CN101574058B - Barbadosnut plantlet tissue culture rapid propagation and rooting method - Google Patents
Barbadosnut plantlet tissue culture rapid propagation and rooting method Download PDFInfo
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- CN101574058B CN101574058B CN2009100596150A CN200910059615A CN101574058B CN 101574058 B CN101574058 B CN 101574058B CN 2009100596150 A CN2009100596150 A CN 2009100596150A CN 200910059615 A CN200910059615 A CN 200910059615A CN 101574058 B CN101574058 B CN 101574058B
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Abstract
The invention discloses a barbadosnut plantlet tissue culture rapid propagation and rooting method, which comprises the steps as follows: a germ of a barbadosnut seed after disinfection and sterilization is placed in a basic culture medium with the pH value of 5.4 to 6.0, and cultured to germinate into a sterile plantlet after 5 days at the culture temperature of 25 to 35 DEG C, with the illumination intensity of 1500 to 3000lx and the illumination time of 12h/d; stem segments with single bud of the sterile plantlet are placed in the rapid propagation culture medium, and cultured for 7days so as to induce out cluster buds; and the cluster buds growing to 2 to 3cm are cut and inoculate to a rooting culture medium, start to root after being cultured for another 5 days, and then grow into 2 to 3 cm strong stocks after 15 days, thus being capable of being transplanted. Only 66 days are needed for the rooting of the first propagation plantlet from the seed germination, therefore, the propagation is rapid; the hormone concentration of the culture process is low, the contact time is short, and the variation probability is low; 4.25 plantets can be obtained by utilizing one seed for 3 weeks, therefore, the propagation coefficient is high; and the types of used agents are single, the usage amount is small, the price is cheap and the cost is low.
Description
Technical field
The invention belongs to Jatropha curcas Plant Tissue Breeding quick propagating technology field, the method that is specifically related to a kind of barbadosnut plantlet tissue culture rapid propagation and takes root.
Technical background
Jatropha curcas is an Euphorbiaceae Jatropha curcas platymiscium, mainly is distributed in the torrid zone, subtropical zone.Jatropha curcas have loose to become silted up, analgesic effect, often be used to treat traumatic injury, the skin convulsion is itched and enterogastritis.Recent study is found, Jatropha curcas removes and also contains anticancer, disease-resistant, pest-resistant composition, have on new drug development outside the potential important value, its fat content is also very high, and the composition of its grease and petrifaction diesel are approaching, add that the Jatropha curcas drought resisting is anti-lean, not high to environmental requirement, thereby Jatropha curcas can be used as the extensive concern that the energy resources plant causes people, many national government and incorporated business all utilize Jatropha curcas at active development, to produce green energy resources.The production of green energy resources, the Jatropha curcas new varieties new lines that requires to cultivate the high oil of high yield, the breeding that requires to strengthen improved seeds as early as possible enlarges, to plant Jatropha curcas on a large scale.Utilizing plant tissue culture technique, is that to accelerate the fast numerous of improved seeds strain effectively be a kind of good approach.
Just begun the research of the tissue culture technique of Jatropha curcas both at home and abroad as far back as 1996, at present, Jatropha curcas blade, cotyledon, epicotyl, hypocotyl (M.Sujatha can have been utilized, N.Mukta.1996.Morphogenesis and plantregeneration from tissue cultures of Jatropha curcas.Plant Cell Tissue Organ Cult, 44:135-141; Lu Weida, Wei Qin, Tang Lin, the face francium, displaying the derivant of .2003 Jatropha curcas callus breeds fast. use and environmental organism journal 9:127-130) and axillalry bud (Lin Juan, Tang Lin displays tissue culture and the plant regeneration of .2002. Jatropha curcas. Plant Physiology Communications 38 (3): 252) launch Study on tissue culture for explant.But, adopt the regeneration rate of the indefinite bud which kind of explant induces not only all very low and the cultivation time is all longer.As (M.Sujatha such as Sujatha, N.Mukta.1996.Morphogenesis and plant regeneration from tissue cultures ofJatropha curcas.Plant Cell Tissue Organ Cult, 44:135-141) adopting regeneration stem section to induce the bud of growing thickly needs 40 days at least; (A.C.Deore. such as Deore, T.S.Johnson.2008.High-frequency plantregeneration from leaf-disc cultures of Jatropha curcas L:an important biodiesel plant.PlantBiotechnol Rep., 2:7-11) plant regeneration that utilizes blade to do for explant also produces the seedling of growing thickly and took for 20 weeks at least; (T.baran Jha such as Jha, P.Mukherjee, M.M.Datta.2007.Somatic embryogenesis in Jatrophacurcas Linn., an important biofuel plant.Plant Biotechnol Rep. 1:135-140) utilizes blade to take 12-16 week for explant obtains regrowth by somatic embryo generation approach; (Lin Juan, Tang Lin display tissue culture and the plant regeneration of .2002. Jatropha curcas to Lin Juan etc.Plant Physiology Communications 38 (3): 252) finish from 1 breeding of seed germination to the and take 70 days.Just present fast numerous level, though the effect of axillalry bud is best, its fast numerous coefficient high energy in 16 weeks reaches 12.3 (M.Sujathal, H.P.S.Makkar and K.Becker.2005.Shootbud proliferation from axillary nodes and leaf sections of non-toxic.Plant GrowthRegulation, 47:83-90), but the time of cultivating is still longer, is inoculated into into the seedling process need more than 70 days from explant.Moreover, they all also exist cultivates the cost problem of higher, as (Lin Juan such as Lin Juan, Tang Lin, display tissue culture and the plant regeneration of .2002. Jatropha curcas. Plant Physiology Communications 38 (3): 252) in the seed germination medium, added the plant hormone of higher concentration, 6-BA concentration causes the Jatropha curcas great majority of initial culture to produce callus up to 2.5mg/L, and the highest frequency that forms complete seedling has only 57.8%; Chen Jinhong etc. (Chen Jinhong, Gao Min, the living .2006. Jatropha curcas of yellow note stem section cultured in vitro and breeding research fast. Guangxi agricultural science .37 (3): 221-223) the combination medium of employing MS+6BA2.5mg/L+0.1～0.5mg/L IBA; (Leeization such as Li Hua, Ceng Ni, Jia Yongjiong, Tang Lin displays the fast numerous and root induction of short axillalry bud branch of .2006. Jatropha curcas. Sichuan University's journal (natural science edition) 43 (5): 1116-1120) then adopted MS+1.0mg/L 6-BA+0.01mg/L IBA+5mg/LAgNO
3Medium; (M.Sujathal such as Sujatha, H.P.S.Makkar and K.Becker.2005.Shoot bud proliferationfrom axillary nodes and leaf sections of non-toxic.Plant Growth Regulation, 47:83-90) preceding 4 weeks are adopted MS+4.5 μ MTDZ, and the back changes it among MS+8.9 μ M BA+2.5 μ M IBA over to all around just can induce the bud of growing thickly.Obviously, these prescriptions are that the hormone dosage or the complexity of additive are all higher.
Summary of the invention
The objective of the invention is at the cost that exists in the existing Jatropha curcas method for tissue culture higher, waste time and energy, problems such as regeneration rate is not high, and the difficult problem of taking root of the Jatropha curcas tissue cultivating seedling that always exists of this area, the method that a kind of barbadosnut plantlet tissue culture rapid propagation is provided and takes root thinks that applying of Jatropha curcas tissue cultivating seedling lays the foundation.
Barbadosnut plantlet tissue culture rapid propagation provided by the invention and the method for taking root, this method is that the jatropha curcas seed that maturation is shelled is carried out sterilization earlier, taking-up embryo wherein places the basal medium of pH5.4-6.0 then, at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivate down that embryo germination is an aseptic seedling after 5 days; The stem-segment with single bud that aseptic seedling is cut places the fast propagating culture medium of pH5.4-6.0, and at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivate after 7 days down can induce the bud of growing thickly; Downcut in the root media of the sub-pH 5.4-6.0 of inoculation growing to the long bud of growing thickly of 2-3cm, at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivates down and began to take root in 5 days, can transplant the strong sprout that root reaches 2-3cm after 15 days, wherein fast propagating culture medium is in basal medium, adding the 6-benzyladenine (6-BA) of 0.1～0.5mg/L and 3-indolebutyric acid (IBA) or the α-Nai Yisuan (NAA) of 0.15～1.0mg/L forms, root media is in basal medium, adding 3-indolebutyric acid or the α-Nai Yisuan of 0.1～1.0mg/L forms, basal medium then is in the MS medium, adds the curing of 25～35g/L sucrose and agar and forms.
Though the sterilization of the jatropha curcas seed that maturation is shelled can adopt conventional method, the inventor finds to handle brownization of seed behind the 1min with the alcohol of volumetric concentration 70-80% under study for action, and sprout time prolongs, and germination rate reduces, and promptly seed has been subjected to injury; And handle 5-10min seed pollution rate height with mass concentration 0.4-0.5% mercury chloride and liquor natrii hypochloritis, can kill and wound seed and handle above 20min, reduce germination rate, so preferably adopt the preferred method for disinfection and sterilization of groping out through the inventor: handled 0.5-1 minute with the alcohol of volumetric concentration 70-80% earlier, with aseptic water washing 2-3 time, again with the mercuric chloride solution of mass concentration 0.4-0.5% handle 10-17 minute thereafter, more preferably handled 12-15 minute or handled 15-20 minute with the liquor natrii hypochloritis of mass concentration 0.4-0.5%, more preferably handled 16-18 minute, use aseptic water washing 2-7 time at last, preferably wash 2-3 time and get final product.Handle with preferred method for disinfection and sterilization, its survival rate can reach more than 95%, and it is good to sprout quality, no browning.
In the inventive method the cultivating aseptic seedling of Jatropha curcas be with the basal medium MS that do not add hormone for well, not only planting percent is up to 90%, and saves cost.In addition, the cultivation temperature by seed embryo cultivation aseptic seedling is preferably 25-29 ℃.
Be preferably 25-29 ℃ by the grow thickly cultivation temperature of bud of cultivating aseptic seedling in the inventive method, and its fast propagating culture medium adds the 3-indolebutyric acid of 0.3～0.5mg/L 6-benzyladenine and 0.15～0.3mg/L and forms preferably in basal medium.
Cultivate by the bud of growing thickly in the inventive method and take root and grow up to strong sprout used root media preferably in basal medium, add the α-Nai Yisuan of 0.1～0.3mg/L and form.
The present invention compared with prior art has the following advantages and good effect:
1, propagation is used the inventive method soon, forms the seedling (stem section 3-5cm is long) that can be used as explant by seed germination and on average takes 30 days; Growing up to the long seedling of growing thickly of 3-5cm after the explant stem-segment with single bud inoculation as 3-5cm length only needs 21 days, and this seedling of growing thickly can be used for the clone again and again, extremely to the n time propagation; During to the long root of needs, take out the high seedling of growing thickly of 2-3cm and be inoculated into root media, experience the root length of sending in 15 days and reach 2-3cm and can transplant.Thereby, if begin to start at from the time of seed germination, form complete seedling to the 1st propagation seedling rooting and take 66 days; Taking root to the 2nd propagation, to form that complete seedling takes be 87 days; Taking root to the 3rd propagation, to form that complete seedling takes be 108 days; The rest may be inferred later on, so compared with prior art, the present invention is more faster than other fast breeding techniques.
2, variation is as mentioned above little, takes weak point because of the present invention cultivates the propagation seedling, and the seed germination process does not need hormone, and explant material contact hormone in incubation time lack, so less relatively by the caused variation probability of hormone.In addition, owing to hormone concentration used in the incubation is also less, especially do not use as 2, the strong hormone that also more easily causes variation simultaneously of callus of induce power such as 4-D, thereby the present invention causes that the probability of variation is very little, more helps keeping the good strains of seeds of former culture materials.
3, the reproduction coefficient height only needs 3 time-of-weeks with the inventive method, can make a seedling that produces behind the seed germination obtain 4.25 seedlings, and average growth coefficient reaches as high as 4.25, thereby can satisfy the demand of planting Jatropha curcas on a large scale.Concrete computational methods are as follows:
Calculate by the highest reproduction coefficient in 1 year of a jatropha curcas seed: in the micropropagation process, seed germination becomes aseptic seedling to need one month, and the rate of increase is 1, and promptly a seed is not bred, and remains a seedling.This aseptic seedling is cut to one section, inoculates back 21 days and can obtain 4.25 seedlings, and the rate of increase is 4.25.Each first quarter moon can obtain 8 seedlings later on, and promptly the rate of increase is 8.So growth coefficient of the present invention can calculate with following formula:
□＝4.25*8n
N=subculture number in the formula, 1 year 12 months, seed germination one month, the explant of inoculation takes 21 days the present age, calculates by one month; Later every propagation generation takes about 45 days, breeds seedling rooting at last and takes by one month and calculate, and therefore, subculture number is 6 times in 1 year.So,
□＝4.25×8
6＝4.25×262144＝1114112。
This jatropha curcas seed can be bred more than 1,000,000 seedlings by the present invention in 1 year in promptly 1 year.
4, the low the inventive method agents useful for same of cost all is a common agents not only, and used kind is single, and consumption is few, low price, thereby cost is low.
Embodiment
Providing embodiment below further specifies so that the present invention is made; but given embodiment can not be interpreted as limiting the scope of the invention, thereby nonessential improvement and adjustment that the professional and technical personnel has done according to the content and the design philosophy of the invention described above also should belong to protection scope of the present invention.
Embodiment 1
The jatropha curcas seed that maturation is shelled washes 5 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 50s, aseptic water washing 3 times with volumetric concentration 75% alcohol earlier, then with mass concentration 0.5% mercuric chloride solution sterilization 15min, aseptic water washing 3 times.
Take out basal medium MS+30g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH 5.4, then 25 ℃ of cultivation temperature, intensity of illumination 1500lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
The aseptic seedling branch is cut into fast propagating culture medium MS+0.1mg/L 6-BA+0.15mg/L IBA+25g/L sucrose+5g/L agar that stem-segment with single bud places pH 5.4,25 ℃ of cultivation temperature, intensity of illumination 1500lx, light application time 12h/d cultivate after 15 days down can induce the bud of growing thickly.Average growth coefficient 1.0.
Be inoculated in root media MS+0.1mg/L NAA+25g/L sucrose+5g/L agar of pH 5.4 growing to the long bud cutting-out of growing thickly of 2-3cm, 25 ℃ of cultivation temperature, intensity of illumination 1500lx, light application time 12h/d cultivates down and began to take root in 7 days, rooting rate reaches 50% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 2
The jatropha curcas seed that maturation is shelled washes 4 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 30s, aseptic water washing 2 times with volumetric concentration 70% alcohol earlier, then with mass concentration 0.4% mercuric chloride solution sterilization 17min, aseptic water washing 3 times.
Take out basal medium MS+30g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.6, then 27 ℃ of cultivation temperature, intensity of illumination 2000lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
The aseptic seedling branch is cut into fast propagating culture medium MS+0.3mg/L 6-BA+0.15mg/L IBA+30g/L sucrose+5g/L agar that stem-segment with single bud places pH5.6,27 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivate after 7 days down can induce the bud of growing thickly.Average growth coefficient 3.25.
Be inoculated in root media MS+0.1mg/L IBA+30g/L sucrose+5g/L agar of pH5.6 growing to the long bud cutting-out of growing thickly of 2-3cm, 27 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivates down and began to take root in 10 days, rooting rate reaches 33.3% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 3
The jatropha curcas seed that maturation is shelled washes 4 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 1min, aseptic water washing 3 times with volumetric concentration 80% alcohol earlier, then with mass concentration 0.5% mercuric chloride solution sterilization 10min, aseptic water washing 2 times.
Take out basal medium MS+35g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.8, then 29 ℃ of cultivation temperature, intensity of illumination 2500lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.5mg/L 6-BA+0.15mg/L IBA+35g/L sucrose+5g/L agar that stem-segment with single bud places pH5.8,29 ℃ of cultivation temperature, intensity of illumination 2500lx, light application time 12h/d cultivate after 7 days down can induce the bud of growing thickly.Average growth coefficient 4.25.
Be inoculated in root media MS+0.2mg/L NAA+35g/L sucrose+5g/L agar of pH5.8 growing to the long bud cutting-out of growing thickly of 2-3cm, 29 ℃ of cultivation temperature, intensity of illumination 2500lx, light application time 12h/d cultivates down and began to take root in 5 days, rooting rate reaches 70% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 4
The jatropha curcas seed that maturation is shelled washes 4 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 40s, aseptic water washing 3 times with volumetric concentration 80% alcohol earlier, then with mass concentration 0.5% mercuric chloride solution sterilization 12min, aseptic water washing 2 times.
Take out basal medium MS+30g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH6.0, then 35 ℃ of cultivation temperature, intensity of illumination 3000lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.5mg/L 6-BA+1.0mg/L IBA+30g/L sucrose+5g/L agar that stem-segment with single bud places pH6.0,35 ℃ of cultivation temperature, intensity of illumination 3000lx, light application time 12h/d cultivate after 11 days down can induce the bud of growing thickly.Average growth coefficient 2.
Be inoculated in root media MS+0.2mg/L IBA+35g/L sucrose+5g/L agar of pH6.0 growing to the long bud cutting-out of growing thickly of 2-3cm, 35 ℃ of cultivation temperature, intensity of illumination 3000lx, light application time 12h/d cultivates down and began to take root in 8 days, rooting rate reaches 46.67% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 5
The jatropha curcas seed that maturation is shelled washes 4 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 1min, aseptic water washing 3 times with volumetric concentration 70% alcohol earlier, then with the mass concentration 0.4% liquor natrii hypochloritis 15min that sterilizes, aseptic water washing 3 times.
Take out basal medium MS+30g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.5, then 32 ℃ of cultivation temperature, intensity of illumination 2500lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.4mg/L 6-BA+0.3mg/L IBA+30g/L sucrose+5g/L agar that stem-segment with single bud places pH5.5,32 ℃ of cultivation temperature, intensity of illumination 2500lx, light application time 12h/d cultivate after 9 days down can induce the bud of growing thickly.Average growth coefficient 3.5.
Be inoculated in root media MS+0.3mg/L NAA+30g/L sucrose+5g/L agar of pH5.5 growing to the long bud cutting-out of growing thickly of 2-3cm, 32 ℃ of cultivation temperature, intensity of illumination 2500lx, light application time 12h/d cultivates down and began to take root in 5 days, rooting rate reaches 83.33% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 6
The jatropha curcas seed that maturation is shelled washes 3 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 1min, aseptic water washing 3 times with volumetric concentration 75% alcohol earlier, then with the mass concentration 0.5% liquor natrii hypochloritis 20min that sterilizes, aseptic water washing 3 times.
Take out basal medium MS+35g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.8, then 28 ℃ of cultivation temperature, intensity of illumination 2000lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.5mg/L 6-BA+0.15mg/L NAA+35g/L sucrose+5g/L agar that stem-segment with single bud places pH5.8,28 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivate after 7 days down can induce the bud of growing thickly.Average growth coefficient 4.07.
Be inoculated in root media MS+0.1mg/L NAA+35g/L sucrose+5g/L agar of pH5.8 growing to the long bud cutting-out of growing thickly of 2-3cm, 28 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivates down and began to take root in 6 days, rooting rate reaches 50% after 15 days, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 7
The jatropha curcas seed that maturation is shelled washes 3 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 30s, aseptic water washing 2 times with volumetric concentration 80% alcohol earlier, then with the mass concentration 0.45% liquor natrii hypochloritis 16min that sterilizes, aseptic water washing 6 times.
Take out basal medium MS+25g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.8, then 28 ℃ of cultivation temperature, intensity of illumination 2000lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.3mg/L 6-BA+0.3mg/LNAA+25g/L sucrose+5g/L agar that stem-segment with single bud places pH5.8,28 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivate after 12 days down can induce the bud of growing thickly.Average growth coefficient 2.5.
Be inoculated in root media MS+0.1mg/L NAA+25g/L sucrose+5g/L agar of pH5.8 growing to the long bud cutting-out of growing thickly of 2-3cm, 28 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivates down and began to take root in 15 days, base portion produces a large amount of callus, rooting rate reaches 25%, and root reaches can transplant the strong sprout of 2-3cm.
Embodiment 8
The jatropha curcas seed that maturation is shelled washes 3 times repeatedly with clear water, after soaking 3h with clear water again, move into superclean bench and handle 40s, aseptic water washing 3 times with volumetric concentration 75% alcohol earlier, then with the mass concentration 0.5% liquor natrii hypochloritis 18min that sterilizes, aseptic water washing 3 times.
Take out basal medium MS+30g/L sucrose+5g/L agar that embryo in the seed of sterilization back places pH5.6, then 26 ℃ of cultivation temperature, intensity of illumination 2000lx cultivates down by light application time 12h/d that embryo germination is an aseptic seedling after 5 days.
Aseptic seedling is cut into fast propagating culture medium MS+0.1mg/L 6-BA+1.0mg/L NAA+30g/L sucrose+5g/L agar that stem-segment with single bud places pH5.6,26 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivate after 15 days down can induce the bud of growing thickly.Average growth coefficient 1.07.
Be inoculated in root media MS+1.0mg/L IBA+30g/L sucrose+5g/L agar of pH5.6 growing to the long bud cutting-out of growing thickly of 2-3cm, 26 ℃ of cultivation temperature, intensity of illumination 2000lx, light application time 12h/d cultivates down and began to take root in 15 days, base portion produces a large amount of callus, rooting rate reaches 20%, and root reaches can transplant the strong sprout of 2-3cm.
Claims (10)
1. a barbadosnut plantlet tissue culture rapid propagation and the method for taking root, this method is that the jatropha curcas seed that maturation is shelled is carried out sterilization earlier, taking-up embryo wherein places the basal medium of pH5.4-6.0 then, at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivate down that embryo germination is an aseptic seedling after 5 days; The stem-segment with single bud that aseptic seedling is cut places the fast propagating culture medium of pH5.4-6.0, and at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivate after 7 days down and promptly induce the bud of growing thickly; Cut in the root media that is inoculated in pH5.4-6.0 growing to the long bud of growing thickly of 2-3cm, at cultivation temperature 25-35 ℃, intensity of illumination 1500-3000lx, light application time 12h/d cultivate down and began to take root in 5 days, and promptly transplant the strong sprout that root reaches 2-3cm after 15 days; Wherein fast propagating culture medium is to add the 6-benzyladenine of 0.1～0.5mg/L and the 3-indolebutyric acid of 0.15～1.0mg/L in basal medium, or adds the 6-benzyladenine of 0.1～0.5mg/L and the α-Nai Yisuan composition of 0.15～1.0mg/L in basal medium; Root media is in basal medium, adds the 3-indolebutyric acid of 0.1～1.0mg/L or the α-Nai Yisuan of 0.1～1.0mg/L and forms; Basal medium then is in the MS medium, adds the curing of 25～35g/L sucrose and agar and forms.
2. barbadosnut plantlet tissue culture rapid propagation according to claim 1 and the method for taking root, the sterilization of this method is to handle 0.5-1 minute with the alcohol of volumetric concentration 70-80% earlier, with aseptic water washing 2-3 time, again with the mercuric chloride solution of mass concentration 0.4-0.5% handle 10-17 minute or handled 15-20 minute, use aseptic water washing 2-7 time at last with the liquor natrii hypochloritis of mass concentration 0.4-0.5% thereafter.
3. barbadosnut plantlet tissue culture rapid propagation according to claim 1 and the method for taking root, the sterilization of this method is to handle 0.5-1 minute with the alcohol of volumetric concentration 70-80% earlier, with aseptic water washing 2-3 time, again with the mercuric chloride solution of mass concentration 0.4-0.5% handle 12-15 minute or handled 16-18 minute, use aseptic water washing 2-3 time at last with the liquor natrii hypochloritis of mass concentration 0.4-0.5% thereafter.
4. according to claim 1 or 2 or 3 described barbadosnut plantlet tissue culture rapid propagations and the method for taking root, the cultivation temperature by the embryo culture aseptic seedling of seed in this method is 25-29 ℃.
5. according to claim 1 or 2 or 3 described barbadosnut plantlet tissue culture rapid propagations and the method for taking root, the cultivation temperature of cultivating the bud of growing thickly by aseptic seedling in this method is 25-29 ℃, and its fast propagating culture medium is in basal medium, adds the 6-benzyladenine of 0.3～0.5mg/L and the 3-indolebutyric acid of 0.15～0.3mg/L and forms.
6. barbadosnut plantlet tissue culture rapid propagation according to claim 4 and the method for taking root, the cultivation temperature of cultivating the bud of growing thickly by aseptic seedling in this method is 25-29 ℃, and its fast propagating culture medium is in basal medium, adds the 6-benzyladenine of 0.3～0.5mg/L and the 3-indolebutyric acid of 0.15～0.3mg/L and forms.
7. according to claim 1 or 2 or 3 described barbadosnut plantlet tissue culture rapid propagations and the method for taking root, cultivate in this method that to grow thickly the blastogenesis root and to grow up to strong sprout used root media be in basal medium, add the α-Nai Yisuan of 0.1～0.3mg/L and form.
8. barbadosnut plantlet tissue culture rapid propagation according to claim 4 and the method for taking root cultivate in this method that to grow thickly the blastogenesis root and to grow up to strong sprout used root media be in basal medium, add the α-Nai Yisuan of 0.1～0.3mg/L and form.
9. barbadosnut plantlet tissue culture rapid propagation according to claim 5 and the method for taking root cultivate in this method that to grow thickly the blastogenesis root and to grow up to strong sprout used root media be in basal medium, add the α-Nai Yisuan of 0.1～0.3mg/L and form.
10. barbadosnut plantlet tissue culture rapid propagation according to claim 6 and the method for taking root cultivate in this method that to grow thickly the blastogenesis root and to grow up to strong sprout used root media be in basal medium, add the α-Nai Yisuan of 0.1～0.3mg/L and form.
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| CN103283592B (en) * | 2013-04-17 | 2014-11-12 | 华南农业大学 | Method for direct regeneration of adventitious bud from Jatropha curcas petiole explant |
| CN103430841B (en) * | 2013-08-02 | 2014-11-12 | 华南农业大学 | Method for promoting jatropha curcas explant to regenerate adventitious buds |
| CN103416307B (en) * | 2013-08-05 | 2015-05-20 | 四川大学 | Rapid propagation method for directly inducing buds by using jatropha curcas L. seed leaf regenerated roots |
| CN103749295B (en) * | 2013-12-31 | 2015-12-09 | 四川大学 | A kind of method of Jatropha curcas regrowth extraneous root |
| CN106857247B (en) * | 2017-01-16 | 2019-11-08 | 华南农业大学 | A kind of method that Jatropha curcas aseptic seedling respectively organizes callus induction |
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| 林娟，等.麻疯树的组织培养及植株再生.《植物生理学通讯》.2002,第38卷(第3期),252. * |
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