 | Explant for detoxification of plant tissue culture and detoxification method of explant |
| CN107409669A (en) * | 2017-04-22 | 2017-12-01 | 蚌埠市乔峰农业蔬菜专业合作社 | A kind of sweet potato is heat-treated poison-removing method |
| CN111226790A (en) * | 2020-01-07 | 2020-06-05 | 徐世彦 | Detoxification method for apple virus diseases |
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