CN107549016A - The rooting method and rooting induction culture medium of a kind of soybean transgene tissue-cultured seedling - Google Patents
The rooting method and rooting induction culture medium of a kind of soybean transgene tissue-cultured seedling Download PDFInfo
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- CN107549016A CN107549016A CN201710966331.4A CN201710966331A CN107549016A CN 107549016 A CN107549016 A CN 107549016A CN 201710966331 A CN201710966331 A CN 201710966331A CN 107549016 A CN107549016 A CN 107549016A
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Abstract
The invention belongs to plant biotechnology field, more particularly to the rooting method and rooting induction culture medium of a kind of soybean transgene tissue-cultured seedling, rooting induction medium component includes the 2.5g/L of MS powder 2.0, the 0.4mg/L of methyl α-naphthyl acetate 0.2, the 15g/L of sucrose 10, the 8g/L of agar 6, and pH value is 5.6 5.8.Rooting method is to scale off soybean transgene tissue culture elongation seedling using scissors straight snips along basal part of stem, is inoculated on rooting induction culture medium and takes root 34 days, be subsequently transferred in vermiculite soil, capping moisturizing is placed in light culture in greenhouse and stayed overnight.Next day opens lid, and illumination cultivation is viable in greenhouse.The invention enables the survival rate of soybean transgene tissue-cultured seedling to 100%; greatly reduce soybean tissue culture seedling rooting difficulty; rootage duration is grown; difficulty survives, is unable to the problems such as scale is taken root; technology platform and support are provided for soybean biological technology Breeding, present invention could apply to vegetative propagation of soybean transformants plant cultivating and soybean transformants plant etc. is obtained by transgenic technology.
Description
Technical field
The invention belongs to plant biotechnology field, and in particular to a kind of rooting method of soybean transgene tissue-cultured seedling and life
Root induction culture medium.
Background technology
Soybean (Glycine Max (L.) Merr.) is a kind of important oil plant and forage crop, has important economic valency
Value.The soybean breeder carried out using transgenic technology is one of current study hotspot.Compared with other crops, soybean is compared
Hardly possible conversion, is mainly shown as that transformation of soybean efficiency is low, and regeneration frequency is low after conversion, the problems such as difficulty of taking root.Difficulty of wherein taking root is big
Most important link in the regenerative process of beans plant.Nutrient is independently only absorbed by regenerated root, soybean sprout could be caused to be good for
Health is grown.According to the report published an article, taking root for soybean general uses this common three kinds of IAA, IBA or NAA
The induction of hormone.Wherein, when hormone IAA is mainly used in soybean transplantation of seedlings earliest, root is smeared stimulation and taken root；Hereafter, 2008,
The scholar Zhao Qing of University Of Tianjin is taken root using 0.2mg/L IBA inducing soybeans, and its efficiency of taking root is up to 85%；It is 2013, interior
The scholar Hu Weijing of Mongolian university is taken root soon using 0.5mg/L NAA induction discoveries, but root is thicker, without root hair；2014,
The scholar Jiang Guoyong of Qingdao Agricultural University is induced 1-3 days using 0.5mg/L IBA, is subsequently transferred to cultivate 14 in root media
It can grow up to 2-5 bar roots, and rooting rate reaches 100%.Therefore according to existing soybean rooting technique handbook, although can be one
Taken root under fixed condition, but can not realize that soybean scale is taken root, turn into the bottleneck of soybean large-scale molecular breeding, it would be highly desirable to solve.
The content of the invention
In order to solve the above technical problems, the invention provides a kind of rooting method of soybean transgene tissue-cultured seedling, solve
In the prior art soybean take root it is difficult, take root it is slow, the problem of scale is taken root can not be realized, filled up technological gap, will utilize should
Method implements extensive soybean molecular breeding, and important technology platform and support are provided for transgenic research.
The present invention is achieved in that according to an aspect of the present invention, there is provided a kind of inducing culture, composition include MS
Powder 2.0-2.5g/L, methyl α-naphthyl acetate 0.2-0.4mg/L, sucrose 10-15g/L, agar 6-8g/L, pH value 5.6-5.8.
Further, the Fiber differentiation based component includes MS powder 2.2g/L, methyl α-naphthyl acetate 0.3mg/L, sucrose 15g/L, fine jade
Fat 6.5g/L, pH value 5.6-5.8.
According to another aspect of the present invention, there is provided a kind of rooting method of soybean transgene tissue-cultured seedling, including it is as follows
Step：
1) according to above-mentioned recipe configuration rooting induction culture medium；
2) pretreatment and inoculation of soybean transgene tissue-cultured seedling：By length it is 3-4cm's with tweezers on superclean bench
Soybean transgene tissue-cultured seedling is cut with scissors along basal part of stem straight snips, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration take root
In inducing culture；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse,
Hot-house culture temperature is 24-26 DEG C, humidity 50-70%, intensity of illumination 1500-2000lux, light application time 16h, culture
The soybean tissue culture seedlings with root are obtained after 3-4 days；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in temperature
Light culture is stayed overnight in room, and next day opens lid, and illumination cultivation is viable in greenhouse.
Further, soybean transgene tissue-cultured seedling is cut with scissors along basal part of stem straight snips, and by otch insert described in take root and lure
Lead below culture medium liquid level at 0.3-0.5cm.
Further, the soybean transgene tissue-cultured seedling being inserted into the rooting induction culture medium cultivates 3-4 in greenhouse
My god, the root that length is 3-5mm length can be obtained.
Compared with prior art, the advantage of the invention is that：The time required to the rooting method short (3-4 days), and it can obtain
Obtain 100% take root efficiency and 100% survival rate.Compared with existing rooting technique, the rooting technique system of the invention established
Rootage duration can greatly be reduced, soybean scale is taken root, for soybean transgene research provides important technology platform with
Support.
Brief description of the drawings
Below in conjunction with the accompanying drawings and embodiment the present invention is further detailed explanation：
Fig. 1 is the rooted seedling of the method for soybean (kind JACK) the application present invention；
Fig. 2 is that the bottom of the method for soybean (kind JACK) the application present invention is taken root figure；
Fig. 3 is the rooted seedling of the method for soybean (kind WILLIAMS82) the application present invention；
Fig. 4 is that the bottom of the method for soybean (kind WILLIAMS82) the application present invention is taken root figure.
Embodiment
In order to make the purpose , technical scheme and advantage of the present invention be clearer, below in conjunction with the accompanying drawings and embodiment, it is right
The present invention is further elaborated.It should be appreciated that specific embodiment described herein is used only for explaining the present invention, and
It is not used in the restriction present invention.
In order to solve, soybean transgene tissue culture seedling rooting of the prior art is difficult, rootage duration is long, can not take root on a large scale
The problem of, the invention provides a kind of soybean transgene tissue culture seedling-growing method, is set out using soybean transgene tissue culture elongation seedling as research
Point, quantity is induced in 3-4d, length is 3-5mm root, and inductivity is 100%；That cultivates in this way is big
Beans transgenosis tissue-cultured seedling, which can be transferred directly in vermiculite, to be grown, survival rate to 100%.Material therefor of the present invention, medicine
Product are commercially available.
Embodiment 1
With soybean (kind JACK) for subjects
1) configuration of rooting induction culture medium：Including MS powder 2.0g/L, methyl α-naphthyl acetate 0.2mg/L, sucrose 10g/L, agar 6g/
L, pH value 5.6-5.8；
2) on superclean bench, with tweezers by the soybean transgene tissue-cultured seedling scissors that length is 3-4cm along basal part of stem
Straight snips is cut, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration rooting induction culture medium liquid level below at 0.3-0.5cm；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse,
Hot-house culture temperature is 24 DEG C, humidity 50%, intensity of illumination 1500lux, light application time 16h, and culture obtains after 3-4 days
Strip length is the soybean tissue culture seedlings of 3-5mm roots；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in temperature
Light culture is stayed overnight in room, and next day opens lid, and illumination cultivation is viable in greenhouse.
Observation is learnt after one week, using the soybean survival rate that this method is taken root to 100%, with reference to figure 1 and Fig. 2.
Embodiment 2
With soybean (kind WILLIAMS82) for subjects
1) configuration of rooting induction culture medium：Including MS powder 2.2g/L, methyl α-naphthyl acetate 0.3mg/L, sucrose 13g/L, agar 7g/
L, pH value 5.6-5.8；
2) on superclean bench, with tweezers by the soybean transgene tissue-cultured seedling scissors that length is 3-4cm along basal part of stem
Straight snips is cut, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration rooting induction culture medium liquid level below at 0.3-0.5cm；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse,
Hot-house culture temperature is 25 DEG C, humidity 60%, intensity of illumination 1700lux, light application time 16h, and culture obtains after 3-4 days
Strip length is the soybean tissue culture seedlings of 3-5mm roots；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in temperature
Light culture is stayed overnight in room, and next day opens lid, and illumination cultivation is viable in greenhouse.
Observation is learnt after one week, using the soybean survival rate that this method is taken root to 100%, with reference to figure 3 and Fig. 4.
Embodiment 3,
With soybean (kind WILLIAMS82) for subjects
1) configuration of rooting induction culture medium：Including MS powder 2.5g/L, methyl α-naphthyl acetate 0.4mg/L, sucrose 15g/L, agar 8g/
L, pH value 5.6-5.8；
2) on superclean bench, with tweezers by the soybean transgene tissue-cultured seedling scissors that length is 3-4cm along basal part of stem
Straight snips is cut, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration rooting induction culture medium liquid level below at 0.3-0.5cm；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse,
Hot-house culture temperature is 26 DEG C, humidity 70%, intensity of illumination 2000lux, light application time 16h, and culture obtains after 3-4 days
Strip length is the soybean tissue culture seedlings of 3-5mm roots；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in temperature
Light culture is stayed overnight in room, and next day opens lid, and illumination cultivation is viable in greenhouse.
Observation is learnt after one week, utilizes the soybean survival rate that this method is taken root to 100%.
Embodiment 4
With soybean (kind JACK) for subjects
1) configuration of rooting induction culture medium：Including MS powder 2.2g/L, methyl α-naphthyl acetate 0.3mg/L, sucrose 15g/L, agar
6.5g/L, heating are completely dissolved, and regulation Medium's PH Value is 5.8, autoclaving, tissue culture is poured into when temperature is reduced to 60 DEG C
Box, cooling is standby, and aforesaid operations are carried out under aseptic conditions；
2) on superclean bench, with tweezers by the soybean transgene tissue-cultured seedling scissors that length is 3-4cm along basal part of stem
Straight snips is cut, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration rooting induction culture medium liquid level below at 0.3-0.5cm；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse,
Hot-house culture temperature is 26 DEG C, humidity 70%, intensity of illumination 2000lux, light application time 16h, and culture obtains after 3-4 days
The soybean tissue culture seedlings for the root that strip length is 3-5mm, number is indefinite；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in temperature
Light culture is stayed overnight in room, and next day opens lid, and illumination cultivation is viable in greenhouse.
Observation is learnt after one week, utilizes the soybean survival rate that this method is taken root to 100%.
Embodiment 5
The places different from embodiment 4 are, when step 1) configures inducing culture, MS powder is dissolved in 1 liter of distilled water
2g, add methyl α-naphthyl acetate 0.3mg, sucrose 10g, agar 7g, other steps are same as Example 4, after one week observe soybean into
Motility rate is 100%.
Embodiment 6
The places different from embodiment 4 are, when step 1) configures inducing culture, MS powder is dissolved in 1 liter of distilled water
2.5g, methyl α-naphthyl acetate 0.4mg, sucrose 15g, agar 8g are added, other steps are same as Example 4, and soybean is observed after one week
Survival rate is 100%.
Claims (5)
1. a kind of rooting induction culture medium of soybean transgene tissue-cultured seedling, it is characterised in that the Fiber differentiation based component includes
MS powder 2.0-2.5g/L, methyl α-naphthyl acetate 0.2-0.4mg/L, sucrose 10-15g/L, agar 6-8g/L, pH value 5.6-5.8.
2. according to the rooting induction culture medium of the soybean transgene tissue-cultured seedling described in claim 1, it is characterised in that the induction
Medium component includes MS powder 2.2g/L, methyl α-naphthyl acetate 0.3mg/L, sucrose 15g/L, agar 6.5g/L, pH value 5.6-5.8.
3. a kind of rooting method of soybean transgene tissue-cultured seedling, it is characterised in that comprise the following steps：
1) according to any described recipe configuration rooting induction culture medium of claim 1 or 2；
2) pretreatment and inoculation of soybean transgene tissue-cultured seedling：On superclean bench, the soybean for being 3-4cm by length with tweezers
Transgenosis tissue-cultured seedling is cut with scissors along basal part of stem straight snips, the tissue-cultured seedling inserting step 1 that then will be cut) in configuration rooting induction
In culture medium；
3) root induction：The rooting induction culture medium that soybean transgene tissue-cultured seedling has been inoculated with step 2) is placed in greenhouse, greenhouse
Cultivation temperature is 24-26 DEG C, humidity 50-70%, intensity of illumination 1500-2000lux, light application time 16h, cultivates 3-4
The soybean tissue culture seedlings with root are obtained after it；
4) hardening：The soybean tissue culture seedlings of raw good root in step 3) are transferred directly in vermiculite soil, capping moisturizing is placed in greenhouse
Light culture is stayed overnight, and next day opens lid, and illumination cultivation is viable in greenhouse.
4. according to the rooting method of the soybean transgene tissue-cultured seedling described in claim 3, it is characterised in that soybean transgene tissue culture
Seedling is cut with scissors along basal part of stem straight snips, and otch is inserted below the rooting induction culture medium liquid level at 0.3-0.5cm.
5. according to the rooting method of the soybean transgene tissue-cultured seedling described in claim 3, it is characterised in that be inserted into described take root
Soybean transgene tissue-cultured seedling in inducing culture is cultivated 3-4 days in greenhouse, can obtain the root that length is 3-5mm length.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101173297A (en) * | 2007-10-10 | 2008-05-07 | 吉林省农业科学院 | High-efficiency genetic transformation method for soybean immaturity seed lobe regeneration system with auxiliary vacuum permeation |
| CN101176427A (en) * | 2007-12-06 | 2008-05-14 | 中国科学院东北地理与农业生态研究所 | Method for regenerating plant strain using soybean cotyledonary node |
| CN102174562A (en) * | 2011-01-28 | 2011-09-07 | 河北农业大学 | Application of novel rooting method in soybean transgenic technology |
-
2017
- 2017-10-17 CN CN201710966331.4A patent/CN107549016A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101173297A (en) * | 2007-10-10 | 2008-05-07 | 吉林省农业科学院 | High-efficiency genetic transformation method for soybean immaturity seed lobe regeneration system with auxiliary vacuum permeation |
| CN101176427A (en) * | 2007-12-06 | 2008-05-14 | 中国科学院东北地理与农业生态研究所 | Method for regenerating plant strain using soybean cotyledonary node |
| CN102174562A (en) * | 2011-01-28 | 2011-09-07 | 河北农业大学 | Application of novel rooting method in soybean transgenic technology |
Non-Patent Citations (1)
| Title |
|---|
| 鲍顺淑等: "可控环境下大豆组培苗的生根过程", 《中国农业工程学会2005年学术年会论文集》 * |
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Application publication date: 20180109 |