CN112772414A - Method for improving rapid propagation efficiency of apple rootstock tissue culture seedlings - Google Patents
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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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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
Abstract
The invention discloses a method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings, which comprises the steps of explant inoculation, primary rapid subculture, replying subculture, secondary rapid subculture and time-lapse subculture. By adopting the method, the multiplication coefficient is about 6.11 times and the height is about 5.23cm in 20-25 days during the time-shrinking subculture, the same multiplication coefficient can be kept to be more than 5 generations, the growth speed and the multiplication coefficient of the apple rootstock tissue culture seedling are greatly improved, the time of 1/3-1/2 of the traditional apple rootstock tissue culture is shortened, and the high-quality and high-efficiency production of the apple rootstock tissue culture seedling can be accelerated.
Description
Technical Field
The invention relates to the technical field of plant tissue culture, in particular to a method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings by double fast one recovery.
Background
The apple cultivation mainly exists in the form of grafting scion cultivation nursery stocks on stocks, and the stocks have important influences on growth and development of apples, fruit quality and yield, resistance to various biological and abiotic stresses and the like. Therefore, efficient breeding of apple rootstocks is always a research focus at home and abroad.
The tissue culture technology opens up a new way for the rapid propagation of the apple rootstock. At present, a large number of apple rootstocks and new varieties establish a rapid propagation system through a tissue culture approach. Researches such as Rong (281569) and (281569) find that the proportion of the plant growth regulator suitable for propagation culture of the dwarf rootstock, namely dwarf rootstock No. 6, is 6-BA1.0mg/L and 0.05-0.10 mg/L of NAA, and the propagation coefficient reaches 4.53 times per 4 weeks; sunqingrong and the like take the cold-resistant half-dwarf apple rootstock '54-118' as a test material, and researches show that a suitable proliferation culture medium of the aseptic seedling is QL +6-BA1mg/L + IBA0.3mg/L +30g/L sucrose, and the proliferation coefficient is 4.2 times.
However, the existing apple rootstock tissue culture technology generally has the problems of low efficiency, long culture period and the like, the value-added coefficient of about 4 times cannot meet the rapid propagation expanding requirement of large-scale production, certain manpower and material resources are wasted, and the popularization of the apple rootstock virus-free seedlings is not facilitated.
Disclosure of Invention
Aiming at the prior art, the invention aims to provide a method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings.
In order to achieve the purpose, the invention adopts the following technical scheme:
a method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings by double rapid recovery comprises the following steps:
(1) inoculating the apple rootstock explant into an induction culture medium, and culturing for 20-25 days to form an adventitious bud cluster;
the induction culture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar;
(2) cutting the adventitious bud cluster formed after the culture in the step (1), inoculating the adventitious bud cluster into a primary rapid subculture medium, and continuously subculturing for 3 generations, wherein each generation is cultured for 10-15 days;
the primary rapid subculture medium is an MS culture medium added with 1mg/L of 6-BA, 0.2mg/L of IBA, 30g/L of sucrose and 7.5g/L of agar;
(3) inoculating the tissue culture seedlings subjected to continuous subculture in the step (2) into a recovery subculture medium, and carrying out subculture for 1 generation for 20-25 days;
the recovery subculture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar;
(4) inoculating the tissue culture seedling obtained after 1 generation of culture in the recovery subculture medium in the step (3) into a secondary rapid subculture medium, and continuously subculturing for 3 generations for 10-15 days each generation;
the secondary rapid subculture medium is an MS culture medium added with 1mg/L of 6-BA, 0.2mg/L of IBA, 30g/L of sucrose and 7.5g/L of agar;
(5) inoculating the tissue culture seedling after the secondary quick subculture for 3 generations in the step (4) into a time-shrinking subculture medium, and carrying out subculture for 1-5 generations, wherein each generation is cultured for 20-25 days;
the time-lapse subculture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar.
Preferably, in the step (1), the apple rootstock explant is prepared by the following method:
selecting strong tender branches of apple rootstocks, cutting the tender branches into 2.5-3.5cm stem sections with new shoots, washing with clear water, soaking for 30s with 75% alcohol, sterilizing with 0.05% sodium hypochlorite solution for 10min, and washing with sterile water for 3 times, 30s each time; a beveled incision was made 0.5cm above the bottom of the stem section using a sterilized scalpel.
Preferably, in the step (1), the culture conditions of the apple rootstock explants in the induction medium are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Preferably, in step (2), the conditions for one continuous subculture are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Preferably, in step (3), the conditions for the secondary subculture are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Preferably, in step (4), the conditions for the secondary continuous subculture are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Preferably, in step (5), the time-lapse subculture conditions are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
The invention has the beneficial effects that:
compared with the traditional 2-step subculture technology (namely explant inoculation-subculture, wherein the explant and the subculture medium are MS culture mediums added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar) of the apple rootstock, the propagation coefficient is about 4 times after about 40 days, the method for tissue culture and rapid propagation of the apple rootstock can obviously improve the rapid propagation efficiency of the apple rootstock, and the method for tissue culture and rapid propagation of the apple rootstock can culture the apple rootstock tissue culture seedlings with the height of about 5.23cm and the increment coefficient of about 6.11 times after 20-25 days of the subculture during subsequent contraction, and can keep the same increment coefficient above 5 generations, the method not only greatly improves the growth speed and the increment coefficient of the apple rootstock tissue culture seedling, but also shortens the tissue culture 1/3-1/2 time of the traditional apple rootstock, and is particularly suitable for high-quality and high-efficiency production of the large-scale apple rootstock tissue culture seedling.
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FIG. 1A shows the growth of an apple rootstock 12-2 after two rapid subcultures in the culture medium according to an embodiment of the present invention in a first generation of time-lapse subcultures (one day before the first ten-day subculture in 8 months of 2020);
fig. 1B shows the growth of the apple rootstock 12-2 provided by the embodiment of the present invention after being cultured in the second rapid subculture medium, and then cultured in the time-shortened medium for five generations (one day before 11 months of 2020).
FIG. 1C shows the growth of apple rootstock 12-2 in common subculture medium (MS-based, with other components at 30g/L sucrose, 7.5g/L agar, 6-BA0.6mg/L agar, 0.2mg/L IBA0, pH 5.8, and same set of culture room conditions as A, B) for 20 days (from the middle and last of 5 months in 2020 to the middle and last of 6 months in 2020).
Detailed Description
It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.
As introduced in the background art, the problems of low increment coefficient, long culture period and the like generally exist in the conventional apple rootstock tissue culture technology, certain manpower and material resources are wasted, and the popularization of the apple rootstock virus-free seedlings is not facilitated.
Based on the above, the invention aims to provide a method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings, after the apple rootstock explants are inoculated on an induction culture medium, then inoculating the strain into a high-concentration 6-BA rapid subculture medium for continuous subculture for 3 generations, inoculating the strain into a low-concentration 6-BA recovery subculture medium for recovery culture for 1 generation, then carrying out secondary continuous subculture for 3 generations in the high-concentration 6-BA rapid subculture medium, then inoculating the strain into a lower-concentration 6-BA time-reducing subculture medium, then continuously carrying out multi-generation (more than 5 generations) subculture by using the time-reducing subculture medium, after 20-25 days of time-reducing subculture, can cultivate the apple rootstock tissue culture seedling with the height of about 5.23cm and the increment coefficient of about 6.11 times, and can keep the same increment coefficient to more than 5 generations.
In a specific embodiment of the invention, the method for improving the rapid propagation efficiency of the apple rootstock tissue culture seedling comprises the following steps:
(1) shearing strong twigs from the rootstocks, shearing the twigs into stem segments with young shoots of about 3cm serving as collected strong explants, washing the strong explants under clear water for one hour, soaking the strong explants for 30s on an ultra-clean workbench which is subjected to ultraviolet sterilization for 30min by using 75% alcohol, then disinfecting the strong explants for 10min by using 0.05% sodium hypochlorite solution, and washing the strong shoots for 3 times by using sterile water for 30s each time. A beveled incision was made approximately 0.5cm above the bottom of each stem section using a sterilized scalpel, and the resulting cut was inoculated into an induction medium containing 1L of MS medium, in this order, sucrose at 30g/L, agar at 7.5g/L, 6-BA at 0.6mg/L, IBA 0.2.2 mg/L, and pH at 5.8. Inoculating 5 explants in each bottle of induction culture medium, wherein the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
(2) After 20-25 days, the adventitious bud cluster growing after the culture of the induction culture medium is obliquely cut into about 2cm stem segments on the bottom of each stem segment by using a sterilized debristling knife on an ultra-clean workbench sterilized by ultraviolet for 30min, the stem segments are inoculated on a rapid subculture medium, the rapid subculture medium is based on 1L MS culture medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 1mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8. 5 bud clusters are inoculated in each bottle of induction culture medium, the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
And 3 generations of continuous subculture, wherein the key of the requirement lies in that high-concentration 6-BA is continuously used to make the tissue culture seedling forcibly dedifferentiate capability, maintain the state of plant stem cells, improve the division capability of the tissue culture seedling, and carry out the next generation of rapid subculture after the stem tip of the tissue culture seedling of each generation grows out for 10-15 days on average.
(3) The key point of the requirement is that after 3 successive generations, the stem tip growing out is inoculated into a lower-concentration 6-BA recovery subculture medium for culturing for 1 generation for about 20-25 days, and the slight vitrification problem of the tissue culture seedling appears after 3 successive generations, so that in order to prevent the serious vitrification problem, the tissue culture seedling needs to be cultured for 1 generation in the lower-concentration 6-BA recovery subculture medium. The recovery subculture medium is based on 1L MS medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 0.6mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8.
(4) And (3) inoculating the grown adventitious bud cluster cultured by the recovery subculture medium into the rapid subculture medium again for secondary continuous subculture, wherein the key point of the requirement is to perform secondary dedifferentiation treatment on the tissue culture seedling recovered without vitrification phenomenon to obtain the stem cell subculture tissue culture seedling. And 3 generations of secondary continuous subculture are carried out, each generation of culture is cultured for 10-15 days on average, and the next generation of rapid subculture is carried out after stem tips of tissue culture seedlings of each generation grow. Rapid subculture medium synchronization step (2), namely: based on 1L MS culture medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 1mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8.
(5) The key point of the requirement is that the high propagation potential of the stem cell subculture seedling is exerted under the condition of low concentration 6-BA at the moment, so that the stem cell subculture seedling achieves the effects of high growth speed and high propagation coefficient. The time-lapse subculture medium is based on 1L MS medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 0.6mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8.
When the number of the generations is more than 5, the time-shrinking successive transfer culture medium keeps the effect of rapid successive transfer during shrinking, one generation of culture can be completed in 20-25 days, each adventitious bud cluster can grow to about 5cm on average, the increment coefficient reaches about 6 times, multiple successive transfer treatments on the time-shrinking successive transfer culture medium can be carried out according to the test requirements or the number requirements of seedlings, and the number of the apple rootstock tissue culture seedlings is increased.
Generally speaking, as the number of subcultures increases, the physiological diseases of the apple rootstock tissue culture seedlings increase, and the increment coefficient becomes lower. The increment coefficient of the currently reported apple rootstock tissue culture seedling is about 4 times, but the increment coefficient of about 4 times cannot meet the rapid propagation requirement of large-scale production. Based on the above, the invention originally creates a '5-step' tissue culture technology (namely explant inoculation-primary rapid subculture-recovery subculture-secondary rapid subculture-time shrinking culture), and in the '5-step' tissue culture technology, all the steps supplement each other and are an organic whole; wherein:
and (3) through induction culture, the inoculated explants rapidly grow adventitious bud clusters.
Through one-time rapid subculture, the tissue culture seedling is forced to have dedifferentiation capacity by using high-concentration 6-BA, the state of the plant stem cell is maintained, and the division capacity of the tissue culture seedling is improved; the generation of one-time rapid subculture is more critical, and if the culture generation is too small, the improvement degree of the division capability of the tissue culture seedling is limited; if the culture generation is too many, the tissue culture seedling is easy to have vitrification phenomenon; multiple experiments show that the rapid subculture is suitable for 3 generations.
And correcting the tissue culture seedlings subjected to the primary rapid subculture by the recovery subculture so as to prevent serious vitrification.
Then, inoculating the grown adventitious bud cluster cultured by the recovery subculture medium into the rapid subculture medium again for secondary continuous subculture; the tissue culture seedling which is recovered to have no vitrification phenomenon can be subjected to secondary dedifferentiation treatment through secondary continuous subculture, and the stem cell subculture tissue culture seedling with high propagation potential is obtained.
And finally, inoculating the tissue culture seedling after the secondary continuous subculture into a time-shrinking subculture medium with lower concentration of 6-BA, and exerting the high propagation expansion potential of the stem cell subculture tissue culture seedling under the condition of lower concentration of 6-BA, so that the effects of high growth speed and large multiplication coefficient are achieved.
By the '5-step' tissue culture technology, rapid and continuous subculture can be realized on a time-lapse subculture medium, the increment coefficient of each subculture can be kept about 6 times, the problem that the increment coefficient of the apple rootstock tissue culture seedling is reduced along with the increase of the subculture times is solved, and the rapid propagation efficiency of the apple rootstock tissue culture seedling is greatly improved.
In order to make the technical solutions of the present application more clearly understood by those skilled in the art, the technical solutions of the present application will be described in detail below with reference to specific embodiments.
The test materials used in the examples of the present invention, which were not specifically described, were all those conventional in the art and commercially available.
Example 1: method for improving rapid propagation efficiency of apple rootstock tissue culture seedlings
Firstly, promoting budding:
in 5 months in 2020, selecting strong twigs of apple rootstocks 12-2 self-bred by an inventor through an in-situ breeding technology, cutting the twigs into stem sections with young shoots of about 3cm, washing the stem sections with clear water for one hour, soaking the stem sections with 75% alcohol on an ultra-clean workbench which is subjected to ultraviolet sterilization for 30min, disinfecting the stem sections with 0.05% sodium hypochlorite solution for 10min, and washing the stem sections with sterile water for 3 times for 30s each time. Using a sterilized scalpel, a bevel cut was made at the top of about 0.5cm from the bottom of each stem section and inoculated into an induction medium containing, in order, sucrose at 30g/L, agar at 7.5g/L, 6-BA at 0.6mg/L, IBA 0.2.2 mg/L, and pH at 5.8, based on 1L of MS medium. Inoculating 5 explants in each bottle of induction culture medium, wherein the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Secondly, continuous subculture for one time:
in the middle and last ten days of 5 months in 2020, beveling 12-2 adventitious bud clusters cultured by an induction culture medium into stem segments of about 2cm, inoculating the stem segments to a rapid subculture medium for first rapid subculture for 3 generations, and respectively carrying out second and third rapid subculture on 12-2 stem tips growing on the rapid subculture medium in the early 6 months and the middle and upper 6 days of 6 months in 2020. The culture medium is based on 1L MS culture medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 1.0mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8. 5 bud clusters are inoculated in each bottle of induction culture medium, the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Thirdly, replying for subculture:
in the middle and last ten days of 6 months in 2020, 12-2 stem tips subjected to the third rapid subculture are inoculated into a recovery subculture medium, the medium is based on 1L of MS medium, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 0.6mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8. 5 bud clusters are inoculated in each bottle of induction culture medium, the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
Fourthly, secondary continuous subculture:
in the first ten days of 7 months in 2020, inoculating 12-2 adventitious bud clumps subjected to the reversion subculture into a rapid subculture medium, carrying out secondary continuous subculture for 3 generations, respectively carrying out secondary and third rapid subculture in the middle and last days of 7 months and in the beginning of 8 months in 2020, continuously subculturing in a tissue culture chamber under the same condition at the same species concentration in the medium.
Fifthly, time-lapse rapid propagation:
in the middle and last ten days of 7 months in 2020, 12-2 stem tips subjected to secondary continuous subculture are inoculated into a time-lapse subculture medium, the medium is based on MS, the concentrations of other components are 30g/L of sucrose, 7.5g/L of agar and 0.6mg/L, IBA 0.2.2 mg/L of 6-BA in sequence, and the pH value is 5.8. 5 bud clusters are inoculated in each bottle of induction culture medium, the temperature of a tissue culture room is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
In the middle-upper 8, 9 and 10 months of 2020, subculture was continued on the medium with reduced growth (5 generations, 20-25 days per generation), each generation was as long as about 5.23cm, and the increment coefficients were all 6.11 times (Table 1).
Table 1: time-lapse rapid propagation culture result
Sixthly, subsequent culture
The obtained tissue culture seedling can be inoculated into a rooting culture medium for rooting culture; the rooting culture medium is 1/2MS culture medium added with 0.2 mg/L6-BA, 1.0mg/L IBA, 20g/L sucrose and 7.5g/L agar; and after rooting, carrying out subsequent tests such as plug transplantation and the like.
The above description is only a preferred embodiment of the present application and is not intended to limit the present application, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present application shall be included in the protection scope of the present application.
Claims (7)
1. A method for improving the rapid propagation efficiency of apple rootstock tissue culture seedlings by double rapid recovery is characterized by comprising the following steps:
(1) inoculating the apple rootstock explant into an induction culture medium, and culturing for 20-25 days to form an adventitious bud cluster;
the induction culture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar;
(2) cutting the adventitious bud cluster formed after the culture in the step (1), inoculating the adventitious bud cluster into a primary rapid subculture medium, and continuously subculturing for 3 generations, wherein each generation is cultured for 10-15 days;
the primary rapid subculture medium is an MS culture medium added with 1mg/L of 6-BA, 0.2mg/L of IBA, 30g/L of sucrose and 7.5g/L of agar;
(3) inoculating the tissue culture seedlings subjected to continuous subculture in the step (2) into a recovery subculture medium, and carrying out subculture for 1 generation for 20-25 days;
the recovery subculture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar;
(4) inoculating the tissue culture seedling obtained after 1 generation of culture in the recovery subculture medium in the step (3) into a secondary rapid subculture medium, and continuously subculturing for 3 generations for 10-15 days each generation;
the secondary rapid subculture medium is an MS culture medium added with 1mg/L of 6-BA, 0.2mg/L of IBA, 30g/L of sucrose and 7.5g/L of agar;
(5) inoculating the tissue culture seedling after the secondary quick subculture for 3 generations in the step (4) into a time-shrinking subculture medium, and carrying out subculture for 1-5 generations, wherein each generation is cultured for 20-25 days;
the time-lapse subculture medium is an MS culture medium added with 0.6 mg/L6-BA, 0.2mg/L IBA, 30g/L sucrose and 7.5g/L agar.
2. The method according to claim 1, wherein in step (1), the apple rootstock explant is prepared by the following method:
selecting strong tender branches of apple rootstocks, cutting the tender branches into 2.5-3.5cm stem sections with new shoots, washing with clear water, soaking for 30s with 75% alcohol, sterilizing with 0.05% sodium hypochlorite solution for 10min, and washing with sterile water for 3 times, 30s each time; a beveled incision was made 0.5cm above the bottom of the stem section using a sterilized scalpel.
3. The method according to claim 1, wherein in step (1), the culture conditions of the apple rootstock explants in the induction medium are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
4. The method according to claim 1, wherein in the step (2), the conditions of the successive subcultures are: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
5. The method according to claim 1, wherein in the step (3), the conditions for the recovery culture are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
6. The method according to claim 1, wherein in the step (4), the conditions of the secondary continuous subculture are as follows: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
7. The method according to claim 1, wherein in step (5), the time-lapse subculture conditions are: the temperature is 25 +/-2 ℃, the illumination time is 10h/d, and the illumination intensity is 1000 lx.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101965797A (en) * | 2010-10-25 | 2011-02-09 | 江苏省农业科学院 | Quick propagation seedling-breeding method for reducing vitrifaction in multiple times of subculture of rosaceous plant |
| CN106134997A (en) * | 2016-07-06 | 2016-11-23 | 中国农业科学院郑州果树研究所 | The group training fast seedling-cultivating method of apple rootstock M26 |
| CN106234212A (en) * | 2015-12-07 | 2016-12-21 | 内蒙古自治区农牧业科学研究院 | A kind of method for tissue culture controlling Fructus Fragariae Ananssae Vitrification |
| CN106613934A (en) * | 2015-10-28 | 2017-05-10 | 北京市农林科学院 | Method for rapidly propagating apple rootstock SH6 |
| CN108401907A (en) * | 2018-04-25 | 2018-08-17 | 河南省农业科学院 | A kind of Apples Dwarf Stocks M9T337 tissue cultures volume production method |
-
2021
- 2021-01-08 CN CN202110022203.0A patent/CN112772414B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101965797A (en) * | 2010-10-25 | 2011-02-09 | 江苏省农业科学院 | Quick propagation seedling-breeding method for reducing vitrifaction in multiple times of subculture of rosaceous plant |
| CN106613934A (en) * | 2015-10-28 | 2017-05-10 | 北京市农林科学院 | Method for rapidly propagating apple rootstock SH6 |
| CN106234212A (en) * | 2015-12-07 | 2016-12-21 | 内蒙古自治区农牧业科学研究院 | A kind of method for tissue culture controlling Fructus Fragariae Ananssae Vitrification |
| CN106134997A (en) * | 2016-07-06 | 2016-11-23 | 中国农业科学院郑州果树研究所 | The group training fast seedling-cultivating method of apple rootstock M26 |
| CN108401907A (en) * | 2018-04-25 | 2018-08-17 | 河南省农业科学院 | A kind of Apples Dwarf Stocks M9T337 tissue cultures volume production method |
Non-Patent Citations (3)
| Title |
|---|
| JAIME A.TEIXEIRA DA SILVA等: "In vitro tissue culture of apple and other Malus species: recent advances and applications", 《PLANTA》 * |
| 李淑艳等: "不同继代培养次数对苹果八棱海棠离体培养和遗传转化的影响", 《果树学报》 * |
| 赵亮明等: "苹果砧木组织培养与快繁技术研究", 《西北农业学报》 * |
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