CN102144547A - Method for quickly breeding and transplanting grape stock unit - Google Patents
Method for quickly breeding and transplanting grape stock unit Download PDFInfo
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- CN102144547A CN102144547A CN201110007348XA CN201110007348A CN102144547A CN 102144547 A CN102144547 A CN 102144547A CN 201110007348X A CN201110007348X A CN 201110007348XA CN 201110007348 A CN201110007348 A CN 201110007348A CN 102144547 A CN102144547 A CN 102144547A
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Abstract
The invention discloses a technology for quickly breeding and transplanting a grape stock unit. The technology establishes a quick-breeding system of the grape stock unit through the isolated culture of grape stocks, and comprises material selection, material washing, culture starting, augmentation culture, rooting culture and seedling transplant, wherein the survive rate can reach above 90%. The establishment of the method for quickly breeding and transplanting the grape stock unit solves the problems that the grape stocks have hybrid sterility and cultured seedlings are hard to survive, and the method is suitable for commercially producing grape stock seedlings.
Description
Technical field
The present invention relates to grape rootstock crossbreed tissue-culturing rapid propagation and transplanting method, belong to biological technical field, be specially a kind of grape rootstock tissue-culturing rapid propagation and transplanting method.
Background technology
In recent years, China's grape industry development is swift and violent, and Table Grape output occupies first place in the world, and the wine grape scale occupies the 5th in the world.On planting type, present by traditional own-rooted tree to the grafting development trend of transition gradually, screen and introduced many good stocks, comprising the SO of wide adaptability
4With kinds such as Bei Da.Grape rootstock SO
4(Selection Oppenheim NO.4) is the state-run grape fruit tree of German Oppenheim research institute, forms with winter grape * riverside grape seed selection.Tree vigo(u)r is strong, and comprehensive proterties tool such as cold-resistant, drought resisting, moisture-resistant waterlogging is good, and grafting affinity is strong.The grape rootstock shellfish reaches that (Vitis riparia * V.labrusca) is the America kind, originates in the U.S., is the crossbreed of fox grape and riverside grape.Tree vigo(u)r is strong, cold-resistant, drought-enduring, waterlogging, barren-resistant, easily infected virus not, and grafting affinity is strong.Because the development of grape grafting cultivation, supply falls short of demand for good stock seedling, and therefore, the reproductive efficiency that the raising grape rootstock is grown seedlings is very necessary.Group culturation rapid propagating technology just in time solves this difficult problem.
Summary of the invention
The present invention provides and can improve the reproductive efficiency that grape rootstock is grown seedlings just at above technical problem, and a kind of grape rootstock tissue-culturing rapid propagation and the transplanting method of effective way is provided for the scale breeding of grape rootstock.
Concrete technical scheme of the present invention is as follows:
A kind of grape rootstock tissue culture and rapid propagation method may further comprise the steps:
(A) material is selected: gathering grape rootstock semi-lignified shoot is explant, removes blade, stays petiole, and branch is cut into 5 ~ 6cm length;
(B) material cleans: running water flowing water flushing branch is after 1 hour, with 75% alcohol-pickled 30 seconds, uses 0.1% HgCl again
2Sterilization 7 ~ 8min, sterile water wash 3 times cuts into the stem section of being with internode about 2cm after the sterilization;
(C) start to cultivate: with 1/2MS or MS or GS is minimal medium, add the various combination of indolebutyric acid (IBA), methyl (NAA) and the 6-benzylaminopurine (6-BA) of variable concentrations therein, be mixed with startup medium ⑴-⑺ number of grape rootstock cultured in vitro, wherein indolebutyric acid concentration is: 0.1 mg/L, 0.2 mg/L and 0.3 mg/L; Methyl concentration is: 0.02 mg/L, 0.1 mg/L and 0.2 mg/L; 6-benzylaminopurine concentration is: 0.01 mg/L and 2.0 mg/L; Stem section after handling is inoculated in starts for following ⑴-⑺ number on the medium:
（1）1/2MS?+?IBA?0.1?mg/L
（2）1/2MS?+?IBA?0.2?mg/L
（3）1/2MS?+?IBA?0.3?mg/L
（4）MS?+?6-BA?2.0?mg/L?+?NAA?0.2?mg/L
（5）MS?+?6-BA?2.0?mg/L?+?NAA?0.02?mg/L
（6）GS?+?IBA?0.1?mg/L?+?NAA?0.1?mg/L
（7）GS?+?IBA?0.1?mg/L?+?NAA?0.1?mg/L?+?6-BA?0.01?mg/L
Start on the medium in (1)-(7) number, the result represents: existing bud has the differentiation of root again on 25 days left and right sides ⑴-⑶ medium, and best of breed is ⑵ 1/2MS+IBA 0.2 mg/L medium; ⑷ produce the bud of growing thickly after inducing generation callus subculture on and the ⑸ medium earlier; ⑹ and ⑺ effect are undesirable;
(D) enrichment culture: the callus of inducing on ⑷ and the ⑸ medium transferred in ⑷ MS+6-BA 2.0 mg/L+NAA 0.2 mg/L medium about 35 days can produce the bud of growing thickly;
(E) culture of rootage: behind the bud height of seedling 2 ~ 3cm of growing thickly that treats to grow in ⑷ MS+6-BA 2.0 mg/L+NAA 0.2 mg/L medium, change over to and cultivate the generation that root was just arranged in about 25 days in ⑵ 1/2MS+IBA 0.2 mg/L medium;
(F) an one-step inducing Cheng Miao: the grape rootstock stem section primary vaccination after the sterilization treatment cultivated 30 days in ⑵ 1/2MS+IBA 0.2 mg/L medium after, not only emerge but also take root.
Add the sucrose of 25g/L in described startup cultivation, enrichment culture and the root media respectively, agar with 5.5 ~ 6g/L solidifies, the pH value is 5.8, and places the culturing room of 25 ± 2 ℃ of temperature, light application time 14h/d, intensity of illumination 1500-2000lx to cultivate postvaccinal culture materials.
In described grape rootstock cultured in vitro, what the material washing and sterilizing time can be according to material is tender always, suitably adjusts 0.1% HgCl
2And sterilization time.
A kind of grape rootstock transplanting method, with grape rootstock bottle seedling plant up to more than the 5cm, the aseptic seedling of the long 3-8cm of root, radical 5-7 root is indoor (or in booth) natural daylight lower refining seedling 3-5 days, after the uncork hardening again 7 days, the agar of getting on the clean root of seedling bottle outlet is transplanted in the perlite, the lid plastic film keeps blade humidity, avoid the high light direct projection, about 25 ℃ of room temperatures.Heel in 15-20 days in the perlite, but just transplant planting is in loose ground, survival rate behind the slow seedling, can spray 0.2% foliage fertilizer more than 90%.
In described grape rootstock transplanting process, test-tube plantlet bottle outlet rear blade is preserved moisture.
Good effect of the present invention is embodied in: can improve the reproductive efficiency that grape rootstock is grown seedlings, for the scale breeding of grape rootstock provides effective way.
Embodiment
The present invention will be further described according to specific embodiment below.
Embodiment one:
Gather grape rootstock SO the morning on May 5th, 2009
4Reach shoot with shellfish, remove blade, stay petiole, cut into the long branch of 5 ~ 6cm, the running water flushing is standby after 1 hour.The explant that cleans up is placed on the superclean bench, and material was used 0.1% HgCl again with 75% alcohol-pickled 30 seconds
2Sterilization 7 ~ 8min, sterile water wash 3 times cuts into it stem section of being with internode about 2cm then, is inoculated in to start on the medium 25 ± 2 ℃ of cultivation temperature, light application time 14h/d, intensity of illumination 1500 ~ 2000lx for ⑴-⑺ number.
With the callus of inducing on ⑷ and the ⑸ medium, switching is as on ⑷ MS+6-BA 2.0 mg/L+NAA 0.2 mg/L medium, and cultivation can produce the bud of growing thickly in about 35 days under 25 ± 2 ℃ of temperature, light application time 14h/d, intensity of illumination 1500 ~ 2000lx.
Behind the bud height of seedling 2 ~ 3cm of growing thickly to be grown, change over to and cultivate the generation that root was just arranged in about 25 days in ⑵ 1/2MS+IBA 0.2 mg/L medium.Condition of culture is cultivated with starting.
Grape rootstock one one-step inducing Cheng Miao, grape rootstock is seeded in ⑵ 1/2MS+IBA 0.2 mg/L medium and cultivates after 30 days, not only emerges but also take root.Condition of culture is cultivated with starting.
With more than the height of seedling 5cm in the bottle, the aseptic seedling of the long 3-8cm of root, radical 5-7 root is indoor (or in booth) natural daylight lower refining seedling 3-5 days, after the uncork hardening again 7 days, the agar of getting on the clean root of seedling bottle outlet is transplanted in the perlite, the lid plastic film keeps blade humidity, avoid the high light direct projection, about 25 ℃ of room temperatures.Heel in 15-20 days in the perlite, but just transplant planting is in loose ground, survival rate behind the slow seedling, can spray 0.2% foliage fertilizer more than 90%.
Embodiment two:
Gather grape rootstock SO the morning on July 1st, 2009
4Reach the semi-lignified shoot with shellfish, remove blade, stay petiole, cut into the long branch of 5 ~ 6cm, the running water flushing is standby after 1 hour.The explant that cleans up is placed on the superclean bench, and material was used 0.1% HgCl again with 75% alcohol-pickled 30 seconds
2Sterilization 7 ~ 8min, sterile water wash 3 times cuts into it stem section of being with internode about 2cm then, is inoculated in ⑵ 1/2MS+IBA 0.2 mg/L+25g/L sucrose+agar 5.5g/L medium and cultivates after 30 days, not only emerges but also take root.
Treat that its test-tube plantlet grows to more than the 5cm in the ⑵ medium after, cut off big blade, keep vanelets, and cut into the stem section of band internode about 2-3cm, continue to transfer in the ⑵ medium, can become seedling about 25 days.Cultivation cycle is about 25 days, 25 ± 2 ℃ of cultivation temperature, light application time 14h/d, intensity of illumination 1500 ~ 2000lx.
More than height of seedling 5cm in the bottle, the long 3-8cm of root, the aseptic seedling of radical 5-7 root was indoor (or in booth) natural daylight lower refining seedling 3-5 days, after the uncork hardening again 7 days, the agar of getting on the clean root of seedling bottle outlet is transplanted in the perlite, the lid plastic film keeps blade humidity, avoids the high light direct projection, about 25 ℃ of room temperatures.Heel in 15-20 days in the perlite, but just transplant planting is in loose ground, survival rate behind the slow seedling, can spray 0.2% foliage fertilizer more than 90%.
Embodiment three:
Gather grape rootstock SO the morning on June 20th, 2010
4Reach the semi-lignified shoot with shellfish, remove blade, stay petiole, cut into the long branch of 5 ~ 6cm, the running water flushing is standby after 1 hour.The explant that cleans up is placed on the superclean bench, and material was used 0.1% HgCl again with 75% alcohol-pickled 30 seconds
2Sterilization 7 ~ 8min, sterile water wash 3 times cuts into it stem section of being with internode about 2cm then, is inoculated in ⑵ 1/2MS+IBA 0.2 mg/L+25g/L sucrose+agar 5.5g/L medium and cultivates after 30 days, not only emerges but also take root.
Treat that its test-tube plantlet grows to more than the 5cm in the ⑵ medium after, cut off big blade, keep vanelets, and cut into the stem section of band internode about 2-3cm, continue to transfer in the ⑵ medium, can become seedling about 25 days.Cultivation cycle is about 25 days.25 ± 2 ℃ of cultivation temperature, light application time 14h/d, intensity of illumination 1500 ~ 2000lx.
With more than the height of seedling 5cm in the bottle, the aseptic seedling of the long 3-8cm of root, radical 5-7 root is indoor (or in booth) natural daylight lower refining seedling 3-5 days, after the uncork hardening again 7 days, the agar of getting on the clean root of seedling bottle outlet is transplanted in the perlite, the lid plastic film keeps blade humidity, avoid the high light direct projection, about 25 ℃ of room temperatures.Heel in 15-20 days in the perlite, but just transplant planting is in loose ground, survival rate behind the slow seedling, can spray 0.2% foliage fertilizer more than 90%.
Claims (5)
1. grape rootstock tissue culture and rapid propagation method is characterized in that may further comprise the steps:
(A) material is selected: gathering grape rootstock semi-lignified shoot is explant, removes blade, stays petiole, and branch is cut into 5 ~ 6cm length;
(B) material cleans: running water flowing water flushing branch is after 1 hour, with 75% alcohol-pickled 30 seconds, uses 0.1% HgCl again
2Sterilization 7 ~ 8min, sterile water wash 3 times cuts into the stem section of being with internode about 2cm after the sterilization;
(C) start to cultivate: with 1/2MS or MS or GS is minimal medium, add the various combination of indolebutyric acid (IBA), methyl (NAA) and the 6-benzylaminopurine (6-BA) of variable concentrations therein, be mixed with startup medium ⑴-⑺ number of grape rootstock cultured in vitro, wherein indolebutyric acid concentration is: 0.1 mg/L, 0.2 mg/L and 0.3 mg/L; Methyl concentration is: 0.02 mg/L, 0.1 mg/L and 0.2 mg/L; 6-benzylaminopurine concentration is: 0.01 mg/L and 2.0 mg/L; Stem section after handling is inoculated in starts for following ⑴-⑺ number on the medium;
（1）1/2MS?+?IBA?0.1?mg/L
（2）1/2MS?+?IBA?0.2?mg/L
（3）1/2MS?+?IBA?0.3?mg/L
（4）MS?+?6-BA?2.0?mg/L?+?NAA?0.2?mg/L
（5）MS?+?6-BA?2.0?mg/L?+?NAA?0.02?mg/L
（6）GS?+?IBA?0.1?mg/L?+?NAA?0.1?mg/L
（7）GS?+?IBA?0.1?mg/L?+?NAA?0.1?mg/L?+?6-BA?0.01?mg/L
The result shows: existing bud has the differentiation of root again on 25 days left and right sides ⑴-⑶ medium, and best of breed is ⑵ 1/2MS+IBA 0.2 mg/L medium; ⑷ produce the bud of growing thickly after inducing generation callus subculture on and the ⑸ medium earlier; ⑹ and ⑺ effect are undesirable;
(D) enrichment culture: the callus of inducing on ⑷ and the ⑸ medium transferred in ⑷ MS+6-BA 2.0 mg/L+NAA 0.2 mg/L medium about 35 days can produce the bud of growing thickly;
(E) culture of rootage: behind the bud height of seedling 2 ~ 3cm of growing thickly that treats to grow in ⑷ MS+6-BA 2.0 mg/L+NAA 0.2 mg/L medium, change over to and cultivate the generation that root was just arranged in about 25 days in ⑵ 1/2MS+IBA 0.2 mg/L medium;
(F) an one-step inducing Cheng Miao: the grape rootstock stem section primary vaccination after the sterilization treatment cultivated 30 days in ⑵ 1/2MS+IBA 0.2 mg/L medium after, not only emerge but also take root.
2. a kind of grape rootstock tissue culture and rapid propagation method according to claim 1, it is characterized in that: the sucrose that adds 25g/L in described startup cultivation, enrichment culture and the root media respectively, agar with 5.5-6g/L solidifies, the pH value is 5.8, and places the culturing room of 25 ± 2 ℃ of temperature, light application time 14h/d, intensity of illumination 1500-2000lx to cultivate postvaccinal culture materials.
3. grape rootstock transplanting method, it is characterized in that: with bottle seedling plant behind the grape rootstock tissue-culturing rapid propagation up to more than the 5cm, the aseptic seedling of the long 3-8cm of root, radical 5-7 root is indoor (or in booth) natural daylight lower refining seedling 3-5 days, after the uncork hardening again 7 days, the agar of getting on the clean root of seedling bottle outlet is transplanted in the perlite, the lid plastic film keeps blade humidity, avoid the high light direct projection, about 25 ℃ of room temperatures, heel in 15-20 days in the perlite, just but transplant planting is in loose ground, survival rate behind the slow seedling, can spray 0.2% foliage fertilizer more than 90%.
4. grape rootstock tissue culture and rapid propagation method according to claim 1 is characterized in that: in described grape rootstock cultured in vitro, what the material washing and sterilizing time can be according to material is tender always, suitably adjusts 0.1% HgCl
2And sterilization time.
5. grape rootstock tissue culture and rapid propagation method according to claim 1 is characterized in that: in described grape rootstock transplanting process, test-tube plantlet bottle outlet rear blade is preserved moisture.
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Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102668991A (en) * | 2012-06-14 | 2012-09-19 | 宁夏农垦西夏王实业有限公司 | Application of penicillin to simple test-tube breeding of grapes and novel technology for test-tube breeding of grapes |
| CN102763593A (en) * | 2012-07-16 | 2012-11-07 | 福建省农业科学院农业工程技术研究所 | Method for rapidly obtaining loose calluses of grapes and for long-term succeeding maintenance of grapes |
| CN103109728A (en) * | 2013-03-10 | 2013-05-22 | 通化师范学院 | Rapid seedling culturing method of pinus sylvestris in tube |
| CN104542298A (en) * | 2015-01-26 | 2015-04-29 | 武汉市林业果树科学研究所 | Grape tissue regeneration culture method |
| CN105145269A (en) * | 2015-07-12 | 2015-12-16 | 青岛乐义现代农业科技示范基地有限公司 | Efficient seedling cultivating method for leafbuds of Yunshan 2# grape novel variety |
| CN106900553A (en) * | 2017-03-13 | 2017-06-30 | 玉林师范学院 | A kind of tissue culture and rapid propagation method of medicinal and edible plant Erythropalum Scandens Blume among the people |
| CN106922539A (en) * | 2017-04-28 | 2017-07-07 | 玉林师范学院 | A kind of tissue culture and rapid propagation method of Chinese medicine cissus assamica |
| CN106937601A (en) * | 2017-04-28 | 2017-07-11 | 福建省农业科学院农业工程技术研究所 | A kind of method for improving grape test tube seedling transplanting survival rate |
| CN106982736A (en) * | 2017-04-28 | 2017-07-28 | 李茂兰 | A kind of breeding method of Tissue culture the seedling of grape |
| CN107333655A (en) * | 2017-08-17 | 2017-11-10 | 宁夏农垦西夏王实业有限公司葡萄苗木分公司 | A kind of micro- numerous screening and culturing medium of alkali resistance grape rootstock and micro- numerous screening technique |
| CN110402816A (en) * | 2019-07-25 | 2019-11-05 | 河北农业大学 | A kind of system and its application method increasing grape test tube transplantation of seedlings output of seedling |
| CN112438202A (en) * | 2019-08-28 | 2021-03-05 | 中国农业大学 | Tissue culture and rapid propagation method of grape rootstock |
| CN114698549A (en) * | 2022-04-18 | 2022-07-05 | 新疆农业大学 | Tissue culture medium and tissue culture method for rapid propagation of grape rootstock stem |
| CN116548306A (en) * | 2023-05-12 | 2023-08-08 | 山西大学 | Single-bud stem induction callus and tissue culture rapid propagation method for grape at Chardonnay |
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Cited By (18)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102668991A (en) * | 2012-06-14 | 2012-09-19 | 宁夏农垦西夏王实业有限公司 | Application of penicillin to simple test-tube breeding of grapes and novel technology for test-tube breeding of grapes |
| CN102668991B (en) * | 2012-06-14 | 2013-10-23 | 宁夏农垦西夏王实业有限公司 | Application of penicillin to simple test-tube breeding of grapes and novel technology for test-tube breeding of grapes |
| CN102763593A (en) * | 2012-07-16 | 2012-11-07 | 福建省农业科学院农业工程技术研究所 | Method for rapidly obtaining loose calluses of grapes and for long-term succeeding maintenance of grapes |
| CN103109728A (en) * | 2013-03-10 | 2013-05-22 | 通化师范学院 | Rapid seedling culturing method of pinus sylvestris in tube |
| CN103109728B (en) * | 2013-03-10 | 2013-12-18 | 通化师范学院 | Rapid seedling culturing method of pinus sylvestris in tube |
| CN104542298A (en) * | 2015-01-26 | 2015-04-29 | 武汉市林业果树科学研究所 | Grape tissue regeneration culture method |
| CN105145269A (en) * | 2015-07-12 | 2015-12-16 | 青岛乐义现代农业科技示范基地有限公司 | Efficient seedling cultivating method for leafbuds of Yunshan 2# grape novel variety |
| CN106900553A (en) * | 2017-03-13 | 2017-06-30 | 玉林师范学院 | A kind of tissue culture and rapid propagation method of medicinal and edible plant Erythropalum Scandens Blume among the people |
| CN106922539A (en) * | 2017-04-28 | 2017-07-07 | 玉林师范学院 | A kind of tissue culture and rapid propagation method of Chinese medicine cissus assamica |
| CN106937601A (en) * | 2017-04-28 | 2017-07-11 | 福建省农业科学院农业工程技术研究所 | A kind of method for improving grape test tube seedling transplanting survival rate |
| CN106982736A (en) * | 2017-04-28 | 2017-07-28 | 李茂兰 | A kind of breeding method of Tissue culture the seedling of grape |
| CN106937601B (en) * | 2017-04-28 | 2019-09-20 | 福建省农业科学院农业工程技术研究所 | A method of improving grape test tube seedling transplanting survival rate |
| CN107333655A (en) * | 2017-08-17 | 2017-11-10 | 宁夏农垦西夏王实业有限公司葡萄苗木分公司 | A kind of micro- numerous screening and culturing medium of alkali resistance grape rootstock and micro- numerous screening technique |
| CN110402816A (en) * | 2019-07-25 | 2019-11-05 | 河北农业大学 | A kind of system and its application method increasing grape test tube transplantation of seedlings output of seedling |
| CN112438202A (en) * | 2019-08-28 | 2021-03-05 | 中国农业大学 | Tissue culture and rapid propagation method of grape rootstock |
| CN114698549A (en) * | 2022-04-18 | 2022-07-05 | 新疆农业大学 | Tissue culture medium and tissue culture method for rapid propagation of grape rootstock stem |
| CN114698549B (en) * | 2022-04-18 | 2023-05-23 | 新疆农业大学 | Tissue culture medium and tissue culture method for rapid propagation of grape stock stem segments |
| CN116548306A (en) * | 2023-05-12 | 2023-08-08 | 山西大学 | Single-bud stem induction callus and tissue culture rapid propagation method for grape at Chardonnay |
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Application publication date: 20110810 |