CN102577981A - Method for strengthening and rooting tissue culture seedlings of transgenic peanuts - Google Patents
Method for strengthening and rooting tissue culture seedlings of transgenic peanuts Download PDFInfo
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- CN102577981A CN102577981A CN2012100794204A CN201210079420A CN102577981A CN 102577981 A CN102577981 A CN 102577981A CN 2012100794204 A CN2012100794204 A CN 2012100794204A CN 201210079420 A CN201210079420 A CN 201210079420A CN 102577981 A CN102577981 A CN 102577981A
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Abstract
The invention provides a method for strengthening and rooting tissue culture seedlings of transgenic peanuts. The method has a good seedling strengthening effect and is high in rooting rate and rooting quality. The method comprises the following steps of: (1) transferring peanut tissue culture seedlings on which cluster buds are induced and differentiated into a seedling strengthening culture medium A, culturing for 20 days, and replacing the culture medium once every 10 days, wherein a component of the seedling strengthening culture medium A is a Murashige and Skoog (MS) culture medium, and used hormone is cytokinin 6-benzylaminopurine (6-BA); (2) dividing the tissue culture seedlings with the heights of more than 1.5cm into single plants, transferring into a seedling strengthening culture medium B, culturing for 20 to 25 days, and replacing the culture medium once every 10 days, wherein a component of the seedling strengthening culture medium B is the MS culture medium, and used hormone is cytokinin 6-BA at the concentration of 0.4mg/L and auxin naphthyl acetic acid (NAA) at the concentration of 0.1mg/L; and (3) transferring into a rooting culture medium when the seedlings with the heights of 2.5 to 4cm, and rooting, wherein a component of the rooting culture medium is a one/second MS culture medium, and used hormone is auxin indolebutyric acid (IBA) at the concentration of 3mg/L and auxin NAA at the concentration of 0.3mg/L.
Description
Technical field
The present invention relates to the training seedling-growing method of cultivating peanut, especially a kind of transgenosis peanut tissue cultivating seedling strong sprout, the method for taking root.
Background technology
Utilize method for tissue culture to carry out peanut and expand numerous following advantage that has: at first, but the short time breed in a large number, to breeding rare kind and present peanut transgenosis work has crucial meaning; Secondly, it is little to take up room, and does not receive area, season limit; In addition, condition of culture is artificially controlled, and the condition homogeneous is convenient to stably carry out cultivating and producing.
The link of taking root is key one ring of whole peanut group training process, has accepted inducing of early stage and has broken up the link and the transplanting link in later stage.The quality that tissue cultivating seedling is taken root has directly determined the survival rate that the later stage transplants.And the quality of taking root directly receives to break up early stage the influence of seedling quality, if seedling is short and small, thin and delicate, the quality of taking root is also poor.Therefore must be through strong sprout.The raising mutual restriction of strong sprout and growth coefficient, if the simple growth coefficient of pursuing, the differentiation seedling certainly will be short and small, thin and delicate, if the simple quality of pursuing seedling influences growth coefficient again, the differentiation seedling reduced.Must be through selecting the suitable basic element of cell division and the kind and the variable concentrations proportioning of growth hormone, to realize the dual purpose in propagation and strong sprout.At present with regard to peanut group training, also there are not a cover effective strong sprout, rooting method, existing method can't realize breeding the purpose with strong sprout synchronously, and rooting rate is low, and it is of poor quality to take root, and has a strong impact on the survival rate that the later stage transplants.Therefore, setting up one, to overlap strong sprout, the system of taking root efficiently be to improve the key of peanut tissue culture efficient under the present condition.
Summary of the invention
The invention provides a kind of strong sprout effective, rooting rate is high, the measured transgenosis peanut of the matter of taking root tissue cultivating seedling strong sprout, the method for taking root.
Realize the transgenosis peanut tissue cultivating seedling strong sprout of the object of the invention, the method for taking root, comprise the steps:
(1) will induce the peanut tissue cultivating seedling that differentiates the bud of growing thickly to transfer to last 20 day of strong seedling culture base A, change a subculture in 10 days, the component of said strong seedling culture base A is the MS medium, and used hormone is basic element of cell division 6-BA;
(2) will grow to the above tissue cultivating seedling of 1.5cm and be divided into individual plant; Moved to strong seedling culture base B last 20 ~ 25 days; Changed a subculture in 10 days, the composition of said strong seedling culture base B is the MS medium, and used hormone is the basic element of cell division 6-BA of 0.4 mg/L and the growth hormone NAA of 0.1mg/L;
(3) treat that seedling grows to 2.5 ~ 4cm, move to root media and take root that the composition of said root media is 1/2 MS medium, used hormone is growth hormone IBA and NAA, and concentration is respectively 3mg/L and 0.3mg/L.
The present invention further provides the optimal technical scheme of above-mentioned peanut tissue cultivating seedling strong sprout, rooting method:
As preferably, the working concentration of the culture medium A in the said step (1) is reduced to 1mg/L from the 3mg/L of recovery media and inducing culture, and medium thickness is 1.5cm.
As preferably; Said will induce the peanut tissue cultivating seedling that differentiates the bud of growing thickly to transfer to strong seedling culture base A to go up the time; To grow preferably, bud is divided into individual plant with the aseptic operation cutter; And excision bottom contact the pitchy part that forms with medium, the medium 2-3mm cultivation of submerging of bottom stem end, with some as yet not the bud of moulding be divided into several bud clumps cultivations.
As preferably, the thickness of the medium B in the said step (2) is 1.5cm.
As preferably, said choosing grown to the above tissue cultivating seedling individual plant of 1.5cm and moved on the strong seedling culture base B, and the excision bottom contact pitchy part of formation with medium, the medium 2-3mm cultivation of submerging of bottom stem end.
As preferably, the thickness of the root media in the said step (3) is 2 ~ 2.5cm.
As preferably, said will the process grown to the high tissue cultivating seedling of 2.5 ~ 4cm strong sprout and moved on the root media 15 ~ 30 days, waits to bear the root more than 3, the long 2cm that surpasses of root, and succulent stem surpasses two internodes, can refine seedling, transplanting.
As preferably; After the culture of rootage to 15 of said step (3) day, if the situation of taking root does not still reach requirement, the 0.6 ~ 1cm that need fall again on the former medium upper strata root medium of improving people's living condition; Newly add medium and need use water-bath cold to 34 ~ 36 ℃, tissue culture bottle is tilted slowly pour into along a bottle wall.
As preferably, said step (1) and the used intensity of illumination of step (2) are 4000Lx, and the stage intensity of illumination of taking root is adjusted to 5000Lx.
The beneficial effect of transgenosis peanut tissue cultivating seedling of the present invention strong sprout, the method for taking root is following:
Peanut tissue cultivating seedling of the present invention strong sprout, the method for taking root, the peanut tissue cultivating seedling growing way of acquisition is vigorous, and rooting rate reaches 100%, and adventive root is sturdy, the capillary root flourishing, has effectively improved transplanting survival rate.Solved peanut tissue cultivating seedling take root difficulty, the poor problem of taking root, social benefit and scientific research are worth significantly.
Embodiment
Embodiment 1
To educate 26 buds of growing thickly through the 51 strain peanut flowers of inducing differentiation transfers to strong seedling culture base A and go up to cultivate 20 days; During transfer; To grow preferably, bud is divided into individual plant; And contact the pitchy part that forms with medium with aseptic operation cutter excision bottom, the medium 2 ~ 3mm cultivation of submerging of bottom stem end, and with some as yet not the bud of moulding be divided into several bud clumps cultivations.The component of strong seedling culture base A is to add 1mg/L basic element of cell division 6-BA (concentration is 3mg/L in recovery, the inducing culture), PH=5.8 in the MS medium.Changed a subculture in 10 days, growth is divided into the individual plant cultivation preferably once more in the bud of will growing thickly.After strong seedling culture base A cultivation; To grow to the above transgenic seedling individual plant of 1.5cm and move to last 25 day of strong seedling culture base B; Same excision bottom contact the pitchy part that forms with medium, the medium 2 ~ 3mm cultivation of submerging of bottom stem end, 10 days replacing one subcultures.The composition of strong seedling culture base B is to add 0.4 mg/L basic element of cell division 6-BA, 0.1mg/L growth hormone NAA, PH=5.8 in the MS medium.Treat that seedling grows to 2.5 ~ 4cm, move to root media and take root.The composition of root media is the 1/2MS medium, and PH=5.8, used hormone are growth hormone IBA and NAA, and concentration is respectively 3mg/L and 0.3mg/L.
Through strong sprout, 249 strain tissue cultivating seedling get into takes root the stage, and rooting rate is 100%, and to moving to root media 25 days; 99% tissue cultivating seedling all bears the root more than 3, and root is long to surpass 2cm, and adventive root is more sturdy; The capillary root is flourishing, reaches the transplanting requirement, and transplanting survival rate reaches more than 92%.
And common tissue cultivating seedling strong sprout, the method for taking root have only 46% tissue cultivating seedling to bear the root more than 3, and root is long generally between 1 ~ 1.5 centimetre.Adventive root a little less than, the capillary root is undeveloped, in time can transplant, transplanting survival rate also is lower than 50%.
Embodiment 2
To educate 17 buds of growing thickly through the 47 strain peanut flowers of inducing differentiation transfers to strong seedling culture base A and go up to cultivate 20 days; During transfer; To grow preferably, bud is divided into individual plant; And contact the pitchy part that forms with medium with aseptic operation cutter excision bottom, the medium 2 ~ 3mm cultivation of submerging of bottom stem end, and with some as yet not the bud of moulding be divided into several bud clumps cultivations.The component of strong seedling culture base A is to add 1mg/L basic element of cell division 6-BA (concentration is 3mg/L in recovery, the inducing culture), PH=5.8 in the MS medium.Changed a subculture in 10 days, growth is divided into the individual plant cultivation preferably once more in the bud of will growing thickly.After strong seedling culture base A cultivation; To grow to the above transgenic seedling individual plant of 1.5cm and move to last 20 day of strong seedling culture base B; Same excision bottom contact the pitchy part that forms with medium, the medium 2 ~ 3mm cultivation of submerging of bottom stem end, 10 days replacing one subcultures.The composition of strong seedling culture base B is to add 0.4 mg/L basic element of cell division 6-BA, 0.1mg/L growth hormone NAA, PH=5.8 in the MS medium.Treat that seedling grows to 2.5 ~ 4cm, move to root media and take root.The composition of root media is the 1/2MS medium, and PH=5.8, used hormone are growth hormone IBA and NAA, and concentration is respectively 3mg/L and 0.3mg/L.
Through strong sprout, 225 strain tissue cultivating seedling get into takes root the stage, and rooting rate is 100%, and bears the root more than 3 to moving to 20 days 98% tissue cultivating seedling of root media, and root is long to surpass 2cm, and adventive root is more sturdy, and the capillary root is flourishing, reaches the transplanting requirement.
And common tissue cultivating seedling strong sprout, the method for taking root have only 46% tissue cultivating seedling to bear the root more than 3, and root is long generally between 1 ~ 1.5 centimetre.Adventive root a little less than, the capillary root is undeveloped, in time can transplant, transplanting survival rate also is lower than 50%.
Embodiment recited above describes preferred implementation of the present invention; Be not that scope of the present invention is limited; Design under the spiritual prerequisite not breaking away from the present invention; Various distortion and improvement that the common engineers and technicians in this area make technical scheme of the present invention all should fall in the definite protection domain of claims of the present invention.
Claims (9)
1. transgenosis peanut tissue cultivating seedling strong sprout, the method for taking root comprise the steps:
(1) will induce the peanut tissue cultivating seedling that differentiates the bud of growing thickly to transfer to last 20 day of strong seedling culture base A, change a subculture in 10 days, the component of said strong seedling culture base A is the MS medium, and used hormone is basic element of cell division 6-BA;
(2) will grow to the above tissue cultivating seedling of 1.5cm and be divided into individual plant; Moving to strong seedling culture base B went up 20-25 days; Changed a subculture in 10 days, the composition of said strong seedling culture base B is the MS medium, and used hormone is the basic element of cell division 6-BA of 0.4 mg/L and the growth hormone NAA of 0.1mg/L;
(3) treat that seedling grows to 2.5 ~ 4cm, move to root media and take root that the composition of said root media is 1/2 MS medium, used hormone is growth hormone IBA and NAA, and concentration is respectively 3mg/L and 0.3mg/L.
2. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root, it is characterized in that: the culture medium A thickness in the said step (1) is 1.5cm.
3. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root; It is characterized in that: said will induce the peanut tissue cultivating seedling that differentiates the bud of growing thickly to transfer to strong seedling culture base A to go up the time; To grow preferably, bud is divided into individual plant with the aseptic operation cutter; And excision bottom contact the pitchy part that forms with medium, the medium 2 ~ 3mm cultivation of submerging of bottom stem end, with some as yet not the bud of moulding be divided into several bud clumps cultivations.
4. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root, it is characterized in that: the thickness of the medium B in the said step (2) is 1.5cm.
5. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root; It is characterized in that: the said tissue cultivating seedling individual plant that grows to more than the 1.5cm of choosing moves on the strong seedling culture base B; Excision bottom contact the pitchy part of formation with medium, the medium 2 ~ 3mm cultivation of submerging of bottom stem end.
6. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root, it is characterized in that: the thickness of the root media in the said step (3) is 2 ~ 2.5cm.
7. transgenosis peanut tissue cultivating seedling according to claim 1 strong sprout, the method for taking root; It is characterized in that: said will the process grown to the high tissue cultivating seedling of 2.5 ~ 4cm strong sprout and moved on the root media 15 ~ 30 days; Wait to bear the root more than 3; The long 2cm that surpasses of root, succulent stem surpasses two internodes, can refine seedling, transplanting.
8. transgenosis peanut tissue cultivating seedling according to claim 7 strong sprout, the method for taking root; It is characterized in that: after the culture of rootage to 15 of said step (3) day; If the situation of taking root does not still reach requirement; 0.6 ~ the 1cm that need be on former medium upper strata falls the again root medium of improving people's living condition newly adds medium and need use water-bath cold to 34 ~ 36 ℃, tissue culture bottle is tilted slowly pour into along a bottle wall.
9. according to the arbitrary described transgenosis peanut tissue cultivating seedling of claim 1 ~ 8 strong sprout, the method for taking root, it is characterized in that: said step (1) and the used intensity of illumination of step (2) are 4000Lx, and the stage intensity of illumination of taking root is adjusted to 5000Lx.
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Cited By (3)
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| CN103651132A (en) * | 2013-12-06 | 2014-03-26 | 山东省农业科学院生物技术研究中心 | Rapid rooting medium for peanut tissue culture seedlings and culture method |
| CN106386493A (en) * | 2016-09-28 | 2017-02-15 | 广西大学 | Immature-embryo culture method for red-peel peanuts |
| CN110786245A (en) * | 2019-12-16 | 2020-02-14 | 山东省花生研究所 | Peanut tissue culture seedling root induction culture medium and preparation method and application thereof |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103651132A (en) * | 2013-12-06 | 2014-03-26 | 山东省农业科学院生物技术研究中心 | Rapid rooting medium for peanut tissue culture seedlings and culture method |
| CN103651132B (en) * | 2013-12-06 | 2015-06-10 | 山东省农业科学院生物技术研究中心 | Rapid rooting medium for peanut tissue culture seedlings and culture method |
| CN106386493A (en) * | 2016-09-28 | 2017-02-15 | 广西大学 | Immature-embryo culture method for red-peel peanuts |
| CN110786245A (en) * | 2019-12-16 | 2020-02-14 | 山东省花生研究所 | Peanut tissue culture seedling root induction culture medium and preparation method and application thereof |
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Address after: 266607, Shandong Province, Qingdao City, Laixi Province town shop No. 23 Hing Road Applicant after: Shandong Peanut Inst. Address before: 266100, No. 64, Wangcheng Road, Laixi, Shandong, Qingdao Applicant before: Shandong Peanut Inst. |
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