CN108575763A - A kind of China fir stem section inducing culture and abductive approach - Google Patents
A kind of China fir stem section inducing culture and abductive approach Download PDFInfo
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- CN108575763A CN108575763A CN201810805591.8A CN201810805591A CN108575763A CN 108575763 A CN108575763 A CN 108575763A CN 201810805591 A CN201810805591 A CN 201810805591A CN 108575763 A CN108575763 A CN 108575763A
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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
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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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Abstract
The present invention relates to China fir adventitious bud tissue cultures, in particular to a kind of China fir stem section inducing culture and abductive approach.A kind of composition of China fir stem section inducing culture includes：DCR minimal mediums, 6 BA and NAA；A concentration of 0.58~0.62mg/L of 6 BA in the China fir stem section inducing culture, a concentration of 0.2~0.3mg/L of NAA.Explant stem section is subjected to Fiber differentiation in the culture medium of 6 BA and NAA compositions that are matched suitable for minimal medium and suitable concentration, has many advantages, such as that inductivity is high, germination rate is high, adventitious bud number is more, tissue cultures is made to have higher success rate.
Description
Technical field
The present invention relates to China fir adventitious bud field of tissue culture, in particular to a kind of China fir stem section inducing culture
And abductive approach.
Background technology
The full name of tissue culture is Plant Tissue Breeding, broadly also known as cultured in vitro, refers to that institute is isolated from plant
Tissue, organ or the cell etc. needed carries out cultivating final acquisition intact plant under artificial manipulation then by sterile working
One technology.In a narrow sense refer to then being cultivated using the tissue, such as forming layer, mesophyll tissue etc. of plant various pieces
Plant is obtained, and the culture carried out by generating callus in incubation can be referred to, then passes through callus
It is differentiated to form new plant again.The China fir tissue culture technology in China develops very fast, but in the way of development inevitably
Encounter many urgent problems to be solved, this just need we carry out largely test go to solve.
Seeds one of of the China fir as fast growing, while being also our southern Major Tree Species Planteds.As afforestation is advised
The continuous expansion of mould, China fir tissue culture technology already become important nursery means.China fir tissue cultures have had been established one at present
Basic fundamental system, but the clone seeds newly selected are still needed to carry out research and experiment.
Most basic in China fir tissue culture is induction, for tissue culture industrialization, selects suitable material and hormone appropriate
Proportioning becomes the pith of induction.Hormone combinations optimize the problem that China Fir seedling shortage can be effectively relieved.But for
The kind newly selected studies deficiency to it, still needs to study different exogenous hormone processing to China fir tissue culture adventitious bud inducing
It influences, a reliable reference frame is provided for China fir tissue culture and propagation in scale.
In view of this, special propose the present invention.
Invention content
The first object of the present invention is to provide a kind of China fir stem section inducing culture, for growth and material and excellent 64
Number elite tree carries out tissue cultures, and hormone combination is reasonable in the culture medium, has good inducing action to China fir adventitious bud, has
Have the advantages that inductivity is high, germination rate is high, bud number is more.
The second object of the present invention is to provide a kind of China fir stem section abductive approach, and this method setting is reasonable, behaviour
Make simply, pollution rate is low, and induction success rate is high.
In order to realize that the above-mentioned purpose of the present invention, spy use following technical scheme：
A kind of China fir stem section inducing culture, the composition of the culture medium include：DCR minimal mediums, 6-BA and NAA；
A concentration of 0.58~0.62mg/L of 6-BA in the China fir stem section inducing culture, a concentration of 0.2~0.3mg/L of NAA.
A kind of China fir stem section abductive approach, including：Explant sterilizing, stem section inoculation and Fiber differentiation；The culture uses
Above-mentioned China fir stem section inducing culture is cultivated.
Growth of Chinese Fir and material and excellent No. 64 are picked out in forth generation germplasm resource bank by Fujian Yang Kou state-owned forest farms
Elite tree, using the stem section of its current year life base portion coppice shoot as explant, in the 6- matched suitable for minimal medium and suitable concentration
It is cultivated in the culture medium of BA and NAA compositions, has many advantages, such as that inductivity is high, adventitious bud number is more, make tissue cultures success rate
Higher.
A kind of China fir stem section inducing culture, the composition of the culture medium include：DCR minimal mediums, 6-BA and NAA；
A concentration of 0.58~0.62mg/L of 6-BA in the China fir stem section inducing culture, a concentration of 0.2~0.3mg/L of NAA.
Most basic in China fir tissue culture is induction, how to realize that the first step of high yield is to obtain that height can be possessed
The tissue-cultured seedling of inductivity needs to carry out largely to test to find out optimal hormone combination combination.
DCR minimal mediums are that culture solution is commonly used in tissue culture, it can provide required nutriment such as a large amount of members for plant
Element, trace element, molysite and organic substance etc..In addition, common minimal medium also has MS, 1/2MS, 3/4MS etc., it is different
The type and content of nutriment contained by minimal medium are different, then have very for explant rudiment, bud number and budding time etc.
Big influence.Therefore, the selection of minimal medium is also highly important in tissue culture, and it is suitable to be obtained by experimental verification
The minimal medium of plant growth.
6-BA and NAA is auxin.6-BA, that is, 6- benzyl aminoadenines, alias：6-benzyl aminopurine, cell division
Element, English common name：6-Benzylaminopurine is the plant growth regulator to safety of human and livestock.It, which has, promotes cell
Division promotes undifferentiated tissue differentiation, promotes the accumulation of biological substance in vivo, the effects that promoting lateral bud, prevent aging.Just
Because of that 6-BA becomes indispensable compound in plant tissue and cell culture.Another important feature of 6-BA is
Poor mobility in plant, physiological action be confined to treatment site and its near.To consider to handle in practical applications
Method and treatment site.In the application, crop/floristics, usage and concentration, processing time and position difference, are shown
The effect or reaction come is also not quite similar, such as function and effect are poor when low concentration, and excessive concentrations may will produce adverse effect,
So needing to select suitable concentration according to without factors such as floristics, processing time and treatment sites in the application.Such as at this
Apply in different embodiments, concentration can also be 0.60mg/L etc., and result is almost the same within the scope of this.
NAA, that is, 1- methyl α-naphthyl acetates, English common name 1-naphthlcetic acid are chemistry of the people according to natural hormone
Structure, a kind of artificial synthesized broad spectrum type plant growth substance can promote cell division and expand, to promote seed to send out
Root, adventitious bud sprout, cuttage root-taking, and raising, which is bloomed, bears fruit.It is the same with other hormones, have in low concentration and growth is promoted to make
With high concentration has inhibiting effect, and different plant Different Organs are different to the susceptibility of auxin, so when in use because of basis
Floristics and suitable concentration is selected using position.Such as in the different embodiment of the application, concentration can also be
0.22mg/L, 0.25mg/L, 0.28mg/L, 0.28mg/L etc..
Preferably, the composition of the culture medium further includes：Sucrose, carragheen and activated carbon, wherein a concentration of the 28 of sucrose
A concentration of 0.8~1.2g/L of~32g/L, a concentration of 7~9g/L of carragheen, activated carbon.
Sucrose can provide carbon source and the energy as carbohydrate for plant, moreover it is possible to maintain the osmotic pressure of culture solution.Card
Glue i.e. carrageenan, galactolipin and Anhydrogalactose is drawn to be formed by polysaccharide sulfuric acid and 3,6- Anhydrogalactoses are straight
Chain polymerization object is formed, and coagulator use is done.Activated carbon is had an effect by suction-operated, can adsorb having in culture medium
Evil substance, the 5 hydroxymethyl furfural generated in high pressure sterilization such as phenol, quinones and the sucrose secreted in impurity, incubation
Deng.But its simultaneously may also Absorption Growth adjust element, nutrients etc., therefore suitable concentration should be found when in use.Such as at this
Apply in different embodiments, the concentration of sucrose can also be 29g/L, 30g/L, 31g/L (result is almost the same within the scope of this)；
The concentration of carragheen can also be 7.5g/L, 8g/L, 8.5g/L (result is almost the same within the scope of this)；The concentration of activated carbon may be used also
Think 0.9g/L, 1.0g/L, 1.1g/L (result is almost the same within the scope of this).
A kind of China fir stem section abductive approach, including：Explant sterilizing, stem section inoculation and Fiber differentiation；The Fiber differentiation
It is cultivated using above-mentioned China fir stem section inducing culture.
Preferably, the explant is the stem section of China fir base portion coppice shoot；
Preferably, the China fir base portion coppice shoot sampling time is June, July or November.
The induction of China fir adventitious bud is very important the selection of material.The hardness of some China firs itself is higher, it
Spray hardness compare it is more hard with other sprays, then being just not easy to induce.Outside the higher China fir of degree of lignification
Implant is more difficult to generate callus, so, it, should be with the lower base portion of degree of lignification of current year life when choosing explant
Material of the coppice shoot as induction.
The infection rate for the explant taken when June, July or these three month November is minimum, and fine day is selected when taking, can
Substantially reduce infection rate.
Preferably, after the sterilizing, the China fir base portion coppice shoot is cut into a length of 2.0~3.0cm length containing needle
For the stem section of 0.8~1.2cm.
It is inoculated with after sterilizing, inoculation need to carry out in an aseptic environment, and inoculation is first first handled explant.It will go out
Explant after bacterium is pressed from both sides out from container with tweezers, after with knife further processing is carried out to explant：Explant is sheared
The stem section for being 0.8~1.2cm for a length of 2.0~3.0cm length containing needle.
Preferably, the explant, which sterilizes, includes：The alcohol for being 70%~75% with percent by volume impregnates 20s~50s,
Then 8~16min is impregnated with a concentration of 0.08mg/mL~0.12mg/mL mercuric chloride.
Preferably, described be seeded in gnotobasis carries out.
The many because being known as of China fir explant can be polluted, such as：China fir explant acquisition time, cleaning method, disinfection
Whether equipment, sterile environment and the consummation of the gimmick of operator etc..China fir explant sterilizes as China fir tissue culture most critical
One step, it is particularly important.
Alcohol is most common surface disinfectant, best with 70%~75% alcohol bactericidal effect, 95% or absolute alcohol
Phage surface protein matter fast dewatering can be made to solidify, form one layer of desciccator diaphragm, prevent alcohol to continue to penetrate into, bactericidal effect is big
It is big to reduce.Alcohol has stronger penetration power, so that bacterium protein is denaturalized, bactericidal effect is good.But alcohol kills vegetable material
Wound effect is also very big, and the growth of long soaking time, vegetable material will be affected, or even be killed by alcohol, and when use answers
Stringent control time.But alcohol is unable to thorough disinfection, is not used alone generally, is mostly used cooperatively with other disinfectants.Mercuric chloride is
Mercury chloride is white crystal, particle or powder, can inhibit the activity of bacterium enzyme containing sulfydryl, makes bacterial metabolism obstacle, can also be with egg
It binds directly in vain, makes albuminous degeneration.Mercuric chloride common a concentration of 0.1%, i.e. 0.1mg/mL.It such as in various embodiments, can
25s is impregnated for 70% alcohol, 0.1% mercuric chloride impregnates 10min, and 75% alcohol impregnates 40s, and 0.1% mercuric chloride impregnates 14min etc..
The tissue-cultured seedling being inoculated with, once having spent this period, can generally be polluted in the 7th to the 14th infection rate highest
Rate will drastically decline.The reason of pollution, is roughly divided into two kinds of culture medium pollution and material contamination.Most culture medium pollution
All be when inoculation with caused by extraneous contact, seldom partially due to when high-temperature sterilization does not have sterilizing in place caused,
Therefore, it in inoculation, has to ensure the sterile of workbench, to improve inductivity.And the reason of material contamination, is then complex,
It is not thorough enough when may be disinfection, it is also possible to long or to be exposed in air too long very much with blade contact when being inoculated with, it is also possible to
Bottle cap pollution etc..In order to reduce pollution rate, inductivity, the completion for needing each step all extreme cares careful are improved.
Preferably, the inoculation is that the stem section is all linked into the China fir stem section inducing culture.
Inoculation is that sterilized good explant segment or fritter are put into the process of culture medium.
Preferably, the temperature of the culture be 24~26 DEG C, intensity of illumination be 1500~2000lx, light application time be 12~
14h/d。
Culture refers to that culture materials are placed on culturing room's (having illumination, temperature condition) is inner, is allowed to grow, the mistake of Differentiation
Journey.The suitable cultivation temperature of different plants is different with intensity of illumination and time.As in various embodiments, the condition of culture is also
Can be temperature it be 25 DEG C, intensity of illumination 1800lx, light application time 13h/d；Temperature is 26 DEG C, and intensity of illumination is
2000lx, light application time 12h/d；Temperature is 24 DEG C, intensity of illumination 1600lx, and light application time is 14h/d etc..
Preferably, the China fir is No. 64 trees in the forth generation germplasm resource bank of Fujian Yang Kou state-owned forest farms.
No. 64 trees are growth and material and excellent elite tree.
Compared with prior art, beneficial effects of the present invention are：
The molding of (1) species depends primarily on genotypic and environment, in the present solution, growth and material and excellent China fir
The wooden elite tree represents excellent genotype, and the China fir stem section inducing culture of the proportioning containing hormon is then environment, this two
Person is combined with each other, the preferred plan that finally can be derived that best environmental condition to be induced as China fir explant.
(2) the application has obtained the concentration of suitable 6-BA and NAA by many experiments and data analysis, in this range
It is interior, the induction of adventitious bud can be effectively facilitated, and excessively there will be inhibiting effect.
(3) the application is with the growth picked out in the forth generation germplasm resource bank of Fujian Yang Kou state-owned forest farms and material and excellent
No. 64 elite trees of China fir be genetic stocks, by many experiments and data analysis obtained various concentration exogenous hormone processing pair
The influence of China fir tissue culture Stem tip induction provides a reliable reference frame for China fir tissue culture, has social benefit and scientific research valence
Value.
Specific implementation mode
Embodiment of the present invention is described in detail below in conjunction with embodiment, but those skilled in the art will
Understand, the following example is merely to illustrate the present invention, and is not construed as limiting the scope of the invention.It is not specified in embodiment specific
Condition person carries out according to conventional conditions or manufacturer's recommended conditions.Reagents or instruments used without specified manufacturer is
It can be with conventional products that are commercially available.
In the application specific implementation mode, the calculation of inductivity, germination rate and average bud number and range analysis and
Meaning is as follows：
K2=at DCR or 0.8mg/l6-BA or 0.2mg/lNAA everywhere in reason the sum of result
Induce number：Successfully induce the bottle number of adventitious bud；
Inoculation number：The bottle number being inoculated with；
Rudiment bottle number：There is the bottle number of rudiment；
Rudiment sum：Untainted the sum of the rudiment number that can successfully induce adventitious bud or induce adventitious bud；
It is bigger, it is bigger on test index influence, reflect the influence of each small factors on test indicators in three main factors
Size.
R：It is bigger, it is bigger on test index influence, it is more important, it can reflect the primary-slave relation of three main factors.
MS used in this application is shown in Table 1 and table respectively with DCR minimal medium mother liquor formulas and auxin mother liquor formula
2。
Table 1MS and DCR mother liquor formula tables
Embodiment 1
In the morning of a certain fair weather in June, chooses in the forth generation germplasm resource bank of Fujian Yang Kou state-owned forest farms and select
The growth gone out is explant with the current year of material and excellent No. 64 elite trees of China fir life base portion coppice shoot, removes extra blade, flowing water
Sterile water wash is used after rinsing well 2 times, 25s is impregnated with 70% alcohol, then cleaned 2 times with asepsis, then with 0.1% liter
Mercury impregnates 10min, with sterile water wash 5 times.The explant that sterilizing is completed is placed on spare in vial.
China fir stem section inducing culture is prepared, and required article is put into workbench and is sterilized 30 minutes with ultraviolet lamp, behaviour
Work person's handle is sterilized with alcohol swab, and tweezers and knife are impregnated with alcohol, after the calcination on alcolhol burner, by explant tweezers from glass
Pressed from both sides out in glass bottle, after with knife the stem section blade of explant is switched to about 1cm, bottle cap is twisted, and (bottle cap needs the first calcination under alcolhol burner
For a moment).Tweezers and scalpel are required for being sterilized with alcolhol burner after being inoculated with every time.It is carried out on culture medium after being all inoculated with
It marks, the selected blaze of China fir of each bottle, inoculation date and inoculation people in note, in order to record data.The culture that will be inoculated with
Base is put into culturing room and is cultivated, and culture room temperature is maintained at 25 DEG C or so, intensity of illumination 1800lx, light application time 13h/d.
It is recorded every three days to pollution number, rudiment number, the bud number of five days docking seedlings.Every group of experiment is 3 times parallel.Logarithm
According to progress range analysis.
Embodiment 2
In the morning of a certain fair weather in July, chooses in the forth generation germplasm resource bank of Fujian Yang Kou state-owned forest farms and select
The growth gone out is explant with the current year of material and excellent No. 64 elite trees of China fir life base portion coppice shoot, removes extra blade, flowing water
Sterile water wash is used after rinsing well 2 times, 40s is impregnated with 75% alcohol, then cleaned 2 times with asepsis, then with 0.11%
Mercuric chloride impregnates 14min, with sterile water wash 5 times.The explant that sterilizing is completed is placed on spare in vial.
China fir stem section inducing culture is prepared, and required article is put into workbench and is sterilized 30 minutes with ultraviolet lamp, behaviour
Work person's handle is sterilized with alcohol swab, and tweezers and knife are impregnated with alcohol, after the calcination on alcolhol burner, by explant tweezers from glass
Pressed from both sides out in glass bottle, after with knife the stem section blade of explant is switched to about 1cm, bottle cap is twisted, and (bottle cap needs the first calcination under alcolhol burner
For a moment).Tweezers and scalpel are required for being sterilized with alcolhol burner after being inoculated with every time.It is carried out on culture medium after being all inoculated with
It marks, the selected blaze of China fir of each bottle, inoculation date and inoculation people in note, in order to record data.The culture that will be inoculated with
Base is put into culturing room and is cultivated, and culture room temperature is maintained at 26 DEG C or so, intensity of illumination 2000lx, light application time 12h/d.
It is recorded every three days to pollution number, rudiment number, the bud number of five days docking seedlings.Every group of experiment is 3 times parallel.Logarithm
According to progress range analysis.
Embodiment 3
In the morning of a certain fair weather in November, chooses in the forth generation germplasm resource bank of Fujian Yang Kou state-owned forest farms and select
The growth gone out is explant with the current year of material and excellent No. 64 elite trees of China fir life base portion coppice shoot, removes extra blade, flowing water
Sterile water wash is used after rinsing well 2 times, 40s is impregnated with 75% alcohol, then cleaned 2 times with asepsis, then with 0.1% liter
Mercury impregnates 14min, with sterile water wash 5 times.The explant that sterilizing is completed is placed on spare in vial.
China fir stem section inducing culture is prepared, and required article is put into workbench and is sterilized 30 minutes with ultraviolet lamp, behaviour
Work person's handle is sterilized with alcohol swab, and tweezers and knife are impregnated with alcohol, after the calcination on alcolhol burner, by explant tweezers from glass
Pressed from both sides out in glass bottle, after with knife the stem section blade of explant is switched to about 1cm, bottle cap is twisted, and (bottle cap needs the first calcination under alcolhol burner
For a moment).Tweezers and scalpel are required for being sterilized with alcolhol burner after being inoculated with every time.It is carried out on culture medium after being all inoculated with
It marks, the selected blaze of China fir of each bottle, inoculation date and inoculation people in note, in order to record data.
The culture medium being inoculated with is put into culturing room to cultivate, culture room temperature is maintained at 24 DEG C or so, intensity of illumination
1600lx, light application time 14h/d.
It is recorded every three days to pollution number, rudiment number, the bud number of five days docking seedlings.Every group of experiment is 3 times parallel.Logarithm
According to progress range analysis.
Experimental example 1
The influence of the pairs of inductivity of different China fir stem section inducing culture groups.
Based on embodiment 3, it is related to the change of three experiment parameters：Minimal medium, 6-BA, NAA, China fir stem section lure
It is constant to lead other compositions such as sucrose, carragheen and activated carbon additive amount in culture medium, and experimental method is the same as embodiment 3.
Wherein culture solution is divided into 1/2MS, MS and DCR, and the concentration of 6-BA is divided into 0.6mg/L, 0.8mg/L, 1.2mg/L, NAA
Concentration be divided into 0.1mg/L, 0.2mg/L, 0.3mg/L, specific proportioning is as shown in the table：
3 not isogeneous induction China fir stem section inducing culture L9 (3 of table3) formula orthogonal test table
Each culture medium is a processing, each 30 bottles of processing inoculation, one explant of every bottle of inoculation, and experiment repeats 3
It is secondary.
It is for statistical analysis to experimental data, it the results are shown in Table 4.
The impact analysis table of the pairs of inductivity of the different China fir stem section inducing culture groups of table 4
According to table 4 it is found that No. 64 China fir elite tree stems of various concentration pair of variety classes minimal medium and 6-BA, NAA
Section inductivity has a great impact.No. 64 China fir elite tree stem sections are in processing 5：DCR+0.8mg/L 6-BA+0.3mg/L NAA and
Processing 4：Inductivity highest under DCR+0.6mg/L 6-BA+0.2mg/LNAA, reaches 90.9%；ByValue is it is found that basic training
The optimal level for supporting base is the second horizontal DCR, and the optimal level of 6-BA is the second horizontal 0.8mg/L, and the optimal level of NAA is the
Two horizontal 0.2mg/L, therefore, theoretically the best China fir stem section inducing culture of No. 64 China fir elite tree inductivities is DCR+
0.8mg/L 6-BA+0.2mg/L NAA.But from same factorValue can be seen that the first level of 6-BA concentration
(72.5%) and second horizontal (73.7%) difference is not notable and significantly higher than third is horizontal (62.7%)；The of NAA concentration
Two horizontal (74.8%) and horizontal (74.0) difference of third is not notable, but is significantly higher than first level (60.2%).According to R values,
By ordered arrangement from big to small：Minimal medium>NAA>6-BA.That is minimal medium is that No. 64 China firs of influence are selected
Set the leading factor of stem section inductivity.Consider, the best China fir stem section inducing culture conducive to the induction of No. 64 elite trees is：
DCR+0.6~0.8mg/L6-BA+0.2~0.3mg/L NAA.
Experimental example 2
Influence of the different China fir stem section inducing cultures to germination rate.
Experimental setup and method are the same as experimental example 1.
It is for statistical analysis to experimental data, it the results are shown in Table 5.
The impact analysis table of the pairs of germination rate of the different China fir stem section inducing culture groups of table 5
According to table 5 it is found that No. 64 China fir elite tree base portion coppice shoot stem sections are handling the germination rate under 1,2,4,5,7,8 most
Height reaches 100%；ByIt is found that the optimal level of minimal medium is first level 1/2MS, the optimal level of 6-BA is value
The optimal level of second level 0.6 or 0.8mg/L, NAA are the horizontal 0.3mg/L of third, but make a concrete analysis of each factor
Difference is not notable (96.7%, 96.3%, 96.3%) between value can be seen that three levels of minimal medium, and the three of NAA concentration
Also not notable (96.3%, 96.3%, 96.7%) is differed between a level, and the concentration of 6-BA does not have in 0.6mg/L and 0.8mg/L
Variant (being 100.0%).According to R values, by ordered arrangement from big to small：6-BA>NAA>Minimal medium, therefore, 6-
BA is the leading factor for influencing No. 64 China fir elite tree stem section germination rates.Therefore, in conjunction withValue and R values obtain being conducive to No. 64 essences
Selecting the best China fir stem section inducing culture of tree base portion coppice shoot is：1/2MS or DCR or 3/4MS+0.6~0.8mg/L6-BA+0.1
~0.3mg/L NAA.
Experimental example 3
Influence of the different China fir stem section inducing cultures to bud number.
Experimental setup and method are the same as experimental example 1.
It is for statistical analysis to experimental data, it the results are shown in Table 6.
Impact analysis table of the different China fir stem section inducing cultures of table 6 to bud number
According to table 6 it is found that No. 64 China fir elite tree base portion coppice shoot stem sections are in processing 4：DCR+0.6mg/L 6-BA+0.2mg/
Average bud number under L NAA is most, reaches 6.4/bottle；ByValue is it is found that the optimal level of minimal medium is the second level
The optimal level of DCR, 6-BA are first level 0.6mg/L, and the optimal level of NAA is the second horizontal 0.2mg/L, therefore, theoretical
The be averaged best China fir stem section inducing culture of bud number of upper No. 64 elite trees is DCR+0.6mg/L6-BA+0.2mg/L NAA.Root
According to R values, by ordered arrangement from big to small：Minimal medium>6-BA>NAA, therefore, minimal medium are to influence No. 64 China firs
Elite tree stem section is averaged the leading factor of bud number.To NAA concentrationValue is further analyzed, optimal level (3.94) and
Third level (3.47) difference is not notable, and the R values in conjunction with NAA concentration are minimum, it is thus determined that being conducive to No. 64 elite tree base portion coppice shoots
The best China fir stem section inducing culture of bud number is：DCR+0.6mg/L6-BA+0.2~0.3mg/L NAA.
Inductivity highest in No. 64 China fir elite tree base portion coppice shoot stem section inductions has been respectively obtained by the above experimental example, has been sprouted
The most culture medium composition of bud rate highest and averagely bud number.Conducive to the best training of No. 64 China fir elite tree base portion coppice shoot stem sections induction
Foster base is：DCR+0.6~0.8mg/L 6-BA+0.2~0.3mg/LNAA；It is sprouted conducive to No. 64 China fir elite tree base portion coppice shoot stem sections
The optimal medium of bud is：1/2MS or DCR or 3/4MS+0.6~0.8mg/L 6-BA+0.1~0.3mg/L NAA；Conducive to 64
The optimal medium of number China fir elite tree base portion coppice shoot stem section bud number is：DCR+0.6mg/L 6-BA+0.2~0.3mg/LNAA.
It can be selected according to demand.Considering inductivity, germination rate and bud number, best medium is：DCR+0.6mg/L 6-BA+
0.2~0.3mg/L NAA.
Finally it should be noted that：The above embodiments are only used to illustrate the technical solution of the present invention., rather than its limitations；To the greatest extent
Present invention has been described in detail with reference to the aforementioned embodiments for pipe, but it will be understood by those of ordinary skill in the art that：Its
It still can be with technical scheme described in the above embodiments is modified, either to which part or all technical features
Carry out equivalent replacement；And these modifications or replacements, various embodiments of the present invention skill that it does not separate the essence of the corresponding technical solution
The range of art scheme.
Claims (10)
1. a kind of China fir stem section inducing culture, which is characterized in that the composition of the culture medium includes：DCR minimal mediums, 6-
BA and NAA；
A concentration of 0.58~0.62mg/L of 6-BA in the China fir stem section inducing culture, a concentration of 0.2~0.3mg/ of NAA
L。
2. China fir stem section inducing culture according to claim 1, which is characterized in that the China fir stem section inducing culture
Composition further include：Sucrose, carragheen and activated carbon, wherein a concentration of 28~32g/L of sucrose, carragheen a concentration of 7~
A concentration of 0.8~1.2g/L of 9g/L, activated carbon.
3. a kind of China fir stem section abductive approach, which is characterized in that including：Explant sterilizing, stem section inoculation and Fiber differentiation；
The Fiber differentiation is cultivated using the China fir stem section inducing culture described in claim 2.
4. China fir stem section abductive approach according to claim 3, which is characterized in that the explant is China fir base portion coppice shoot
Stem section；
Preferably, the China fir base portion coppice shoot sampling time is June, July or November.
5. China fir stem section abductive approach according to claim 4, which is characterized in that after the sterilizing, by the China fir
Base portion coppice shoot is cut into the stem section that a length of 2.0~3.0cm length containing needle is 0.8~1.2cm.
6. China fir stem section abductive approach according to claim 3, which is characterized in that the explant, which sterilizes, includes：Use body
The alcohol that product percentage is 70%~75% impregnates 20s~50s, is then soaked with a concentration of 0.08mg/mL~0.12mg/mL mercuric chloride
Steep 8~16min.
7. China fir stem section abductive approach according to claim 3, which is characterized in that it is described be seeded in gnotobasis into
Row.
8. China fir stem section abductive approach according to claim 3, which is characterized in that the inoculation is that the stem section is whole
It is linked into the China fir stem section inducing culture.
9. China fir stem section abductive approach according to claim 3, which is characterized in that the temperature of the culture is 24~26
DEG C, intensity of illumination is 1500~2000lx, and light application time is 12~14h/d.
10. according to claim 3~9 any one of them China fir stem section abductive approach, which is characterized in that the China fir is Fujian
No. 64 trees in the forth generation germplasm resource bank of foreign mouth state-owned forest farms.
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| CN109479713A (en) * | 2018-12-03 | 2019-03-19 | 南京林业大学 | A kind of coppice spout using China fir carries out root induction method as explant |
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| CN108633742A (en) * | 2018-07-18 | 2018-10-12 | 南京林业大学 | A kind of China fir Stem tip induction culture medium and abductive approach |
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| CN109479713A (en) * | 2018-12-03 | 2019-03-19 | 南京林业大学 | A kind of coppice spout using China fir carries out root induction method as explant |
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